WorldWideScience

Sample records for human ct domain

  1. The human ACC2 CT-domain C-terminus is required for full functionality and has a novel twist

    Energy Technology Data Exchange (ETDEWEB)

    Madauss, Kevin P. [Department of Computational and Structural Chemistry, GlaxoSmithKline Inc., Five Moore Drive, Research Triangle Park, NC 27709 (United States); Burkhart, William A.; Consler, Thomas G. [Department of Biochemical Reagents and Assay Development, GlaxoSmithKline Inc., Five Moore Drive, Research Triangle Park, NC 27709 (United States); Cowan, David J. [Department of Chemistry in the Center for Excellence in Metabolic Pathways Drug Discovery, GlaxoSmithKline Inc., Five Moore Drive, Research Triangle Park, NC 27709 (United States); Gottschalk, William K. [Institute for Genome Sciences and Policy and Department of Medicine, Division of Neurology, Duke University, Durham, NC 27708 (United States); Miller, Aaron B. [Department of Computational and Structural Chemistry, GlaxoSmithKline Inc., Five Moore Drive, Research Triangle Park, NC 27709 (United States); Short, Steven A. [Department of Biochemical Reagents and Assay Development, GlaxoSmithKline Inc., Five Moore Drive, Research Triangle Park, NC 27709 (United States); Tran, Thuy B. [Department of Physiology, UNC School of Medicine, University of North Carolina, Chapel Hill, NC 27515 (United States); Williams, Shawn P., E-mail: shawn.p.williams@gsk.com [Department of Computational and Structural Chemistry, GlaxoSmithKline Inc., Five Moore Drive, Research Triangle Park, NC 27709 (United States)

    2009-05-01

    The use of biophysical assays permitted the identification of a specific human ACC2 carboxyl transferase (CT) domain mutant that binds inhibitors and crystallizes in their presence. This mutant led to determination of the human ACC2 CT domain–CP-640186 complex crystal structure, which revealed differences in the inhibitor conformation from the yeast protein complex that are caused by differing residues in the binding pocket. Inhibition of acetyl-CoA carboxylase (ACC) may prevent lipid-induced insulin resistance and type 2 diabetes, making the enzyme an attractive pharmaceutical target. Although the enzyme is highly conserved amongst animals, only the yeast enzyme structure is available for rational drug design. The use of biophysical assays has permitted the identification of a specific C-terminal truncation of the 826-residue human ACC2 carboxyl transferase (CT) domain that is both functionally competent to bind inhibitors and crystallizes in their presence. This C-terminal truncation led to the determination of the human ACC2 CT domain–CP-640186 complex crystal structure, which revealed distinctions from the yeast-enzyme complex. The human ACC2 CT-domain C-terminus is comprised of three intertwined α-helices that extend outwards from the enzyme on the opposite side to the ligand-binding site. Differences in the observed inhibitor conformation between the yeast and human structures are caused by differing residues in the binding pocket.

  2. Differentiation of early from advanced coronary atherosclerotic lesions: systematic comparison of CT, intravascular US, and optical frequency domain imaging with histopathologic examination in ex vivo human hearts.

    Science.gov (United States)

    Maurovich-Horvat, Pál; Schlett, Christopher L; Alkadhi, Hatem; Nakano, Masataka; Stolzmann, Paul; Vorpahl, Marc; Scheffel, Hans; Tanaka, Atsushi; Warger, William C; Maehara, Akiko; Ma, Shixin; Kriegel, Matthias F; Kaple, Ryan K; Seifarth, Harald; Bamberg, Fabian; Mintz, Gary S; Tearney, Guillermo J; Virmani, Renu; Hoffmann, Udo

    2012-11-01

    To establish an ex vivo experimental setup for imaging coronary atherosclerosis with coronary computed tomographic (CT) angiography, intravascular ultrasonography (US), and optical frequency domain imaging (OFDI) and to investigate their ability to help differentiate early from advanced coronary plaques. All procedures were performed in accordance with local and federal regulations and the Declaration of Helsinki. Approval of the local Ethics Committee was obtained. Overall, 379 histologic cuts from nine coronary arteries from three donor hearts were acquired, coregistered among modalities, and assessed for the presence and composition of atherosclerotic plaque. To assess the discriminatory capacity of the different modalities in the detection of advanced lesions, c statistic analysis was used. Interobserver agreement was assessed with the Cohen κ statistic. Cross sections without plaque at coronary CT angiography and with fibrous plaque at OFDI almost never showed advanced lesions at histopathologic examination (odds ratio [OR]: 0.02 and 0.06, respectively; both Padvanced lesions (OR: 2.49, P=.0003; OR: 2.60, P=.002; and OR: 31.2, Padvanced lesions than intravascular US and coronary CT angiography (area under the receiver operating characteristic curve: 0.858 [95% confidence interval {CI}: 0.802, 0.913], 0.631 [95% CI: 0.554, 0.709], and 0.679 [95% CI: 0.618, 0.740]; respectively, Padvanced coronary plaques. © RSNA, 2012

  3. Changing domains in human capital measurement

    Directory of Open Access Journals (Sweden)

    Pharny D. Chrysler-Fox

    2014-09-01

    Research purpose: The aim of the study was to explore and describe changing domains within human capital management to be managed and measured. Motivation for the study: The motivation was to advance the understanding of changing measurement domains to aid practitioners to manage and measure the contribution of the human resource function and employees, in order to unlock and add value and ultimately contribute to the success of an organisation. Research design, approach and method: Unstructured, in-depth interview data of purposively selected cases from a selected panel of human resource practitioners specialising in human capital measurement was thematically analysed in this exploratory-descriptive investigation. Main findings: Findings suggested that seven domains should be managed and measured. These domains highlight new areas of impact and levels of management. In addition, crossdomain relationships in measurement allow for an understanding of the impact and potential value on which to capitalise. Practical/managerial implications: New domains to manage and measure focus the attention of practitioners beyond the transactional performance management paradigm to a transformational approach to influence the business strategy. Higher education institutions need to develop students’ cognitive skills to facilitate systems thinking. Contribution: This study suggests a new approach to managing and measuring the human capital function and the workforce.

  4. Assembly of human C-terminal binding protein (CtBP) into tetramers.

    Science.gov (United States)

    Bellesis, Andrew G; Jecrois, Anne M; Hayes, Janelle A; Schiffer, Celia A; Royer, William E

    2018-06-08

    C-terminal binding protein 1 (CtBP1) and CtBP2 are transcriptional coregulators that repress numerous cellular processes, such as apoptosis, by binding transcription factors and recruiting chromatin-remodeling enzymes to gene promoters. The NAD(H)-linked oligomerization of human CtBP is coupled to its co-transcriptional activity, which is implicated in cancer progression. However, the biologically relevant level of CtBP assembly has not been firmly established; nor has the stereochemical arrangement of the subunits above that of a dimer. Here, multi-angle light scattering (MALS) data established the NAD + - and NADH-dependent assembly of CtBP1 and CtBP2 into tetramers. An examination of subunit interactions within CtBP1 and CtBP2 crystal lattices revealed that both share a very similar tetrameric arrangement resulting from assembly of two dimeric pairs, with specific interactions probably being sensitive to NAD(H) binding. Creating a series of mutants of both CtBP1 and CtBP2, we tested the hypothesis that the crystallographically observed interdimer pairing stabilizes the solution tetramer. MALS data confirmed that these mutants disrupt both CtBP1 and CtBP2 tetramers, with the dimer generally remaining intact, providing the first stereochemical models for tetrameric assemblies of CtBP1 and CtBP2. The crystal structure of a subtle destabilizing mutant suggested that small structural perturbations of the hinge region linking the substrate- and NAD-binding domains are sufficient to weaken the CtBP1 tetramer. These results strongly suggest that the tetramer is important in CtBP function, and the series of CtBP mutants reported here can be used to investigate the physiological role of the tetramer. © 2018 Bellesis et al.

  5. Statistical CT noise reduction with multiscale decomposition and penalized weighted least squares in the projection domain

    International Nuclear Information System (INIS)

    Tang Shaojie; Tang Xiangyang

    2012-01-01

    Purposes: The suppression of noise in x-ray computed tomography (CT) imaging is of clinical relevance for diagnostic image quality and the potential for radiation dose saving. Toward this purpose, statistical noise reduction methods in either the image or projection domain have been proposed, which employ a multiscale decomposition to enhance the performance of noise suppression while maintaining image sharpness. Recognizing the advantages of noise suppression in the projection domain, the authors propose a projection domain multiscale penalized weighted least squares (PWLS) method, in which the angular sampling rate is explicitly taken into consideration to account for the possible variation of interview sampling rate in advanced clinical or preclinical applications. Methods: The projection domain multiscale PWLS method is derived by converting an isotropic diffusion partial differential equation in the image domain into the projection domain, wherein a multiscale decomposition is carried out. With adoption of the Markov random field or soft thresholding objective function, the projection domain multiscale PWLS method deals with noise at each scale. To compensate for the degradation in image sharpness caused by the projection domain multiscale PWLS method, an edge enhancement is carried out following the noise reduction. The performance of the proposed method is experimentally evaluated and verified using the projection data simulated by computer and acquired by a CT scanner. Results: The preliminary results show that the proposed projection domain multiscale PWLS method outperforms the projection domain single-scale PWLS method and the image domain multiscale anisotropic diffusion method in noise reduction. In addition, the proposed method can preserve image sharpness very well while the occurrence of “salt-and-pepper” noise and mosaic artifacts can be avoided. Conclusions: Since the interview sampling rate is taken into account in the projection domain

  6. Analysis of brain CT on 120 patients of human cysticercosis

    International Nuclear Information System (INIS)

    Ma, J.; To, R.; Ri, T.; Ra, S.; Inomata, Taiten; Ogawa, Yasuhiro; Maeda, Tomoo.

    1990-01-01

    A study on brain CT was made in 120 patients of human cysticercosis, which is a rare disease in Japan and clinical symptoms and laboratory data for the diagnosis were also discussed. From the point of therapeutic view, we proposed a new differentiation on brain CT of human cysticercosis, which is divided into two groups according to the alve or dead parasite. Furthermore, we proposed a new type named multiple large and small cysts type on brain CT. The idea of diagnostic standard was made integrating brain CT image, clinical symptoms and labolatory data. (author)

  7. Human-computer interface incorporating personal and application domains

    Science.gov (United States)

    Anderson, Thomas G [Albuquerque, NM

    2011-03-29

    The present invention provides a human-computer interface. The interface includes provision of an application domain, for example corresponding to a three-dimensional application. The user is allowed to navigate and interact with the application domain. The interface also includes a personal domain, offering the user controls and interaction distinct from the application domain. The separation into two domains allows the most suitable interface methods in each: for example, three-dimensional navigation in the application domain, and two- or three-dimensional controls in the personal domain. Transitions between the application domain and the personal domain are under control of the user, and the transition method is substantially independent of the navigation in the application domain. For example, the user can fly through a three-dimensional application domain, and always move to the personal domain by moving a cursor near one extreme of the display.

  8. Construction and Design Kits: Human Problem-Domain Communication

    National Research Council Canada - National Science Library

    Fischer, Gerhard; Lemke, Andreas C

    1987-01-01

    .... To provide the user with the appropriate level of control and a better understanding, we have to replace human-computer communication with human problem-domain communication, which allows users...

  9. Structural and Histone Binding Ability Characterizations of Human PWWP Domains

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Hong; Zeng, Hong; Lam, Robert; Tempel, Wolfram; Amaya, Maria F.; Xu, Chao; Dombrovski, Ludmila; Qiu, Wei; Wang, Yanming; Min, Jinrong (Toronto); (Penn)

    2013-09-25

    The PWWP domain was first identified as a structural motif of 100-130 amino acids in the WHSC1 protein and predicted to be a protein-protein interaction domain. It belongs to the Tudor domain 'Royal Family', which consists of Tudor, chromodomain, MBT and PWWP domains. While Tudor, chromodomain and MBT domains have long been known to bind methylated histones, PWWP was shown to exhibit histone binding ability only until recently. The PWWP domain has been shown to be a DNA binding domain, but sequence analysis and previous structural studies show that the PWWP domain exhibits significant similarity to other 'Royal Family' members, implying that the PWWP domain has the potential to bind histones. In order to further explore the function of the PWWP domain, we used the protein family approach to determine the crystal structures of the PWWP domains from seven different human proteins. Our fluorescence polarization binding studies show that PWWP domains have weak histone binding ability, which is also confirmed by our NMR titration experiments. Furthermore, we determined the crystal structures of the BRPF1 PWWP domain in complex with H3K36me3, and HDGF2 PWWP domain in complex with H3K79me3 and H4K20me3. PWWP proteins constitute a new family of methyl lysine histone binders. The PWWP domain consists of three motifs: a canonical {beta}-barrel core, an insertion motif between the second and third {beta}-strands and a C-terminal {alpha}-helix bundle. Both the canonical {beta}-barrel core and the insertion motif are directly involved in histone binding. The PWWP domain has been previously shown to be a DNA binding domain. Therefore, the PWWP domain exhibits dual functions: binding both DNA and methyllysine histones.

  10. Image Fusion of CT and MR with Sparse Representation in NSST Domain

    Directory of Open Access Journals (Sweden)

    Chenhui Qiu

    2017-01-01

    Full Text Available Multimodal image fusion techniques can integrate the information from different medical images to get an informative image that is more suitable for joint diagnosis, preoperative planning, intraoperative guidance, and interventional treatment. Fusing images of CT and different MR modalities are studied in this paper. Firstly, the CT and MR images are both transformed to nonsubsampled shearlet transform (NSST domain. So the low-frequency components and high-frequency components are obtained. Then the high-frequency components are merged using the absolute-maximum rule, while the low-frequency components are merged by a sparse representation- (SR- based approach. And the dynamic group sparsity recovery (DGSR algorithm is proposed to improve the performance of the SR-based approach. Finally, the fused image is obtained by performing the inverse NSST on the merged components. The proposed fusion method is tested on a number of clinical CT and MR images and compared with several popular image fusion methods. The experimental results demonstrate that the proposed fusion method can provide better fusion results in terms of subjective quality and objective evaluation.

  11. Epitope mapping of the domains of human angiotensin converting enzyme.

    Science.gov (United States)

    Kugaevskaya, Elena V; Kolesanova, Ekaterina F; Kozin, Sergey A; Veselovsky, Alexander V; Dedinsky, Ilya R; Elisseeva, Yulia E

    2006-06-01

    Somatic angiotensin converting enzyme (sACE), contains in its single chain two homologous domains (called N- and C-domains), each bearing a functional zinc-dependent active site. The present study aims to define the differences between two sACE domains and to localize experimentally revealed antigenic determinants (B-epitopes) in the recently determined three-dimensional structure of testicular tACE. The predicted linear antigenic determinants of human sACE were determined by peptide scanning ("PEPSCAN") approach. Essential difference was demonstrated between locations of the epitopes in the N- and C-domains. Comparison of arrangement of epitopes in the human domains with the corresponding sequences of some mammalian sACEs enabled to classify the revealed antigenic determinants as variable or conserved areas. The location of antigenic determinants with respect to various structural elements and to functionally important sites of the human sACE C-domain was estimated. The majority of antigenic sites of the C-domain were located at the irregular elements and at the boundaries of secondary structure elements. The data show structural differences between the sACE domains. The experimentally revealed antigenic determinants were in agreement with the recently determined crystal tACE structure. New potential applications are open to successfully produce mono-specific and group-specific antipeptide antibodies.

  12. Improving Human Resources Recruitment in Maritime Domain

    OpenAIRE

    Surugiu Felicia; Dragomir Cristina

    2011-01-01

    In order to improve maritime human resources recruitment, most crewing and shipping companies implement an internal procedure of recruitment and management of ship’s crew. Based on observations at several crewing companies, this paper presents an example of such procedure meant to ensure that qualified and competent personnel is recruited and all ships in the fleet have crew members understanding their roles, duties and responsibilities and capable of working effectively in teams

  13. Machine vs. human translation of SNOMED CT terms.

    Science.gov (United States)

    Schulz, Stefan; Bernhardt-Melischnig, Johannes; Kreuzthaler, Markus; Daumke, Philipp; Boeker, Martin

    2013-01-01

    In the context of past and current SNOMED CT translation projects we compare three kinds of SNOMED CT translations from English to German by: (t1) professional medical translators; (t2) a free Web-based machine translation service; (t3) medical students. 500 SNOMED CT fully specified names from the (English) International release were randomly selected. Based on this, German translations t1, t2, and t3 were generated. A German and an Austrian physician rated the translations for linguistic correctness and content fidelity. Kappa for inter-rater reliability was 0.4 for linguistic correctness and 0.23 for content fidelity. Average ratings of linguistic correctness did not differ significantly between human translation scenarios. Content fidelity was rated slightly better for student translators compared to professional translators. Comparing machine to human translation, the linguistic correctness differed about 0.5 scale units in favour of the human translation and about 0.25 regarding content fidelity, equally in favour of the human translation. The results demonstrate that low-cost translation solutions of medical terms may produce surprisingly good results. Although we would not recommend low-cost translation for producing standardized preferred terms, this approach can be useful for creating additional language-specific entry terms. This may serve several important use cases. We also recommend testing this method to bootstrap a crowdsourcing process, by which term translations are gathered, improved, maintained, and rated by the user community.

  14. Personality psychology : Domains of knowledge about human nature

    NARCIS (Netherlands)

    Larsen, Randy J.; Buss, David M.; Wismeijer, Andreas; Song, John; van den Berg, Stéphanie Martine

    Using a unique organizational framework that emphasizes six domains of knowledge about human nature, Personality Psychology presents an accessible, contemporary look at personality as a collection of interrelated topics and themes. The book focuses on the scientific basis of our knowledge about

  15. Institutionalizing the Human Domain: Achieving Cross Domain Synergy for Every Day Missions

    Science.gov (United States)

    2017-04-06

    others rely on more primitive and traditional sources of information or are characterized by strong central government censorship . Technology can play...many societies, others rely on more primitive and traditional sources of information or are characterized by strong central government censorship and...Capabilities for Operations in the Human Domain 92 31 Notes 1 The National Military Strategy of the United States of America , 2015

  16. Structure of the first PDZ domain of human PSD-93

    DEFF Research Database (Denmark)

    Fiorentini, Monica; Nielsen, Ann Kallehauge; Kristensen, Ole

    2009-01-01

    The crystal structure of the PDZ1 domain of human PSD-93 has been determined to 2.0 A resolution. The PDZ1 domain forms a crystallographic trimer that is also predicted to be stable in solution. The main contributions to the stabilization of the trimer seem to arise from interactions involving...... the PDZ1-PDZ2 linker region at the extreme C-terminus of PDZ1, implying that the oligomerization that is observed is not of biological significance in full-length PSD-93. Comparison of the structures of the binding cleft of PSD-93 PDZ1 with the previously reported structures of PSD-93 PDZ2 and PDZ3...

  17. Limited angle CT reconstruction by simultaneous spatial and Radon domain regularization based on TV and data-driven tight frame

    Science.gov (United States)

    Zhang, Wenkun; Zhang, Hanming; Wang, Linyuan; Cai, Ailong; Li, Lei; Yan, Bin

    2018-02-01

    Limited angle computed tomography (CT) reconstruction is widely performed in medical diagnosis and industrial testing because of the size of objects, engine/armor inspection requirements, and limited scan flexibility. Limited angle reconstruction necessitates usage of optimization-based methods that utilize additional sparse priors. However, most of conventional methods solely exploit sparsity priors of spatial domains. When CT projection suffers from serious data deficiency or various noises, obtaining reconstruction images that meet the requirement of quality becomes difficult and challenging. To solve this problem, this paper developed an adaptive reconstruction method for limited angle CT problem. The proposed method simultaneously uses spatial and Radon domain regularization model based on total variation (TV) and data-driven tight frame. Data-driven tight frame being derived from wavelet transformation aims at exploiting sparsity priors of sinogram in Radon domain. Unlike existing works that utilize pre-constructed sparse transformation, the framelets of the data-driven regularization model can be adaptively learned from the latest projection data in the process of iterative reconstruction to provide optimal sparse approximations for given sinogram. At the same time, an effective alternating direction method is designed to solve the simultaneous spatial and Radon domain regularization model. The experiments for both simulation and real data demonstrate that the proposed algorithm shows better performance in artifacts depression and details preservation than the algorithms solely using regularization model of spatial domain. Quantitative evaluations for the results also indicate that the proposed algorithm applying learning strategy performs better than the dual domains algorithms without learning regularization model

  18. Xe enhanced CT in the human newborn infant

    International Nuclear Information System (INIS)

    Fuse, Yozen; Nemoto, Yuko; Shimizu, Mitsumasa; Uga, Naoki; Tada, Hiroshi; Fujii, Toshi; Machida, Keiichi; Kikuchi, Hideo; Izumi, Shigemitsu.

    1990-01-01

    With a cranial computed tomography (CT) using stable xenon gas as a diffusible tracer, we measured regional cerebral blood flow (rCBF) in 6 newborn infants with a variety of neurological abnormalities. Gestational ages and birthweights were 35 to 43 weeks and 2436 to 3540 g, respectively. Four infants exhibited hypoxic-ischemic encephalopathy (HIE), one infant had subdural hemorrhage and the other one was the infant with hyponatremia. A baseline CT was done during denitrogenation by 100% oxygen breathing and then a mixture of 35% xenon and 65% oxygen was breathed for 6 minutes. Six scans were obtained during the inhalation period then the infant was returned to breathing 100% oxygen and additional 7 scans were taken. Four samples of arterial blood were collected every 2 minutes before and during inhalation of the xenon gas. A rCBF was calculated with the changes of Hounsfield units in brain tissue and arterial blood. Relatively high blood flows in the region of the basal nuclei as well as decreased flows in the occipital white matter were observed in the infants with HIE. In an infant with subdural hemorrhage, the blood flows were markedly reduced in the areas adjacent to the lesion, including the basal nuclei, and frontal white matter in the opposite hemisphere. Xenon-enhanced CT by inhaling low concentration of the xenon gas enables to measure rCBF in the human newborn infants without no obvious side effect. (author)

  19. Understanding the Canadian adult CT head rule trial: use of the theoretical domains framework for process evaluation

    Directory of Open Access Journals (Sweden)

    Curran Janet A

    2013-02-01

    Full Text Available Abstract Background The Canadian CT Head Rule was prospectively derived and validated to assist clinicians with diagnostic decision-making regarding the use of computed tomography (CT in adult patients with minor head injury. A recent intervention trial failed to demonstrate a decrease in the rate of head CTs following implementation of the rule in Canadian emergency departments. Yet, the same intervention, which included a one-hour educational session and reminders at the point of requisition, was successful in reducing cervical spine imaging rates in the same emergency departments. The reason for the varied effect of the intervention across these two behaviours is unclear. There is an increasing appreciation for the use of theory to conduct process evaluations to better understand how strategies are linked with outcomes in implementation trials. The Theoretical Domains Framework (TDF has been used to explore health professional behaviour and to design behaviour change interventions but, to date, has not been used to guide a theory-based process evaluation. In this proof of concept study, we explored whether the TDF could be used to guide a retrospective process evaluation to better understand emergency physicians’ responses to the interventions employed in the Canadian CT Head Rule trial. Methods A semi-structured interview guide, based on the 12 domains from the TDF, was used to conduct telephone interviews with project leads and physician participants from the intervention sites in the Canadian CT Head Rule trial. Two reviewers independently coded the anonymised interview transcripts using the TDF as a coding framework. Relevant domains were identified by: the presence of conflicting beliefs within a domain; the frequency of beliefs; and the likely strength of the impact of a belief on the behaviour. Results Eight physicians from four of the intervention sites in the Canadian CT Head Rule trial participated in the interviews. Barriers

  20. Scanning technology with multi-slice helical CT in security inspection domain

    International Nuclear Information System (INIS)

    Wang Jue; Wang Fuquan; Jiang Zenghui

    2008-01-01

    The paper analyzes the technology conditions of security inspection in home and abroad, and expatiates technology of spiral CT and how to define CT value etc, with studying on the key technology of spiral CT scanning way (X-RAY, detector, technology of pulley etc) and mutual relation. By comparing the present products of security inspection, the conclusion was drawn that it is inevitable to develop the tendency of security inspection area with the checking and discerning the substance by using the technology of multi-layer spiral CT. (authors)

  1. Application of social domain of human mind in water management

    Science.gov (United States)

    Piirimäe, Kristjan

    2010-05-01

    Currently, researches dispute whether a human reasons domain-generally or domain-specifically (Fiddick, 2004). The theory of several intuitive reasoning programmes in human mind suggests that the main driver to increase problem-solving abilities is social domain (Byrne & Bates, 2009). This theory leads to an idea to apply the social domain also in environmental management. More specifically, environmental problems might be presented through social aspects. Cosmides (1989) proposed that the most powerful programme in our social domain might be ‘cheater detection module' - a genetically determined mental tool whose dedicated function is to unmask cheaters. She even suggested that only cheater detection can enable logical reasoning. Recently, this idea has found experimental proof and specifications (Buchner et al., 2009). From this perspective, a participatory environmental decision support system requires involvement of the representatives of social control such as environmental agencies and NGOs. These evaluators might effectively discover legal and moral inconsistencies, logical errors and other weaknesses in proposals if they are encouraged to detect cheating. Thus, instead of just environmental concerns, the query of an artificial intelligence should emphasize cheating. Following the idea of Cosmides (1989), employment of cheater detectors to EDSS might appear the only way to achieve environmental management which applies correct logical reasoning as well as both, legislative requirements and conservationist moral. According to our hypothesis, representatives of social control can well discover legal and moral inconsistencies, logical errors and and other weaknesses in envirionmental management proposals if encouraged for cheater detection. In our social experiment, a draft plan of measures for sustainable management of Lake Peipsi environment was proposed to representatives of social control, including Ministry of Environment, other environmental authorities

  2. Single-Domain Antibodies As Therapeutics against Human Viral Diseases

    Directory of Open Access Journals (Sweden)

    Yanling Wu

    2017-12-01

    Full Text Available In full-size formats, monoclonal antibodies have been highly successful as therapeutics against cancer and immune diseases. However, their large size leads to inaccessibility of some epitopes and relatively high production costs. As an alternative, single-domain antibodies (sdAbs offer special advantages compared to full-size antibodies, including smaller size, larger number of accessible epitopes, relatively low production costs and improved robustness. Currently, sdAbs are being developed against a number of viruses, including human immunodeficiency virus-1 (HIV-1, influenza viruses, hepatitis C virus (HCV, respiratory syncytial virus (RSV, and enteric viruses. Although sdAbs are very potent inhibitors of viral infections, no sdAbs have been approved for clinical use against virial infection or any other diseases. In this review, we discuss the current state of research on sdAbs against viruses and their potential as therapeutics against human viral diseases.

  3. Comparison among Wavelet filters and others in the frequency domain for reducing Poisson noise in head CT

    International Nuclear Information System (INIS)

    Perez Diaz, M.; Ruiz Gonzalez, Y.; Lorenzo Ginori, J. V.

    2015-01-01

    This paper describes a comparison among some wavelet filters and other most traditional filters in the frequency domain like Median, Wiener and Butter worth to reduce Poisson noise in Computed Tomography (CT) scans. Five slices of CT containing the posterior fossa from an anthropomorphic phantom and from patients were selected. As their original projections contain noise from the acquisition process, some simulated noise-free lesions were added on the images. After that, the whole images were artificially contaminated with Poisson noise over the sinogram-space. The configurations using wavelets drawn from four wavelet families, using various decomposition levels, and different thresholds, were tested in order to determine de-noising performance as well as the rest of the traditional filters. The quality of the resulting images was evaluated by using Contrast to Noise Ratio (CNR), HVS absolute norm (H1), and Structural Similarity Index (SSIM) as quantitative metrics. We have observed that Wavelet filtering is an alternative to be considered for Poisson noise reduction in image processing of posterior fossa images for head CT with similar behavior to Butter worth and better than Median or Wiener filters for the developed experiment. (Author)

  4. CSI Frequency Domain Fingerprint-Based Passive Indoor Human Detection

    Directory of Open Access Journals (Sweden)

    Chong Han

    2018-04-01

    Full Text Available Passive indoor personnel detection technology is now a hot topic. Existing methods have been greatly influenced by environmental changes, and there are problems with the accuracy and robustness of detection. Passive personnel detection based on Wi-Fi not only solves the above problems, but also has the advantages of being low cost and easy to implement, and can be better applied to elderly care and safety monitoring. In this paper, we propose a passive indoor personnel detection method based on Wi-Fi, which we call FDF-PIHD (Frequency Domain Fingerprint-based Passive Indoor Human Detection. Through this method, fine-grained physical layer Channel State Information (CSI can be extracted to generate feature fingerprints so as to help determine the state in the scene by matching online fingerprints with offline fingerprints. In order to improve accuracy, we combine the detection results of three receiving antennas to obtain the final test result. The experimental results show that the detection rates of our proposed scheme all reach above 90%, no matter whether the scene is human-free, stationary or a moving human presence. In addition, it can not only detect whether there is a target indoors, but also determine the current state of the target.

  5. Deep learning methods for CT image-domain metal artifact reduction

    Science.gov (United States)

    Gjesteby, Lars; Yang, Qingsong; Xi, Yan; Shan, Hongming; Claus, Bernhard; Jin, Yannan; De Man, Bruno; Wang, Ge

    2017-09-01

    Artifacts resulting from metal objects have been a persistent problem in CT images over the last four decades. A common approach to overcome their effects is to replace corrupt projection data with values synthesized from an interpolation scheme or by reprojection of a prior image. State-of-the-art correction methods, such as the interpolation- and normalization-based algorithm NMAR, often do not produce clinically satisfactory results. Residual image artifacts remain in challenging cases and even new artifacts can be introduced by the interpolation scheme. Metal artifacts continue to be a major impediment, particularly in radiation and proton therapy planning as well as orthopedic imaging. A new solution to the long-standing metal artifact reduction (MAR) problem is deep learning, which has been successfully applied to medical image processing and analysis tasks. In this study, we combine a convolutional neural network (CNN) with the state-of-the-art NMAR algorithm to reduce metal streaks in critical image regions. Training data was synthesized from CT simulation scans of a phantom derived from real patient images. The CNN is able to map metal-corrupted images to artifact-free monoenergetic images to achieve additional correction on top of NMAR for improved image quality. Our results indicate that deep learning is a novel tool to address CT reconstruction challenges, and may enable more accurate tumor volume estimation for radiation therapy planning.

  6. Effective data-domain noise and streak reduction for X-ray CT

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Zhi; Zamyatin, Alexander A. [Toshiba Medical Research Institute USA, Inc., Vernon Hills, IL (United States); Akino, Naruomi [Toshiba Medical System Corporation, Tokyo (Japan)

    2011-07-01

    Streaks and noise caused by photon starvation can seriously impair the diagnostic value of the CT imaging. Existing processing methods often have several parameters to tune. The parameters can be ad hoc to the data sets. Iterative methods can achieve better results, however, at the cost of more hardware resources or longer processing time. This paper reports a new scheme of adaptive Gaussian filtering, which is based on the diffusion-derived scale-space concept. In scale-space view, filtering by Gaussians of different sizes is similar to decompose the data into a sequence of scales. The scale measure, which is the variance of the filter, should be linearly related to the noise standard deviation instead of the variance of the noise. This is a fundamental deviation in the way of using filters. The new filter has only one parameter that remains stable once tuned. Singlepass processing can usually reach the desired results. (orig.)

  7. Expanding the landscape of chromatin modification (CM-related functional domains and genes in human.

    Directory of Open Access Journals (Sweden)

    Shuye Pu

    2010-11-01

    Full Text Available Chromatin modification (CM plays a key role in regulating transcription, DNA replication, repair and recombination. However, our knowledge of these processes in humans remains very limited. Here we use computational approaches to study proteins and functional domains involved in CM in humans. We analyze the abundance and the pair-wise domain-domain co-occurrences of 25 well-documented CM domains in 5 model organisms: yeast, worm, fly, mouse and human. Results show that domains involved in histone methylation, DNA methylation, and histone variants are remarkably expanded in metazoan, reflecting the increased demand for cell type-specific gene regulation. We find that CM domains tend to co-occur with a limited number of partner domains and are hence not promiscuous. This property is exploited to identify 47 potentially novel CM domains, including 24 DNA-binding domains, whose role in CM has received little attention so far. Lastly, we use a consensus Machine Learning approach to predict 379 novel CM genes (coding for 329 proteins in humans based on domain compositions. Several of these predictions are supported by very recent experimental studies and others are slated for experimental verification. Identification of novel CM genes and domains in humans will aid our understanding of fundamental epigenetic processes that are important for stem cell differentiation and cancer biology. Information on all the candidate CM domains and genes reported here is publicly available.

  8. Iterative image-domain ring artifact removal in cone-beam CT

    Science.gov (United States)

    Liang, Xiaokun; Zhang, Zhicheng; Niu, Tianye; Yu, Shaode; Wu, Shibin; Li, Zhicheng; Zhang, Huailing; Xie, Yaoqin

    2017-07-01

    Ring artifacts in cone beam computed tomography (CBCT) images are caused by pixel gain variations using flat-panel detectors, and may lead to structured non-uniformities and deterioration of image quality. The purpose of this study is to propose a method of general ring artifact removal in CBCT images. This method is based on the polar coordinate system, where the ring artifacts manifest as stripe artifacts. Using relative total variation, the CBCT images are first smoothed to generate template images with fewer image details and ring artifacts. By subtracting the template images from the CBCT images, residual images with image details and ring artifacts are generated. As the ring artifact manifests as a stripe artifact in a polar coordinate system, the artifact image can be extracted by mean value from the residual image; the image details are generated by subtracting the artifact image from the residual image. Finally, the image details are compensated to the template image to generate the corrected images. The proposed framework is iterated until the differences in the extracted ring artifacts are minimized. We use a 3D Shepp-Logan phantom, Catphan©504 phantom, uniform acrylic cylinder, and images from a head patient to evaluate the proposed method. In the experiments using simulated data, the spatial uniformity is increased by 1.68 times and the structural similarity index is increased from 87.12% to 95.50% using the proposed method. In the experiment using clinical data, our method shows high efficiency in ring artifact removal while preserving the image structure and detail. The iterative approach we propose for ring artifact removal in cone-beam CT is practical and attractive for CBCT guided radiation therapy.

  9. Human-Robot Interaction in High Vulnerability Domains

    Science.gov (United States)

    Gore, Brian F.

    2016-01-01

    Future NASA missions will require successful integration of the human with highly complex systems. Highly complex systems are likely to involve humans, automation, and some level of robotic assistance. The complex environments will require successful integration of the human with automation, with robots, and with human-automation-robot teams to accomplish mission critical goals. Many challenges exist for the human performing in these types of operational environments with these kinds of systems. Systems must be designed to optimally integrate various levels of inputs and outputs based on the roles and responsibilities of the human, the automation, and the robots; from direct manual control, shared human-robotic control, or no active human control (i.e. human supervisory control). It is assumed that the human will remain involved at some level. Technologies that vary based on contextual demands and on operator characteristics (workload, situation awareness) will be needed when the human integrates into these systems. Predictive models that estimate the impact of the technologies on the system performance and the on the human operator are also needed to meet the challenges associated with such future complex human-automation-robot systems in extreme environments.

  10. Structure of a two-CAP-domain protein from the human hookworm parasite Necator americanus

    Energy Technology Data Exchange (ETDEWEB)

    Asojo, Oluwatoyin A., E-mail: oasojo@unmc.edu [Pathology and Microbiology Department, 986495 Nebraska Medical Center, Omaha, NE 68198-6495 (United States)

    2011-05-01

    The first structure of a two-CAP-domain protein, Na-ASP-1, from the major human hookworm parasite N. americanus refined to a resolution limit of 2.2 Å is presented. Major proteins secreted by the infective larval stage hookworms upon host entry include Ancylostoma secreted proteins (ASPs), which are characterized by one or two CAP (cysteine-rich secretory protein/antigen 5/pathogenesis related-1) domains. The CAP domain has been reported in diverse phylogenetically unrelated proteins, but has no confirmed function. The first structure of a two-CAP-domain protein, Na-ASP-1, from the major human hookworm parasite Necator americanus was refined to a resolution limit of 2.2 Å. The structure was solved by molecular replacement (MR) using Na-ASP-2, a one-CAP-domain ASP, as the search model. The correct MR solution could only be obtained by truncating the polyalanine model of Na-ASP-2 and removing several loops. The structure reveals two CAP domains linked by an extended loop. Overall, the carboxyl-terminal CAP domain is more similar to Na-ASP-2 than to the amino-terminal CAP domain. A large central cavity extends from the amino-terminal CAP domain to the carboxyl-terminal CAP domain, encompassing the putative CAP-binding cavity. The putative CAP-binding cavity is a characteristic cavity in the carboxyl-terminal CAP domain that contains a His and Glu pair. These residues are conserved in all single-CAP-domain proteins, but are absent in the amino-terminal CAP domain. The conserved His residues are oriented such that they appear to be capable of directly coordinating a zinc ion as observed for CAP proteins from reptile venoms. This first structure of a two-CAP-domain ASP can serve as a template for homology modeling of other two-CAP-domain proteins.

  11. Structure of a two-CAP-domain protein from the human hookworm parasite Necator americanus

    International Nuclear Information System (INIS)

    Asojo, Oluwatoyin A.

    2011-01-01

    The first structure of a two-CAP-domain protein, Na-ASP-1, from the major human hookworm parasite N. americanus refined to a resolution limit of 2.2 Å is presented. Major proteins secreted by the infective larval stage hookworms upon host entry include Ancylostoma secreted proteins (ASPs), which are characterized by one or two CAP (cysteine-rich secretory protein/antigen 5/pathogenesis related-1) domains. The CAP domain has been reported in diverse phylogenetically unrelated proteins, but has no confirmed function. The first structure of a two-CAP-domain protein, Na-ASP-1, from the major human hookworm parasite Necator americanus was refined to a resolution limit of 2.2 Å. The structure was solved by molecular replacement (MR) using Na-ASP-2, a one-CAP-domain ASP, as the search model. The correct MR solution could only be obtained by truncating the polyalanine model of Na-ASP-2 and removing several loops. The structure reveals two CAP domains linked by an extended loop. Overall, the carboxyl-terminal CAP domain is more similar to Na-ASP-2 than to the amino-terminal CAP domain. A large central cavity extends from the amino-terminal CAP domain to the carboxyl-terminal CAP domain, encompassing the putative CAP-binding cavity. The putative CAP-binding cavity is a characteristic cavity in the carboxyl-terminal CAP domain that contains a His and Glu pair. These residues are conserved in all single-CAP-domain proteins, but are absent in the amino-terminal CAP domain. The conserved His residues are oriented such that they appear to be capable of directly coordinating a zinc ion as observed for CAP proteins from reptile venoms. This first structure of a two-CAP-domain ASP can serve as a template for homology modeling of other two-CAP-domain proteins

  12. Expression, purification, crystallization and preliminary X-ray analysis of the human NORE1 SARAH domain

    International Nuclear Information System (INIS)

    Kim, Hye Jin; Hwang, Eunha; Han, Young-Hyun; Choi, Saehae; Lee, Woo Cheol; Kim, Hye-Yeon; Jeon, Young Ho; Cheong, Chaejoon; Cheong, Hae-Kap

    2012-01-01

    The crystallization of the human NORE1 SARAH domain is reported. NORE1 is an important tumour suppressor in human cancers that interacts with the pro-apoptotic protein kinase MST1/2 through SARAH domains. The SARAH domain (residues 366–413) of human NORE1 was expressed in Escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method. The crystal diffracted to 2.7 Å resolution and belonged to space group P6 1 22, with unit-cell parameters a = b = 73.041, c = 66.092 Å, α = β = 90, γ = 120°

  13. NovelFam3000 – Uncharacterized human protein domains conserved across model organisms

    Science.gov (United States)

    Kemmer, Danielle; Podowski, Raf M; Arenillas, David; Lim, Jonathan; Hodges, Emily; Roth, Peggy; Sonnhammer, Erik LL; Höög, Christer; Wasserman, Wyeth W

    2006-01-01

    Background Despite significant efforts from the research community, an extensive portion of the proteins encoded by human genes lack an assigned cellular function. Most metazoan proteins are composed of structural and/or functional domains, of which many appear in multiple proteins. Once a domain is characterized in one protein, the presence of a similar sequence in an uncharacterized protein serves as a basis for inference of function. Thus knowledge of a domain's function, or the protein within which it arises, can facilitate the analysis of an entire set of proteins. Description From the Pfam domain database, we extracted uncharacterized protein domains represented in proteins from humans, worms, and flies. A data centre was created to facilitate the analysis of the uncharacterized domain-containing proteins. The centre both provides researchers with links to dispersed internet resources containing gene-specific experimental data and enables them to post relevant experimental results or comments. For each human gene in the system, a characterization score is posted, allowing users to track the progress of characterization over time or to identify for study uncharacterized domains in well-characterized genes. As a test of the system, a subset of 39 domains was selected for analysis and the experimental results posted to the NovelFam3000 system. For 25 human protein members of these 39 domain families, detailed sub-cellular localizations were determined. Specific observations are presented based on the analysis of the integrated information provided through the online NovelFam3000 system. Conclusion Consistent experimental results between multiple members of a domain family allow for inferences of the domain's functional role. We unite bioinformatics resources and experimental data in order to accelerate the functional characterization of scarcely annotated domain families. PMID:16533400

  14. NovelFam3000 – Uncharacterized human protein domains conserved across model organisms

    Directory of Open Access Journals (Sweden)

    Sonnhammer Erik LL

    2006-03-01

    Full Text Available Abstract Background Despite significant efforts from the research community, an extensive portion of the proteins encoded by human genes lack an assigned cellular function. Most metazoan proteins are composed of structural and/or functional domains, of which many appear in multiple proteins. Once a domain is characterized in one protein, the presence of a similar sequence in an uncharacterized protein serves as a basis for inference of function. Thus knowledge of a domain's function, or the protein within which it arises, can facilitate the analysis of an entire set of proteins. Description From the Pfam domain database, we extracted uncharacterized protein domains represented in proteins from humans, worms, and flies. A data centre was created to facilitate the analysis of the uncharacterized domain-containing proteins. The centre both provides researchers with links to dispersed internet resources containing gene-specific experimental data and enables them to post relevant experimental results or comments. For each human gene in the system, a characterization score is posted, allowing users to track the progress of characterization over time or to identify for study uncharacterized domains in well-characterized genes. As a test of the system, a subset of 39 domains was selected for analysis and the experimental results posted to the NovelFam3000 system. For 25 human protein members of these 39 domain families, detailed sub-cellular localizations were determined. Specific observations are presented based on the analysis of the integrated information provided through the online NovelFam3000 system. Conclusion Consistent experimental results between multiple members of a domain family allow for inferences of the domain's functional role. We unite bioinformatics resources and experimental data in order to accelerate the functional characterization of scarcely annotated domain families.

  15. Correlation between human observer performance and model observer performance in differential phase contrast CT

    International Nuclear Information System (INIS)

    Li, Ke; Garrett, John; Chen, Guang-Hong

    2013-01-01

    Purpose: With the recently expanding interest and developments in x-ray differential phase contrast CT (DPC-CT), the evaluation of its task-specific detection performance and comparison with the corresponding absorption CT under a given radiation dose constraint become increasingly important. Mathematical model observers are often used to quantify the performance of imaging systems, but their correlations with actual human observers need to be confirmed for each new imaging method. This work is an investigation of the effects of stochastic DPC-CT noise on the correlation of detection performance between model and human observers with signal-known-exactly (SKE) detection tasks.Methods: The detectabilities of different objects (five disks with different diameters and two breast lesion masses) embedded in an experimental DPC-CT noise background were assessed using both model and human observers. The detectability of the disk and lesion signals was then measured using five types of model observers including the prewhitening ideal observer, the nonprewhitening (NPW) observer, the nonprewhitening observer with eye filter and internal noise (NPWEi), the prewhitening observer with eye filter and internal noise (PWEi), and the channelized Hotelling observer (CHO). The same objects were also evaluated by four human observers using the two-alternative forced choice method. The results from the model observer experiment were quantitatively compared to the human observer results to assess the correlation between the two techniques.Results: The contrast-to-detail (CD) curve generated by the human observers for the disk-detection experiments shows that the required contrast to detect a disk is inversely proportional to the square root of the disk size. Based on the CD curves, the ideal and NPW observers tend to systematically overestimate the performance of the human observers. The NPWEi and PWEi observers did not predict human performance well either, as the slopes of their CD

  16. Structure of the thioredoxin-fold domain of human phosducin-like protein 2

    International Nuclear Information System (INIS)

    Lou, Xiaochu; Bao, Rui; Zhou, Cong-Zhao; Chen, Yuxing

    2009-01-01

    The X-ray crystal structure of the Trx-fold domain of hPDCL2 was solved at 2.70 Å resolution and resembled the Trx-fold domain of rat phosducin. Human phosducin-like protein 2 (hPDCL2) has been identified as belonging to subgroup II of the phosducin (Pdc) family. The members of this family share an N-terminal helix domain and a C-terminal thioredoxin-fold (Trx-fold) domain. The X-ray crystal structure of the Trx-fold domain of hPDCL2 was solved at 2.70 Å resolution and resembled the Trx-fold domain of rat phosducin. Comparative structural analysis revealed the structural basis of their putative functional divergence

  17. A new pro-migratory activity on human myogenic precursor cells for a synthetic peptide within the E domain of the mechano growth factor

    International Nuclear Information System (INIS)

    Mills, Philippe; Lafreniere, Jean-Francois; Benabdallah, Basma Fattouma; El Fahime, El Mostafa; Tremblay, Jacques-P.

    2007-01-01

    Duchenne muscular dystrophy (DMD) is an inherited disease that leads to progressive muscle wasting. Myogenic precursor cell transplantation is an approach that can introduce the normal dystrophin gene in the muscle fibers of the patients. Unfortunately, these myogenic precursor cells do not migrate well in the muscle and thus many injections have to be done to enable a good graft success. Recent reports have shown that there is extensive splicing of the IGF-1 gene in muscles. The MGF isoform contains a C-terminal 24 amino acids peptide in the E domain (MGF-Ct24E) that has intrinsic properties. It can promote the proliferation while delaying the differentiation of C 2 C 12 cells. Here, we demonstrated that this synthetic peptide is a motogenic factor for human precursor myogenic cells in vitro and in vivo. Indeed, MGF-Ct24E peptide can modulate members of the fibrinolytic and metalloproteinase systems, which are implicated in the migration of myogenic cells. MGF-Ct24E peptide enhances the expression of u-PA, u-PAR and MMP-7 while reducing PAI-1 activity. Moreover, it has no effect on the gelatinases MMP-2 and -9. Those combined effects can favour cell migration. Finally, we present some results suggesting that the MGF-Ct24E peptide induces these cell responses through a mechanism that does not involve the IGF-1 receptor. Thus, this MGF-Ct24E peptide has a new pro-migratory activity on human myogenic precursor cells that may be helpful in the treatment of DMD. Those results reinforce the possibility that the IGF-1Ec isoform may produce an E domain peptide that can act as a cytokine

  18. CT imaging of the internal human ear: Test of a high resolution scanner

    Energy Technology Data Exchange (ETDEWEB)

    Bettuzzi, M., E-mail: matteo.bettuzzi@unibo.it [Department of Physics, University of Bologna and National Institute of Nuclear Physics Section of Bologna (Italy); Brancaccio, R.; Morigi, M.P. [Department of Physics, University of Bologna and National Institute of Nuclear Physics Section of Bologna (Italy); Gallo, A. [Medicine Faculty, Magna Graecia University, Catanzaro and INFN Cosenza (Italy); Strolin, S.; Casali, F. [Department of Physics, University of Bologna and National Institute of Nuclear Physics Section of Bologna (Italy); Lamanna, Ernesto [Medicine Faculty, Magna Graecia University, Catanzaro and INFN Cosenza (Italy); Ariu, Marilu [CEFLA Dental Group, Imola (Italy)

    2011-08-21

    During the course of 2009, in the framework of a project supported by the National Institute of Nuclear Physics, a number of tests were carried out at the Department of Physics of the University of Bologna in order to achieve a good quality CT scan of the internal human ear. The work was carried out in collaboration with the local 'S. Orsola' Hospital in Bologna and a company (CEFLA) already involved in the production and commercialization of a CT scanner dedicated to dentistry. A laboratory scanner with a simple concept detector (CCD camera-lens-mirror-scintillator) was used to see to what extent it was possible to enhance the quality of a conventional CT scanner when examining the internal human ear. To test the system, some conventional measurements were made, such as the spatial resolution calculation with the MTF and dynamic range evaluation. Different scintillators were compared to select the most suitable for the purpose. With 0.5 mm thick structured cesium iodide and a field of view of 120x120 mm{sup 2}, a spatial resolution of 6.5l p/mm at 5% MTF was obtained. The CT of a pair of human head phantoms was performed at an energy of 120 kVp. The first phantom was a rough representation of the human head shape, with soft tissue made of coarse slabs of Lucite. Some inserts, like small aluminum cylinders and cubes, with 1 mm diameter drilled holes, were used to simulate the channels that one finds inside the human inner ear. The second phantom is a plastic PVC fused head with a real human cranium inside. The bones in the cranium are well conserved and the inner ear features, such as the cochlea and semicircular channels, are clearly detectable. After a number of CT tests we obtained good results as far as structural representation and channel detection are concerned. Some images of the 3D rendering of the CT volume are shown below. The doctors of the local hospital who followed our experimentation expressed their satisfaction. The CT was compared to a

  19. Time domain functional NIRS imaging for human brain mapping.

    Science.gov (United States)

    Torricelli, Alessandro; Contini, Davide; Pifferi, Antonio; Caffini, Matteo; Re, Rebecca; Zucchelli, Lucia; Spinelli, Lorenzo

    2014-01-15

    This review is aimed at presenting the state-of-the-art of time domain (TD) functional near-infrared spectroscopy (fNIRS). We first introduce the physical principles, the basics of modeling and data analysis. Basic instrumentation components (light sources, detection techniques, and delivery and collection systems) of a TD fNIRS system are described. A survey of past, existing and next generation TD fNIRS systems used for research and clinical studies is presented. Performance assessment of TD fNIRS systems and standardization issues are also discussed. Main strengths and weakness of TD fNIRS are highlighted, also in comparison with continuous wave (CW) fNIRS. Issues like quantification of the hemodynamic response, penetration depth, depth selectivity, spatial resolution and contrast-to-noise ratio are critically examined, with the help of experimental results performed on phantoms or in vivo. Finally we give an account on the technological developments that would pave the way for a broader use of TD fNIRS in the neuroimaging community. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

  20. Escherichia coli lipoprotein binds human plasminogen via an intramolecular domain

    Directory of Open Access Journals (Sweden)

    Tammy eGonzalez

    2015-10-01

    Full Text Available Escherichia coli lipoprotein (Lpp is a major cellular component that exists in two distinct states, bound-form and free-form. Bound-form Lpp is known to interact with the periplasmic bacterial cell wall, while free-form Lpp is localized to the bacterial cell surface. A function for surface-exposed Lpp has yet to be determined. We hypothesized that the presence of C-terminal lysines in the surface-exposed region of Lpp would facilitate binding to the host zymogen plasminogen, a protease commandeered by a number of clinically important bacteria. Recombinant Lpp was synthesized and the binding of Lpp to plasminogen, the effect of various inhibitors on this binding, and the effects of various mutations of Lpp on Lpp-plasminogen interactions were examined. Additionally, the ability of Lpp-bound plasminogen to be converted to active plasmin was analyzed. We determined that Lpp binds plasminogen via an atypical domain located near the center of mature Lpp that may not be exposed on the surface of intact E. coli according to the current localization model. Finally, we found that plasminogen bound by Lpp can be converted to active plasmin. While the consequences of Lpp binding plasminogen are unclear, these results prompt further investigation of the ability of surface exposed Lpp to interact with host molecules such as extracellular matrix components and complement regulators, and the role of these interactions in infections caused by E. coli and other bacteria.

  1. An Evaluation of the Performance Diagnostic Checklist-Human Services (PDC-HS) Across Domains.

    Science.gov (United States)

    Wilder, David A; Lipschultz, Joshua; Gehrman, Chana

    2018-06-01

    The Performance Diagnostic Checklist-Human Services (PDC-HS) is an informant-based tool designed to assess the environmental variables that contribute to poor employee performance in human service settings. Although the PDC-HS has been shown to effectively identify variables contributing to problematic performance, interventions based on only two of the four PDC-HS domains have been evaluated to date. In addition, the extent to which PDC-HS-indicated interventions are more effective than nonindicated interventions for two domains remains unclear. In the current study, we administered the PDC-HS to supervisors to assess the variables contributing to infrequent teaching of verbal operants and use of a timer by therapists at a center-based autism treatment program. Each of the four PDC-HS domains was identified as contributing to poor performance for at least one therapist. We then evaluated PDC-HS-indicated interventions for each domain. In addition, to assess the predictive validity of the tool, we evaluated various nonindicated interventions prior to implementing a PDC-HS-indicated intervention for two of the four domains. Results suggest that the PDC-HS-indicated interventions were effective across all four domains and were more effective than the nonindicated interventions for the two domains for which they were evaluated. Results are discussed in terms of the utility of the PDC-HS to identify appropriate interventions to manage therapist performance in human service settings.

  2. Influence of lung CT changes in chronic obstructive pulmonary disease (COPD on the human lung microbiome.

    Directory of Open Access Journals (Sweden)

    Marion Engel

    Full Text Available Changes in microbial community composition in the lung of patients suffering from moderate to severe COPD have been well documented. However, knowledge about specific microbiome structures in the human lung associated with CT defined abnormalities is limited.Bacterial community composition derived from brush samples from lungs of 16 patients suffering from different CT defined subtypes of COPD and 9 healthy subjects was analyzed using a cultivation independent barcoding approach applying 454-pyrosequencing of 16S rRNA gene fragment amplicons.We could show that bacterial community composition in patients with changes in CT (either airway or emphysema type changes, designated as severe subtypes was different from community composition in lungs of patients without visible changes in CT as well as from healthy subjects (designated as mild COPD subtype and control group (PC1, Padj = 0.002. Higher abundance of Prevotella in samples from patients with mild COPD subtype and from controls and of Streptococcus in the severe subtype cases mainly contributed to the separation of bacterial communities of subjects. No significant effects of treatment with inhaled glucocorticoids on bacterial community composition were detected within COPD cases with and without abnormalities in CT in PCoA. Co-occurrence analysis suggests the presence of networks of co-occurring bacteria. Four communities of positively correlated bacteria were revealed. The microbial communities can clearly be distinguished by their associations with the CT defined disease phenotype.Our findings indicate that CT detectable structural changes in the lung of COPD patients, which we termed severe subtypes, are associated with alterations in bacterial communities, which may induce further changes in the interaction between microbes and host cells. This might result in a changed interplay with the host immune system.

  3. Influence of lung CT changes in chronic obstructive pulmonary disease (COPD) on the human lung microbiome.

    Science.gov (United States)

    Engel, Marion; Endesfelder, David; Schloter-Hai, Brigitte; Kublik, Susanne; Granitsiotis, Michael S; Boschetto, Piera; Stendardo, Mariarita; Barta, Imre; Dome, Balazs; Deleuze, Jean-François; Boland, Anne; Müller-Quernheim, Joachim; Prasse, Antje; Welte, Tobias; Hohlfeld, Jens; Subramanian, Deepak; Parr, David; Gut, Ivo Glynne; Greulich, Timm; Koczulla, Andreas Rembert; Nowinski, Adam; Gorecka, Dorota; Singh, Dave; Gupta, Sumit; Brightling, Christopher E; Hoffmann, Harald; Frankenberger, Marion; Hofer, Thomas P; Burggraf, Dorothe; Heiss-Neumann, Marion; Ziegler-Heitbrock, Loems; Schloter, Michael; Zu Castell, Wolfgang

    2017-01-01

    Changes in microbial community composition in the lung of patients suffering from moderate to severe COPD have been well documented. However, knowledge about specific microbiome structures in the human lung associated with CT defined abnormalities is limited. Bacterial community composition derived from brush samples from lungs of 16 patients suffering from different CT defined subtypes of COPD and 9 healthy subjects was analyzed using a cultivation independent barcoding approach applying 454-pyrosequencing of 16S rRNA gene fragment amplicons. We could show that bacterial community composition in patients with changes in CT (either airway or emphysema type changes, designated as severe subtypes) was different from community composition in lungs of patients without visible changes in CT as well as from healthy subjects (designated as mild COPD subtype and control group) (PC1, Padj = 0.002). Higher abundance of Prevotella in samples from patients with mild COPD subtype and from controls and of Streptococcus in the severe subtype cases mainly contributed to the separation of bacterial communities of subjects. No significant effects of treatment with inhaled glucocorticoids on bacterial community composition were detected within COPD cases with and without abnormalities in CT in PCoA. Co-occurrence analysis suggests the presence of networks of co-occurring bacteria. Four communities of positively correlated bacteria were revealed. The microbial communities can clearly be distinguished by their associations with the CT defined disease phenotype. Our findings indicate that CT detectable structural changes in the lung of COPD patients, which we termed severe subtypes, are associated with alterations in bacterial communities, which may induce further changes in the interaction between microbes and host cells. This might result in a changed interplay with the host immune system.

  4. Skeletal and total body volumes of human fetuses: assessment of reference data by spiral CT

    International Nuclear Information System (INIS)

    Braillon, Pierre M.; Buenerd, Annie; Bouvier, Raymonde; Lapillonne, Alexandre

    2002-01-01

    Objective: To define reference data for skeletal and total body volumes of normal human fetuses. Materials and methods: Spiral CT was used to assess the skeletal and total body volumes of 31 normal human stillborn infants with gestational age (GA) and body weight (BW) ranging from 14 to 41.5 weeks and 22 to 3,760 g, respectively. CT scans (slice thickness 2.7 mm, pitch 0.7) were performed within the first 24 h after delivery. Precise bone and soft-tissue windows were defined from analysis of the density along the diaphysis of the fetal long bones and from the measurement of a phantom that mimics soft tissues. Lengths and volumes were obtained from 3D reconstructions. The femur lengths measured from CT images (FLct) were compared with those provided by US studies (FLus). Results: Significant correlations (r>0.9) were found between BW, measured volumes of the entire skeleton or head, long-bone lengths, biparietal diameter and GA. Strong linear correlations (r>0.98) were observed between FLct and FLus. Conclusions: Skeletal and total body volume values obtained using spiral CT were significantly correlated with fetal biometric measurements. These data could complement those obtained in obstetric investigations with US. (orig.)

  5. Technical Note: Insertion of digital lesions in the projection domain for dual-source, dual-energy CT.

    Science.gov (United States)

    Ferrero, Andrea; Chen, Baiyu; Li, Zhoubo; Yu, Lifeng; McCollough, Cynthia

    2017-05-01

    To compare algorithms performing material decomposition and classification in dual-energy CT, it is desirable to know the ground truth of the lesion to be analyzed in real patient data. In this work, we developed and validated a framework to insert digital lesions of arbitrary chemical composition into patient projection data acquired on a dual-source, dual-energy CT system. A model that takes into account beam-hardening effects was developed to predict the CT number of objects with known chemical composition. The model utilizes information about the x-ray energy spectra, the patient/phantom attenuation, and the x-ray detector energy response. The beam-hardening model was validated on samples of iodine (I) and calcium (Ca) for a second-generation dual-source, dual-energy CT scanner for all tube potentials available and a wide range of patient sizes. The seven most prevalent mineral components of renal stones were modeled and digital stones were created with CT numbers computed for each patient/phantom size and x-ray energy spectra using the developed beam-hardening model. Each digital stone was inserted in the dual-energy projection data of a water phantom scanned on a dual-source scanner and reconstructed with the routine algorithms in use in our practice. The geometry of the forward projection for dual-energy data was validated by comparing CT number accuracy and high-contrast resolution of simulated dual-energy CT data of the ACR phantom with experimentally acquired data. The beam-hardening model and forward projection method accurately predicted the CT number of I and Ca over a wide range of tube potentials and phantom sizes. The images reconstructed after the insertion of digital kidney stones were consistent with the images reconstructed from the scanner, and the CT number ratios for different kidney stone types were consistent with data in the literature. A sample application of the proposed tool was also demonstrated. A framework was developed and validated

  6. Polycystic ovary syndrome: association of a C/T single nucleotide polymorphism at tyrosine kinase domain of insulin receptor gene with pathogenesis among lean Japanese women.

    Science.gov (United States)

    Kashima, Katsunori; Yahata, Tetsuro; Fujita, Kazuyuki; Tanaka, Kenichi

    2013-01-01

    To assess whether the insulin receptor (INSR) gene contributes to genetic susceptibility to polycystic ovary syndrome (PCOS) in a Japanese population. We ex-amined the frequency of the His 1058 C/T single nucleotide polymorphism (SNP) found in exon 17 of the INSR gene in 61 Japanese PCOS patients and 99 Japanese healthy controls. In addition, we analyzed the association between the genotype of this SNP and the clinical phenotypes. The frequency of the C/C genotype was not significantly different between all PCOS patients (47.5%) and controls (35.4%). However, among the lean cases (body mass index PCOS patients (65.0%) as compared with controls (36.6%). We concluded that the His 1058 C/T polymorphism at the tyrosine kinase domain of the INSR gene had a relationship to the pathogenesis of lean PCOS patients in a Japanese population.

  7. Domain altering SNPs in the human proteome and their impact on signaling pathways.

    Directory of Open Access Journals (Sweden)

    Yichuan Liu

    Full Text Available Single nucleotide polymorphisms (SNPs constitute an important mode of genetic variations observed in the human genome. A small fraction of SNPs, about four thousand out of the ten million, has been associated with genetic disorders and complex diseases. The present study focuses on SNPs that fall on protein domains, 3D structures that facilitate connectivity of proteins in cell signaling and metabolic pathways. We scanned the human proteome using the PROSITE web tool and identified proteins with SNP containing domains. We showed that SNPs that fall on protein domains are highly statistically enriched among SNPs linked to hereditary disorders and complex diseases. Proteins whose domains are dramatically altered by the presence of an SNP are even more likely to be present among proteins linked to hereditary disorders. Proteins with domain-altering SNPs comprise highly connected nodes in cellular pathways such as the focal adhesion, the axon guidance pathway and the autoimmune disease pathways. Statistical enrichment of domain/motif signatures in interacting protein pairs indicates extensive loss of connectivity of cell signaling pathways due to domain-altering SNPs, potentially leading to hereditary disorders.

  8. Structure determination of human Lck unique and SH3 domains by nuclear magnetic resonance spectroscopy

    Directory of Open Access Journals (Sweden)

    Willbold Dieter

    2003-05-01

    Full Text Available Abstract Background Protein tyrosine kinases are involved in signal transduction pathways that regulate cell growth, differentiation, activation and transformation. Human lymphocyte specific kinase (Lck is a 56 kDa protein involved in T-cell- and IL2-receptor signaling. Three-dimensional structures are known for SH3, SH2 and kinase domains of Lck as well as for other tyrosine kinases. No structure is known for the unique domain of any Src-type tyrosine kinase. Results Lck(1–120 comprising unique and SH3 domains was structurally investigated by nuclear magnetic resonance spectroscopy. We found the unique domain, in contrast to the SH3 part, to have basically no defined structural elements. The solution structure of the SH3 part could be determined with very high precision. It does not show significant differences to Lck SH3 in the absence of the unique domain. Minor differences were observed to the X-ray structure of Lck SH3. Conclusion The unique domain of Lck does not contain any defined structure elements in the absence of ligands and membranes. Presence of the unique domain is not relevant to the three-dimensional structure of the Lck SH3 domain.

  9. Megabase replication domains along the human genome: relation to chromatin structure and genome organisation.

    Science.gov (United States)

    Audit, Benjamin; Zaghloul, Lamia; Baker, Antoine; Arneodo, Alain; Chen, Chun-Long; d'Aubenton-Carafa, Yves; Thermes, Claude

    2013-01-01

    In higher eukaryotes, the absence of specific sequence motifs, marking the origins of replication has been a serious hindrance to the understanding of (i) the mechanisms that regulate the spatio-temporal replication program, and (ii) the links between origins activation, chromatin structure and transcription. In this chapter, we review the partitioning of the human genome into megabased-size replication domains delineated as N-shaped motifs in the strand compositional asymmetry profiles. They collectively span 28.3% of the genome and are bordered by more than 1,000 putative replication origins. We recapitulate the comparison of this partition of the human genome with high-resolution experimental data that confirms that replication domain borders are likely to be preferential replication initiation zones in the germline. In addition, we highlight the specific distribution of experimental and numerical chromatin marks along replication domains. Domain borders correspond to particular open chromatin regions, possibly encoded in the DNA sequence, and around which replication and transcription are highly coordinated. These regions also present a high evolutionary breakpoint density, suggesting that susceptibility to breakage might be linked to local open chromatin fiber state. Altogether, this chapter presents a compartmentalization of the human genome into replication domains that are landmarks of the human genome organization and are likely to play a key role in genome dynamics during evolution and in pathological situations.

  10. Epitope mapping of functional domains of human factor V with human and mouse monoclonal antibodies

    International Nuclear Information System (INIS)

    Annamalai, A.E.; Rao, A.K.; Chiu, H.C.; Wang, D.; Dutta-Roy, A.K.; Colman, R.W.

    1986-01-01

    The authors previously described two human monoclonal antibodies (MAbs) which inactivated factor V. The authors have now purified the predominant antibody (H2) on protein A Sepharose using a pH gradient and typed it as IgG 1 ,. Immunoprecipitation of 125 I-human factor Va with H2 demonstrated specificity for the heavy chain (D), Mr = 105,000. The authors compared using ELISA the competitive binding to factor Va, of H2, H1 and two mouse MAbs, B38 (directed to E) and B10 (to activation peptide, Cl). All four antibodies recognized distinct epitopes in factor V with steric overlap in some cases. Factor Xa showed a concentration dependent competition for binding of H1, H2 and B38 but not B10 to factor V/Va in ELISA. All MAbs bound to factor V/Va in the absence of Ca ++ . However, Ca ++ at 8 mM increased the binding of H1 and H2 to 165% and 360% and did not have any effect on the binding of either mouse MAbs. Prothrombin at a concentration of up to 400 μg/ml did not inhibit binding of any of these antibodies. Thus, both the light (E) and heavy (D) chains of factor Va but not the activation peptide (Cl) interact with factor Xa as defined by the MAbs. In addition, sites on both chains for Ca ++ are recognized by particular MAbs (H1 and H2). These studies increase their knowledge of the interactions of factor V domains in the formation of prothrombinase complex

  11. Correlation between model observer and human observer performance in CT imaging when lesion location is uncertain

    Energy Technology Data Exchange (ETDEWEB)

    Leng, Shuai; Yu, Lifeng; Zhang, Yi; McCollough, Cynthia H. [Department of Radiology, Mayo Clinic, 200 First Street Southwest, Rochester, Minnesota 55905 (United States); Carter, Rickey [Department of Biostatistics, Mayo Clinic, 200 First Street Southwest, Rochester, Minnesota 55905 (United States); Toledano, Alicia Y. [Biostatistics Consulting, LLC, 10606 Wheatley Street, Kensington, Maryland 20895 (United States)

    2013-08-15

    Purpose: The purpose of this study was to investigate the correlation between model observer and human observer performance in CT imaging for the task of lesion detection and localization when the lesion location is uncertain.Methods: Two cylindrical rods (3-mm and 5-mm diameters) were placed in a 35 × 26 cm torso-shaped water phantom to simulate lesions with −15 HU contrast at 120 kV. The phantom was scanned 100 times on a 128-slice CT scanner at each of four dose levels (CTDIvol = 5.7, 11.4, 17.1, and 22.8 mGy). Regions of interest (ROIs) around each lesion were extracted to generate images with signal-present, with each ROI containing 128 × 128 pixels. Corresponding ROIs of signal-absent images were generated from images without lesion mimicking rods. The location of the lesion (rod) in each ROI was randomly distributed by moving the ROIs around each lesion. Human observer studies were performed by having three trained observers identify the presence or absence of lesions, indicating the lesion location in each image and scoring confidence for the detection task on a 6-point scale. The same image data were analyzed using a channelized Hotelling model observer (CHO) with Gabor channels. Internal noise was added to the decision variables for the model observer study. Area under the curve (AUC) of ROC and localization ROC (LROC) curves were calculated using a nonparametric approach. The Spearman's rank order correlation between the average performance of the human observers and the model observer performance was calculated for the AUC of both ROC and LROC curves for both the 3- and 5-mm diameter lesions.Results: In both ROC and LROC analyses, AUC values for the model observer agreed well with the average values across the three human observers. The Spearman's rank order correlation values for both ROC and LROC analyses for both the 3- and 5-mm diameter lesions were all 1.0, indicating perfect rank ordering agreement of the figures of merit (AUC

  12. Human epidermal growth factor receptor2 expression in unresectable gastric cancers: Relationship with CT characteristics

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Jeong Sub [Dept. of Radiology, Jeju National University Hospital, Jeju (Korea, Republic of); Kim, Se Hyung; Im, Seock Ah; Kim, Min A; Han, Joon Koo [Seoul National University Hospital, Seoul (Korea, Republic of)

    2017-09-15

    To retrospectively analyze the qualitative CT features that correlate with human epidermal growth factor receptor 2 (HER2)-expression in pathologically-proven gastric cancers. A total of 181 patients with pathologically-proven unresectable gastric cancers with HER2-expression (HER2-positive [n = 32] and negative [n = 149]) were included. CT features of primary gastric and metastatic tumors were reviewed. The prevalence of each CT finding was compared in both groups. Thereafter, binary logistic regression determined the most significant differential CT features. Clinical outcomes were compared using Kaplan-Meier method. HER2-postive cancers showed lower clinical T stage (21.9% vs. 8.1%; p = 0.015), hyperattenuation on portal phase (62.5% vs. 30.9%; p = 0.003), and was more frequently metastasized to the liver (62.5% vs. 32.2%; p = 0.001), than HER2-negative cancers. On binary regression analysis, hyperattenuation of the tumor (odds ratio [OR], 4.68; p < 0.001) and hepatic metastasis (OR, 4.43; p = 0.001) were significant independent factors that predict HER2-positive cancers. Median survival of HER2-positive cancers (13.7 months) was significantly longer than HER2-negative cancers (9.6 months) (p = 0.035). HER2-positive gastric cancers show less-advanced T stage, hyperattenuation on the portal phase, and frequently metastasize to the liver, as compared to HER2-negative cancers.

  13. Human epidermal growth factor receptor2 expression in unresectable gastric cancers: Relationship with CT characteristics

    International Nuclear Information System (INIS)

    Lee, Jeong Sub; Kim, Se Hyung; Im, Seock Ah; Kim, Min A; Han, Joon Koo

    2017-01-01

    To retrospectively analyze the qualitative CT features that correlate with human epidermal growth factor receptor 2 (HER2)-expression in pathologically-proven gastric cancers. A total of 181 patients with pathologically-proven unresectable gastric cancers with HER2-expression (HER2-positive [n = 32] and negative [n = 149]) were included. CT features of primary gastric and metastatic tumors were reviewed. The prevalence of each CT finding was compared in both groups. Thereafter, binary logistic regression determined the most significant differential CT features. Clinical outcomes were compared using Kaplan-Meier method. HER2-postive cancers showed lower clinical T stage (21.9% vs. 8.1%; p = 0.015), hyperattenuation on portal phase (62.5% vs. 30.9%; p = 0.003), and was more frequently metastasized to the liver (62.5% vs. 32.2%; p = 0.001), than HER2-negative cancers. On binary regression analysis, hyperattenuation of the tumor (odds ratio [OR], 4.68; p < 0.001) and hepatic metastasis (OR, 4.43; p = 0.001) were significant independent factors that predict HER2-positive cancers. Median survival of HER2-positive cancers (13.7 months) was significantly longer than HER2-negative cancers (9.6 months) (p = 0.035). HER2-positive gastric cancers show less-advanced T stage, hyperattenuation on the portal phase, and frequently metastasize to the liver, as compared to HER2-negative cancers

  14. Quantifying the impact of µCT-scanning of human fossil teeth on ESR age results.

    Science.gov (United States)

    Duval, Mathieu; Martín-Francés, Laura

    2017-05-01

    Fossil human teeth are nowadays systematically CT-scanned by palaeoanthropologists prior to any further analysis. It has been recently demonstrated that this noninvasive technique has, in most cases, virtually no influence on ancient DNA preservation. However, it may have nevertheless an impact on other techniques, like Electron Spin Resonance (ESR) dating, by artificially ageing the apparent age of the sample. To evaluate this impact, we µCT-scanned several modern enamel fragments following the standard analytical procedures employed by the Dental Anthropology Group at CENIEH, Spain, and then performed ESR dose reconstruction for each of them. The results of our experiment demonstrate that the systematic high-resolution µCT-scanning of fossil hominin remains introduces a nonnegligible X-ray dose into the tooth enamel, equivalent to 15-30 Gy depending on the parameters used. This dose may be multiplied by a factor of ∼8 if no metallic filter is used. However, this dose estimate cannot be universally extrapolated to any µCT-scan experiment but has instead to be specifically assessed for each device and set of parameters employed. The impact on the ESR age results is directly dependent on the magnitude of the geological dose measured in fossil enamel but could potentially lead to an age overestimation up to 40% in case of Late Pleistocene samples, if not taken into consideration. © 2017 Wiley Periodicals, Inc.

  15. Domain-specific rationality in human choices: violations of utility axioms and social contexts.

    Science.gov (United States)

    Wang, X T

    1996-07-01

    This study presents a domain-specific view of human decision rationality. It explores social and ecological domain-specific psychological mechanisms underlying choice biases and violations of utility axioms. Results from both the USA and China revealed a social group domain-specific choice pattern. The irrational preference reversal in a hypothetical life-death decision problem (a classical example of framing effects) was eliminated by providing a small group or family context in which most subjects favored a risky choice option regardless of the positive/negative framing of choice outcomes. The risk preference data also indicate that the subjective scope of small group domain is larger for Chinese subjects, suggesting that human choice mechanisms are sensitive to culturally specific features of group living. A further experiment provided evidence that perceived fairness might be one major factor regulating the choice preferences found in small group (kith-and-kin) contexts. Finally, the violation of the stochastic dominance axiom of the rational theory of choice was predicted and tested. The violations were found only when the "life-death" problem was presented in small group contexts; the strongest violation was found in a family context. These results suggest that human decisions and choices are regulated by domain-specific choice mechanisms designed to solve evolutionary recurrent and adaptively important problems.

  16. Structure of the human histone chaperone FACT Spt16 N-terminal domain

    Energy Technology Data Exchange (ETDEWEB)

    Marcianò, G.; Huang, D. T., E-mail: d.huang@beatson.gla.ac.uk [Cancer Research UK Beatson Institute, Garscube Estate, Switchback Road, Glasgow G61 1BD, Scotland (United Kingdom)

    2016-01-22

    The Spt16–SSRP1 heterodimer is a histone chaperone that plays an important role in regulating chromatin assembly. Here, a crystal structure of the N-terminal domain of human Spt16 is presented and it is shown that this domain may contribute to histone binding. The histone chaperone FACT plays an important role in facilitating nucleosome assembly and disassembly during transcription. FACT is a heterodimeric complex consisting of Spt16 and SSRP1. The N-terminal domain of Spt16 resembles an inactive aminopeptidase. How this domain contributes to the histone chaperone activity of FACT remains elusive. Here, the crystal structure of the N-terminal domain (NTD) of human Spt16 is reported at a resolution of 1.84 Å. The structure adopts an aminopeptidase-like fold similar to those of the Saccharomyces cerevisiae and Schizosaccharomyces pombe Spt16 NTDs. Isothermal titration calorimetry analyses show that human Spt16 NTD binds histones H3/H4 with low-micromolar affinity, suggesting that Spt16 NTD may contribute to histone binding in the FACT complex. Surface-residue conservation and electrostatic analysis reveal a conserved acidic patch that may be involved in histone binding.

  17. Structure of the human histone chaperone FACT Spt16 N-terminal domain

    International Nuclear Information System (INIS)

    Marcianò, G.; Huang, D. T.

    2016-01-01

    The Spt16–SSRP1 heterodimer is a histone chaperone that plays an important role in regulating chromatin assembly. Here, a crystal structure of the N-terminal domain of human Spt16 is presented and it is shown that this domain may contribute to histone binding. The histone chaperone FACT plays an important role in facilitating nucleosome assembly and disassembly during transcription. FACT is a heterodimeric complex consisting of Spt16 and SSRP1. The N-terminal domain of Spt16 resembles an inactive aminopeptidase. How this domain contributes to the histone chaperone activity of FACT remains elusive. Here, the crystal structure of the N-terminal domain (NTD) of human Spt16 is reported at a resolution of 1.84 Å. The structure adopts an aminopeptidase-like fold similar to those of the Saccharomyces cerevisiae and Schizosaccharomyces pombe Spt16 NTDs. Isothermal titration calorimetry analyses show that human Spt16 NTD binds histones H3/H4 with low-micromolar affinity, suggesting that Spt16 NTD may contribute to histone binding in the FACT complex. Surface-residue conservation and electrostatic analysis reveal a conserved acidic patch that may be involved in histone binding

  18. Simplified derivation of stopping power ratio in the human body from dual-energy CT data.

    Science.gov (United States)

    Saito, Masatoshi; Sagara, Shota

    2017-08-01

    The main objective of this study is to propose an alternative parameterization for the empirical relation between mean excitation energies (I-value) and effective atomic numbers (Z eff ) of human tissues, and to present a simplified formulation (which we called DEEDZ-SPR) for deriving the stopping power ratio (SPR) from dual-energy (DE) CT data via electron density (ρ e ) and Z eff calibration. We performed a numerical analysis of this DEEDZ-SPR method for the human-body-equivalent tissues of ICRU Report 46, as objects of interest with unknown SPR and ρ e . The attenuation coefficients of these materials were calculated using the XCOM photon cross-sections database. We also applied the DEEDZ-SPR conversion to experimental DECT data available in the literature, which was measured for the tissue-characterization phantom using a dual-source CT scanner at 80 kV and 140 kV/Sn. It was found that the DEEDZ-SPR conversion enables the calculation of SPR simply by means of the weighted subtraction of an electron-density image and a low- or high-kV CT image. The simulated SPRs were in excellent agreement with the reference values over the SPR range from 0.258 (lung) to 3.638 (bone mineral-hydroxyapatite). The relative deviations from the reference SPR were within ±0.6% for all ICRU-46 human tissues, except for the thyroid that presented a -1.1% deviation. The overall root-mean-square error was 0.21%. Application to experimental DECT data confirmed this agreement within the experimental accuracy, which demonstrates the practical feasibility of the method. The DEEDZ-SPR conversion method could facilitate the construction of SPR images as accurately as a recent DECT-based calibration procedure of SPR parameterization based directly on the CT numbers in a DECT data set. © 2017 American Association of Physicists in Medicine.

  19. Recombinant expression in E. coli of human FGFR2 with its transmembrane and extracellular domains

    Directory of Open Access Journals (Sweden)

    Adam Bajinting

    2017-06-01

    Full Text Available Fibroblast growth factor receptors (FGFRs are a family of receptor tyrosine kinases containing three domains: an extracellular receptor domain, a single transmembrane helix, and an intracellular tyrosine kinase domain. FGFRs are activated by fibroblast growth factors (FGFs as part of complex signal transduction cascades regulating angiogenesis, skeletal formation, cell differentiation, proliferation, cell survival, and cancer. We have developed the first recombinant expression system in E. coli to produce a construct of human FGFR2 containing its transmembrane and extracellular receptor domains. We demonstrate that the expressed construct is functional in binding heparin and dimerizing. Size exclusion chromatography demonstrates that the purified FGFR2 does not form a complex with FGF1 or adopts an inactive dimer conformation. Progress towards the successful recombinant production of intact FGFRs will facilitate further biochemical experiments and structure determination that will provide insight into how extracellular FGF binding activates intracellular kinase activity.

  20. Truncation artifact suppression in cone-beam radionuclide transmission CT using maximum likelihood techniques: evaluation with human subjects

    International Nuclear Information System (INIS)

    Manglos, S.H.

    1992-01-01

    Transverse image truncation can be a serious problem for human imaging using cone-beam transmission CT (CB-CT) implemented on a conventional rotating gamma camera. This paper presents a reconstruction method to reduce or eliminate the artifacts resulting from the truncation. The method uses a previously published transmission maximum likelihood EM algorithm, adapted to the cone-beam geometry. The reconstruction method is evaluated qualitatively using three human subjects of various dimensions and various degrees of truncation. (author)

  1. Data describing the solution structure of the WW3* domain from human Nedd4-1

    Directory of Open Access Journals (Sweden)

    Vineet Panwalkar

    2016-09-01

    Full Text Available The third WW domain (WW3* of human Nedd4-1 (Neuronal precursor cell expressed developmentally down-regulated gene 4-1 interacts with the poly-proline (PY motifs of the human epithelial Na+ channel (hENaC subunits at micromolar affinity. This data supplements the article (Panwalkar et al., 2015 [1]. We describe the NMR experiments used to solve the solution structure of the WW3* domain. We also present NOE network data for defining the rotameric state of side chains of peptide binding residues, and complement this data with χ1 dihedral angles derived from 3J couplings and molecular dynamics simulations data. Keywords: Chemical shift, Neuronal precursor cell expressed developmentally down-regulated gene 4-1, NMR, NOE distance restraints, WW domain

  2. Crystal structure of the N domain of human somatic angiotensin I-converting enzyme provides a structural basis for domain-specific inhibitor design.

    Science.gov (United States)

    Corradi, Hazel R; Schwager, Sylva L U; Nchinda, Aloysius T; Sturrock, Edward D; Acharya, K Ravi

    2006-03-31

    Human somatic angiotensin I-converting enzyme (sACE) is a key regulator of blood pressure and an important drug target for combating cardiovascular and renal disease. sACE comprises two homologous metallopeptidase domains, N and C, joined by an inter-domain linker. Both domains are capable of cleaving the two hemoregulatory peptides angiotensin I and bradykinin, but differ in their affinities for a range of other substrates and inhibitors. Previously we determined the structure of testis ACE (C domain); here we present the crystal structure of the N domain of sACE (both in the presence and absence of the antihypertensive drug lisinopril) in order to aid the understanding of how these two domains differ in specificity and function. In addition, the structure of most of the inter-domain linker allows us to propose relative domain positions for sACE that may contribute to the domain cooperativity. The structure now provides a platform for the design of "domain-specific" second-generation ACE inhibitors.

  3. Overexpression, purification and crystallization of the two C-terminal domains of the bifunctional cellulase ctCel9D-Cel44A from Clostridium thermocellum

    International Nuclear Information System (INIS)

    Najmudin, Shabir; Guerreiro, Catarina I. P. D.; Ferreira, Luís M. A.; Romão, Maria J. C.; Fontes, Carlos M. G. A.; Prates, José A. M.

    2005-01-01

    The two C-terminal domains of the cellulase ctCel9D-Cel44A from C. thermocellum cellulosome have been crystallized in tetragonal space group P4 3 2 1 2 and X-ray diffraction data have been collected to 2.1 and 2.8 Å from native and seleno-l-methionine-derivative crystals, respectively. Clostridium thermocellum produces a highly organized multi-enzyme complex of cellulases and hemicellulases for the hydrolysis of plant cell-wall polysaccharides, which is termed the cellulosome. The bifunctional multi-modular cellulase ctCel9D-Cel44A is one of the largest components of the C. thermocellum cellulosome. The enzyme contains two internal catalytic domains belonging to glycoside hydrolase families 9 and 44. The C-terminus of this cellulase, comprising a polycystic kidney-disease module (PKD) and a carbohydrate-binding module (CBM44), has been crystallized. The crystals belong to the tetragonal space group P4 3 2 1 2, containing a single molecule in the asymmetric unit. Native and seleno-l-methionine-derivative crystals diffracted to 2.1 and 2.8 Å, respectively

  4. Overexpression, purification and crystallization of the two C-terminal domains of the bifunctional cellulase ctCel9D-Cel44A from Clostridium thermocellum

    Energy Technology Data Exchange (ETDEWEB)

    Najmudin, Shabir [REQUIMTE, Departamento de Química, FCT-UNL, 2829-516 Caparica (Portugal); Guerreiro, Catarina I. P. D.; Ferreira, Luís M. A. [CIISA - Faculdade de Medicina Veterinária, Universidade Técnica de Lisboa, Avenida da Universidade Técnica, 1300-477 Lisboa (Portugal); Romão, Maria J. C. [REQUIMTE, Departamento de Química, FCT-UNL, 2829-516 Caparica (Portugal); Fontes, Carlos M. G. A.; Prates, José A. M., E-mail: japrates@fmv.utl.pt [CIISA - Faculdade de Medicina Veterinária, Universidade Técnica de Lisboa, Avenida da Universidade Técnica, 1300-477 Lisboa (Portugal); REQUIMTE, Departamento de Química, FCT-UNL, 2829-516 Caparica (Portugal)

    2005-12-01

    The two C-terminal domains of the cellulase ctCel9D-Cel44A from C. thermocellum cellulosome have been crystallized in tetragonal space group P4{sub 3}2{sub 1}2 and X-ray diffraction data have been collected to 2.1 and 2.8 Å from native and seleno-l-methionine-derivative crystals, respectively. Clostridium thermocellum produces a highly organized multi-enzyme complex of cellulases and hemicellulases for the hydrolysis of plant cell-wall polysaccharides, which is termed the cellulosome. The bifunctional multi-modular cellulase ctCel9D-Cel44A is one of the largest components of the C. thermocellum cellulosome. The enzyme contains two internal catalytic domains belonging to glycoside hydrolase families 9 and 44. The C-terminus of this cellulase, comprising a polycystic kidney-disease module (PKD) and a carbohydrate-binding module (CBM44), has been crystallized. The crystals belong to the tetragonal space group P4{sub 3}2{sub 1}2, containing a single molecule in the asymmetric unit. Native and seleno-l-methionine-derivative crystals diffracted to 2.1 and 2.8 Å, respectively.

  5. Organizing to Understand: How to Operate Effectively in the Human Domain

    Science.gov (United States)

    2015-05-21

    was both entering and creating when it overthrew Saddam Hussein and dismantled the Iraqi government and security forces. The research examines the...sponsored initiative to help tactical and operational level commanders understand the human terrain, the “social, ethnographic , cultural, economic, and...as an intelligence function within TRADOC “as the primary and enduring social science-based human domain research , analysis, and training capability

  6. Identification of a minimal functional linker in human topoisomerase I by domain swapping with Cre recombinase

    DEFF Research Database (Denmark)

    Hougaard, Rikke Frøhlich; Juul, Sissel; Vinther, Maria

    2008-01-01

    . In this study we replace 86 amino acids including the linker domain of the cellular type IB topoisomerase, human topoisomerase I, with four, six, or eight amino acids from the corresponding short loop region in Cre recombinase. In vitro characterization of the resulting chimeras, denoted Cropos, reveals...

  7. Structure and dynamics of the human pleckstrin DEP domain: distinct molecular features of a novel DEP domain subfamily.

    Science.gov (United States)

    Civera, Concepcion; Simon, Bernd; Stier, Gunter; Sattler, Michael; Macias, Maria J

    2005-02-01

    Pleckstrin1 is a major substrate for protein kinase C in platelets and leukocytes, and comprises a central DEP (disheveled, Egl-10, pleckstrin) domain, which is flanked by two PH (pleckstrin homology) domains. DEP domains display a unique alpha/beta fold and have been implicated in membrane binding utilizing different mechanisms. Using multiple sequence alignments and phylogenetic tree reconstructions, we find that 6 subfamilies of the DEP domain exist, of which pleckstrin represents a novel and distinct subfamily. To clarify structural determinants of the DEP fold and to gain further insight into the role of the DEP domain, we determined the three-dimensional structure of the pleckstrin DEP domain using heteronuclear NMR spectroscopy. Pleckstrin DEP shares main structural features with the DEP domains of disheveled and Epac, which belong to different DEP subfamilies. However, the pleckstrin DEP fold is distinct from these structures and contains an additional, short helix alpha4 inserted in the beta4-beta5 loop that exhibits increased backbone mobility as judged by NMR relaxation measurements. Based on sequence conservation, the helix alpha4 may also be present in the DEP domains of regulator of G-protein signaling (RGS) proteins, which are members of the same DEP subfamily. In pleckstrin, the DEP domain is surrounded by two PH domains. Structural analysis and charge complementarity suggest that the DEP domain may interact with the N-terminal PH domain in pleckstrin. Phosphorylation of the PH-DEP linker, which is required for pleckstrin function, could regulate such an intramolecular interaction. This suggests a role of the pleckstrin DEP domain in intramolecular domain interactions, which is distinct from the functions of other DEP domain subfamilies found so far.

  8. The impact of the human DNA topoisomerase II C-terminal domain on activity.

    Directory of Open Access Journals (Sweden)

    Emma L Meczes

    2008-03-01

    Full Text Available Type II DNA topoisomerases (topos are essential enzymes needed for the resolution of topological problems that occur during DNA metabolic processes. Topos carry out an ATP-dependent strand passage reaction whereby one double helix is passed through a transient break in another. Humans have two topoII isoforms, alpha and beta, which while enzymatically similar are differentially expressed and regulated, and are thought to have different cellular roles. The C-terminal domain (CTD of the enzyme has the most diversity, and has been implicated in regulation. We sought to investigate the impact of the CTD domain on activity.We have investigated the role of the human topoII C-terminal domain by creating constructs encoding C-terminally truncated recombinant topoIIalpha and beta and topoIIalpha+beta-tail and topoIIbeta+alpha-tail chimeric proteins. We then investigated function in vivo in a yeast system, and in vitro in activity assays. We find that the C-terminal domain of human topoII isoforms is needed for in vivo function of the enzyme, but not needed for cleavage activity. C-terminally truncated enzymes had similar strand passage activity to full length enzymes, but the presence of the opposite C-terminal domain had a large effect, with the topoIIalpha-CTD increasing activity, and the topoIIbeta-CTD decreasing activity.In vivo complementation data show that the topoIIalpha C-terminal domain is needed for growth, but the topoIIbeta isoform is able to support low levels of growth without a C-terminal domain. This may indicate that topoIIbeta has an additional localisation signal. In vitro data suggest that, while the lack of any C-terminal domain has little effect on activity, the presence of either the topoIIalpha or beta C-terminal domain can affect strand passage activity. Data indicates that the topoIIbeta-CTD may be a negative regulator. This is the first report of in vitro data with chimeric human topoIIs.

  9. Evaluation of automatic image quality assessment in chest CT - A human cadaver study.

    Science.gov (United States)

    Franck, Caro; De Crop, An; De Roo, Bieke; Smeets, Peter; Vergauwen, Merel; Dewaele, Tom; Van Borsel, Mathias; Achten, Eric; Van Hoof, Tom; Bacher, Klaus

    2017-04-01

    The evaluation of clinical image quality (IQ) is important to optimize CT protocols and to keep patient doses as low as reasonably achievable. Considering the significant amount of effort needed for human observer studies, automatic IQ tools are a promising alternative. The purpose of this study was to evaluate automatic IQ assessment in chest CT using Thiel embalmed cadavers. Chest CT's of Thiel embalmed cadavers were acquired at different exposures. Clinical IQ was determined by performing a visual grading analysis. Physical-technical IQ (noise, contrast-to-noise and contrast-detail) was assessed in a Catphan phantom. Soft and sharp reconstructions were made with filtered back projection and two strengths of iterative reconstruction. In addition to the classical IQ metrics, an automatic algorithm was used to calculate image quality scores (IQs). To be able to compare datasets reconstructed with different kernels, the IQs values were normalized. Good correlations were found between IQs and the measured physical-technical image quality: noise (ρ=-1.00), contrast-to-noise (ρ=1.00) and contrast-detail (ρ=0.96). The correlation coefficients between IQs and the observed clinical image quality of soft and sharp reconstructions were 0.88 and 0.93, respectively. The automatic scoring algorithm is a promising tool for the evaluation of thoracic CT scans in daily clinical practice. It allows monitoring of the image quality of a chest protocol over time, without human intervention. Different reconstruction kernels can be compared after normalization of the IQs. Copyright © 2017 Associazione Italiana di Fisica Medica. Published by Elsevier Ltd. All rights reserved.

  10. Comparative kinomics of human and chimpanzee reveal unique kinship and functional diversity generated by new domain combinations

    Directory of Open Access Journals (Sweden)

    Martin Juliette

    2008-12-01

    Full Text Available Abstract Background Phosphorylation by protein kinases is a common event in many cellular processes. Further, many kinases perform specialized roles and are regulated by non-kinase domains tethered to kinase domain. Perturbation in the regulation of kinases leads to malignancy. We have identified and analysed putative protein kinases encoded in the genome of chimpanzee which is a close evolutionary relative of human. Result The shared core biology between chimpanzee and human is characterized by many orthologous protein kinases which are involved in conserved pathways. Domain architectures specific to chimp/human kinases have been observed. Chimp kinases with unique domain architectures are characterized by deletion of one or more non-kinase domains in the human kinases. Interestingly, counterparts of some of the multi-domain human kinases in chimp are characterized by identical domain architectures but with kinase-like non-kinase domain. Remarkably, out of 587 chimpanzee kinases no human orthologue with greater than 95% sequence identity could be identified for 160 kinases. Variations in chimpanzee kinases compared to human kinases are brought about also by differences in functions of domains tethered to the catalytic kinase domain. For example, the heterodimer forming PB1 domain related to the fold of ubiquitin/Ras-binding domain is seen uniquely tethered to PKC-like chimpanzee kinase. Conclusion Though the chimpanzee and human are evolutionary very close, there are chimpanzee kinases with no close counterpart in the human suggesting differences in their functions. This analysis provides a direction for experimental analysis of human and chimpanzee protein kinases in order to enhance our understanding on their specific biological roles.

  11. Crystallization and preliminary crystallographic analysis of human LR11 Vps10p domain

    International Nuclear Information System (INIS)

    Nakata, Zenzaburo; Nagae, Masamichi; Yasui, Norihisa; Bujo, Hideaki; Nogi, Terukazu; Takagi, Junichi

    2010-01-01

    LR11/sorLA contains in its extracellular region a large (∼700-residue) Vps10p domain that is implicated in its intracellular protein-trafficking function. Here, the expression, purification, crystallization and preliminary crystallographic characterization of this domain are described. Low-density lipoprotein receptor (LDLR) relative with 11 binding repeats (LR11; also known as sorLA) is genetically associated with late-onset Alzheimer’s disease and is thought to be involved in neurodegenerative processes. LR11 contains a vacuolar protein-sorting 10 protein (Vps10p) domain. As this domain has been implicated in protein–protein interaction in other receptors, its structure and function are of great biological interest. Human LR11 Vps10p domain was expressed in mammalian cells and the purified protein was crystallized using the hanging-drop vapour-diffusion method. Enzymatic deglycosylation of the sample was critical to obtaining diffraction-quality crystals. Deglycosylated LR11 Vps10p-domain crystals belonged to the hexagonal space group P6 1 22. A diffraction data set was collected to 2.4 Å resolution and a clear molecular-replacement solution was obtained

  12. Lysophosphatidic acid stimulates thrombomodulin lectin-like domain shedding in human endothelial cells

    International Nuclear Information System (INIS)

    Wu Hualin; Lin ChiIou; Huang Yuanli; Chen, Pin-Shern; Kuo, Cheng-Hsiang; Chen, Mei-Shing; Wu, G.C.-C.; Shi, G.-Y.; Yang, H.-Y.; Lee Hsinyu

    2008-01-01

    Thrombomodulin (TM) is an anticoagulant glycoprotein highly expressed on endothelial cell surfaces. Increased levels of soluble TM in circulation have been widely accepted as an indicator of endothelial damage or dysfunction. Previous studies indicated that various proinflammatory factors stimulate TM shedding in various cell types such as smooth muscle cells and epithelial cells. Lysophosphatidic acid (LPA) is a bioactive lipid mediator present in biological fluids during endothelial damage or injury. In the present study, we first observed that LPA triggered TM shedding in human umbilical vein endothelial cells (HUVECs). By Cyflow analysis, we showed that the LPA-induced accessibility of antibodies to the endothelial growth factor (EGF)-like domain of TM is independent of matrix metalloproteinases (MMPs), while LPA-induced TM lectin-like domain shedding is MMP-dependent. Furthermore, a stable cell line expressing TM without its lectin-like domain exhibited a higher cell proliferation rate than a stable cell line expressing full-length TM. These results imply that LPA induces TM lectin-like domain shedding, which might contribute to the exposure of its EGF-like domain for EGF receptor (EGFR) binding, thereby stimulating subsequent cell proliferation. Based on our findings, we propose a novel mechanism for the exposure of TM EGF-like domain, which possibly mediates LPA-induced EGFR transactivation

  13. Human surfactant protein D: SP-D contains a C-type lectin carbohydrate recognition domain.

    Science.gov (United States)

    Rust, K; Grosso, L; Zhang, V; Chang, D; Persson, A; Longmore, W; Cai, G Z; Crouch, E

    1991-10-01

    Lung surfactant protein D (SP-D) shows calcium-dependent binding to specific saccharides, and is similar in domain structure to certain members of the calcium-dependent (C-type) lectin family. Using a degenerate oligomeric probe corresponding to a conserved peptide sequence derived from the amino-terminus of the putative carbohydrate binding domain of rat and bovine SP-D, we screened a human lung cDNA library and isolated a 1.4-kb cDNA for the human protein. The relationship of the cDNA to SP-D was established by several techniques including amino-terminal microsequencing of SP-D-derived peptides, and immunoprecipitation of translation products of transcribed mRNA with monospecific antibodies to SP-D. In addition, antibodies to a synthetic peptide derived from a predicted unique epitope within the carbohydrate recognition domain of SP-D specifically reacted with SP-D. DNA sequencing demonstrated a noncollagenous carboxy-terminal domain that is highly homologous with the carboxy-terminal globular domain of previously described C-type lectins. This domain contains all of the so-called "invariant residues," including four conserved cysteine residues, and shows high homology with the mannose-binding subfamily of C-type lectins. Sequencing also demonstrated an amino-terminal collagenous domain that contains an uninterrupted sequence of 59 Gly-X-Y triplets and that also contains the only identified consensus for asparagine-linked oligosaccharides. The studies demonstrate that SP-D is a member of the C-type lectin family, and confirm predicted structural similarities to conglutinin, SP-D, and the serum mannose binding proteins.

  14. Computational and human observer image quality evaluation of low dose, knowledge-based CT iterative reconstruction

    Energy Technology Data Exchange (ETDEWEB)

    Eck, Brendan L.; Fahmi, Rachid; Miao, Jun [Department of Biomedical Engineering, Case Western Reserve University, Cleveland, Ohio 44106 (United States); Brown, Kevin M.; Zabic, Stanislav; Raihani, Nilgoun [Philips Healthcare, Cleveland, Ohio 44143 (United States); Wilson, David L., E-mail: dlw@case.edu [Department of Biomedical Engineering, Case Western Reserve University, Cleveland, Ohio 44106 and Department of Radiology, Case Western Reserve University, Cleveland, Ohio 44106 (United States)

    2015-10-15

    Purpose: Aims in this study are to (1) develop a computational model observer which reliably tracks the detectability of human observers in low dose computed tomography (CT) images reconstructed with knowledge-based iterative reconstruction (IMR™, Philips Healthcare) and filtered back projection (FBP) across a range of independent variables, (2) use the model to evaluate detectability trends across reconstructions and make predictions of human observer detectability, and (3) perform human observer studies based on model predictions to demonstrate applications of the model in CT imaging. Methods: Detectability (d′) was evaluated in phantom studies across a range of conditions. Images were generated using a numerical CT simulator. Trained observers performed 4-alternative forced choice (4-AFC) experiments across dose (1.3, 2.7, 4.0 mGy), pin size (4, 6, 8 mm), contrast (0.3%, 0.5%, 1.0%), and reconstruction (FBP, IMR), at fixed display window. A five-channel Laguerre–Gauss channelized Hotelling observer (CHO) was developed with internal noise added to the decision variable and/or to channel outputs, creating six different internal noise models. Semianalytic internal noise computation was tested against Monte Carlo and used to accelerate internal noise parameter optimization. Model parameters were estimated from all experiments at once using maximum likelihood on the probability correct, P{sub C}. Akaike information criterion (AIC) was used to compare models of different orders. The best model was selected according to AIC and used to predict detectability in blended FBP-IMR images, analyze trends in IMR detectability improvements, and predict dose savings with IMR. Predicted dose savings were compared against 4-AFC study results using physical CT phantom images. Results: Detection in IMR was greater than FBP in all tested conditions. The CHO with internal noise proportional to channel output standard deviations, Model-k4, showed the best trade-off between fit

  15. SEM and microCT validation for en face OCT imagistic evaluation of endodontically treated human teeth

    Science.gov (United States)

    Negrutiu, Meda L.; Nica, Luminita; Sinescu, Cosmin; Topala, Florin; Ionita, Ciprian; Bradu, Adrian; Petrescu, Emanuela L.; Pop, Daniela M.; Rominu, Mihai; Podoleanu, Adrian Gh.

    2011-03-01

    Successful root canal treatment is based on diagnosis, treatment planning, knowledge of tooth anatomy, endodontic access cavity design, controlling the infection by thorough cleaning and shaping, methods and materials used in root canal obturation. An endodontic obturation must be a complete, three-dimensional filling of the root canal system, as close as possible to cemento-dentinal junction, without massive overfilling or underfilling. There are several known methods which are used to assess the quality of the endodontic sealing, but most are invasive. These lead to the destruction of the samples and often no conclusion could be drawn in respect to the existence of any microleakage in the investigated areas of interest. Using an time domain en-face OCT system, we have recently demonstrated real time thorough evaluation of quality of root canal fillings. The purpose of this in vitro study was to validate the en face OCT imagistic evaluation of endodontically treated human teeth by using scanning electron microscopy (SEM) and microcomputer tomography (μCT). SEM investigations evidenced the nonlinear aspect of the interface between the endodontic filling material and the root canal walls and materials defects in some samples. The results obtained by μCT revealed also some defects inside the root-canal filling and at the interfaces between the material and the root canal walls. The advantages of the OCT method consist in non-invasiveness and high resolution. In addition, en face OCT investigations permit visualization of the more complex stratified structure at the interface between the filling material and the dental hard tissue.

  16. Performance evaluation of phage-displayed synthetic human single-domain antibody libraries: A retrospective analysis.

    Science.gov (United States)

    Henry, Kevin A; Tanha, Jamshid

    2018-05-01

    Fully human synthetic single-domain antibodies (sdAbs) are desirable therapeutic molecules but their development is a considerable challenge. Here, using a retrospective analysis of in-house historical data, we examined the parameters that impact the outcome of screening phage-displayed synthetic human sdAb libraries to discover antigen-specific binders. We found no evidence for a differential effect of domain type (V H or V L ), library randomization strategy, incorporation of a stabilizing disulfide linkage or sdAb display format (monovalent vs. multivalent) on the probability of obtaining any antigen-binding human sdAbs, instead finding that the success of library screens was primarily related to properties of target antigens, especially molecular mass. The solubility and binding affinity of sdAbs isolated from successful screens depended both on properties of the sdAb libraries (primarily domain type) and the target antigens. Taking attrition of sdAbs with major manufacturability concerns (aggregation; low expression) and sdAbs that do not recognize native cell-surface antigens as independent probabilities, we calculate the overall likelihood of obtaining ≥1 antigen-binding human sdAb from a single library-target screen as ~24%. Successful library-target screens should be expected to yield ~1.3 human sdAbs on average, each with average binding affinity of ~2 μM. Copyright © 2018 Elsevier B.V. All rights reserved.

  17. Human Hsp70 molecular chaperone binds two calcium ions within the ATPase domain.

    Science.gov (United States)

    Sriram, M; Osipiuk, J; Freeman, B; Morimoto, R; Joachimiak, A

    1997-03-15

    The 70 kDa heat shock proteins (Hsp70) are a family of molecular chaperones, which promote protein folding and participate in many cellular functions. The Hsp70 chaperones are composed of two major domains. The N-terminal ATPase domain binds to and hydrolyzes ATP, whereas the C-terminal domain is required for polypeptide binding. Cooperation of both domains is needed for protein folding. The crystal structure of bovine Hsc70 ATPase domain (bATPase) has been determined and, more recently, the crystal structure of the peptide-binding domain of a related chaperone, DnaK, in complex with peptide substrate has been obtained. The molecular chaperone activity and conformational switch are functionally linked with ATP hydrolysis. A high-resolution structure of the ATPase domain is required to provide an understanding of the mechanism of ATP hydrolysis and how it affects communication between C- and N-terminal domains. The crystal structure of the human Hsp70 ATPase domain (hATPase) has been determined and refined at 1. 84 A, using synchrotron radiation at 120K. Two calcium sites were identified: the first calcium binds within the catalytic pocket, bridging ADP and inorganic phosphate, and the second calcium is tightly coordinated on the protein surface by Glu231, Asp232 and the carbonyl of His227. Overall, the structure of hATPase is similar to bATPase. Differences between them are found in the loops, the sites of amino acid substitution and the calcium-binding sites. Human Hsp70 chaperone is phosphorylated in vitro in the presence of divalent ions, calcium being the most effective. The structural similarity of hATPase and bATPase and the sequence similarity within the Hsp70 chaperone family suggest a universal mechanism of ATP hydrolysis among all Hsp70 molecular chaperones. Two calcium ions have been found in the hATPase structure. One corresponds to the magnesium site in bATPase and appears to be important for ATP hydrolysis and in vitro phosphorylation. Local changes

  18. N-terminal domains of human DNA polymerase lambda promote primer realignment during translesion DNA synthesis

    Science.gov (United States)

    Taggart, David J.; Dayeh, Daniel M.; Fredrickson, Saul W.; Suo, Zucai

    2014-01-01

    The X-family DNA polymerases λ (Polλ) and β (Polβ) possess similar 5′-2-deoxyribose-5-phosphatelyase (dRPase) and polymerase domains. Besides these domains, Polλ also possesses a BRCA1 C-terminal (BRCT) domain and a proline-rich domain at its N terminus. However, it is unclear how these non-enzymatic domains contribute to the unique biological functions of Polλ. Here, we used primer extension assays and a newly developed high-throughput short oligonucleotide sequencing assay (HT-SOSA) to compare the efficiency of lesion bypass and fidelity of human Polβ, Polλ and two N-terminal deletion constructs of Polλ during the bypass of either an abasic site or a 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodG) lesion. We demonstrate that the BRCT domain of Polλ enhances the efficiency of abasic site bypass by approximately 1.6-fold. In contrast, deletion of the N-terminal domains of Polλ did not affect the efficiency of 8-oxodG bypass relative to nucleotide incorporations opposite undamaged dG. HT-SOSA analysis demonstrated that Polλ and Polβ preferentially generated −1 or −2 frameshift mutations when bypassing an abasic site and the single or double base deletion frequency was highly sequence dependent. Interestingly, the BRCT and proline-rich domains of Polλ cooperatively promoted the generation of −2 frameshift mutations when the abasic site was situated within a sequence context that was susceptible to homology-driven primer realignment. Furthermore, both N-terminal domains of Polλ increased the generation of −1 frameshift mutations during 8-oxodG bypass and influenced the frequency of substitution mutations produced by Polλ opposite the 8-oxodG lesion. Overall, our data support a model wherein the BRCT and proline-rich domains of Polλ act cooperatively to promote primer/template realignment between DNA strands of limited sequence homology. This function of the N-terminal domains may facilitate the role of Polλ as a gap-filling polymerase

  19. Conserved chromosomal positions of dual domains of the ets protooncogene in cats, mice, and humans

    International Nuclear Information System (INIS)

    Watson, D.K.; McWilliams-Smith, M.J.; Kozak, C.

    1986-01-01

    The mammalian protooncogene homologue of the avian v-ets sequence from the E26 retrovirus consists of two sequentially distinct domains located on different chromosomes. Using somatic cell hybrid panels, the authors have mapped the mammalian homologue of the 5' v-ets-domain to chromosome 11 (ETS1) in man, to chromosome 9 (ets-1) in mouse, and to chromosome D1 (ETS1) in the domestic cat. The mammalian homologue of the 3' v-ets domain was similarly mapped to human chromosome 21 (ETS2), to mouse chromosome 16 (Ets-2), and to feline chromosome C2 (ETS2). Both protooncogenes fell in syntenic groups of homologous linked loci that were conserved among the three species. The occurrence of two distinct functional protooncogenes and their conservation of linkage positions in the three mammalian orders indicate that these two genes have been separate since before the evolutionary divergence of mammals

  20. Interactive lung segmentation in abnormal human and animal chest CT scans

    International Nuclear Information System (INIS)

    Kockelkorn, Thessa T. J. P.; Viergever, Max A.; Schaefer-Prokop, Cornelia M.; Bozovic, Gracijela; Muñoz-Barrutia, Arrate; Rikxoort, Eva M. van; Brown, Matthew S.; Jong, Pim A. de; Ginneken, Bram van

    2014-01-01

    Purpose: Many medical image analysis systems require segmentation of the structures of interest as a first step. For scans with gross pathology, automatic segmentation methods may fail. The authors’ aim is to develop a versatile, fast, and reliable interactive system to segment anatomical structures. In this study, this system was used for segmenting lungs in challenging thoracic computed tomography (CT) scans. Methods: In volumetric thoracic CT scans, the chest is segmented and divided into 3D volumes of interest (VOIs), containing voxels with similar densities. These VOIs are automatically labeled as either lung tissue or nonlung tissue. The automatic labeling results can be corrected using an interactive or a supervised interactive approach. When using the supervised interactive system, the user is shown the classification results per slice, whereupon he/she can adjust incorrect labels. The system is retrained continuously, taking the corrections and approvals of the user into account. In this way, the system learns to make a better distinction between lung tissue and nonlung tissue. When using the interactive framework without supervised learning, the user corrects all incorrectly labeled VOIs manually. Both interactive segmentation tools were tested on 32 volumetric CT scans of pigs, mice and humans, containing pulmonary abnormalities. Results: On average, supervised interactive lung segmentation took under 9 min of user interaction. Algorithm computing time was 2 min on average, but can easily be reduced. On average, 2.0% of all VOIs in a scan had to be relabeled. Lung segmentation using the interactive segmentation method took on average 13 min and involved relabeling 3.0% of all VOIs on average. The resulting segmentations correspond well to manual delineations of eight axial slices per scan, with an average Dice similarity coefficient of 0.933. Conclusions: The authors have developed two fast and reliable methods for interactive lung segmentation in

  1. Conservation of the human integrin-type beta-propeller domain in bacteria.

    Directory of Open Access Journals (Sweden)

    Bhanupratap Chouhan

    Full Text Available Integrins are heterodimeric cell-surface receptors with key functions in cell-cell and cell-matrix adhesion. Integrin α and β subunits are present throughout the metazoans, but it is unclear whether the subunits predate the origin of multicellular organisms. Several component domains have been detected in bacteria, one of which, a specific 7-bladed β-propeller domain, is a unique feature of the integrin α subunits. Here, we describe a structure-derived motif, which incorporates key features of each blade from the X-ray structures of human αIIbβ3 and αVβ3, includes elements of the FG-GAP/Cage and Ca(2+-binding motifs, and is specific only for the metazoan integrin domains. Separately, we searched for the metazoan integrin type β-propeller domains among all available sequences from bacteria and unicellular eukaryotic organisms, which must incorporate seven repeats, corresponding to the seven blades of the β-propeller domain, and so that the newly found structure-derived motif would exist in every repeat. As the result, among 47 available genomes of unicellular eukaryotes we could not find a single instance of seven repeats with the motif. Several sequences contained three repeats, a predicted transmembrane segment, and a short cytoplasmic motif associated with some integrins, but otherwise differ from the metazoan integrin α subunits. Among the available bacterial sequences, we found five examples containing seven sequential metazoan integrin-specific motifs within the seven repeats. The motifs differ in having one Ca(2+-binding site per repeat, whereas metazoan integrins have three or four sites. The bacterial sequences are more conserved in terms of motif conservation and loop length, suggesting that the structure is more regular and compact than those example structures from human integrins. Although the bacterial examples are not full-length integrins, the full-length metazoan-type 7-bladed β-propeller domains are present, and

  2. Functional conservation of the hydrophobic domain of polypeptide 3AB between human rhinovirus and poliovirus

    International Nuclear Information System (INIS)

    Towner, Jonathan S.; Brown, David M.; Nguyen, Joseph H.C.; Semler, Bert L.

    2003-01-01

    In this study we exchanged portions of the poliovirus type 1 (PV1) hydrophobic domain within the membrane-associated polypeptide 3AB for the analogous sequences from human rhinovirus 14 (HRV14). The sequence exchanges were based upon a previous report in which the 22 amino acid hydrophobic region was subdivided into two domains, I and II, the latter of which was shown to be required for membrane association (J. Biol. Chem. 271 (1996), 26810). Using these divisions, the HRV14 sequences were cloned into the complete poliovirus type 1 cDNA sequence. RNAs transcribed from these cDNAs were transfected into HeLa cell monolayers and used in HeLa cell-free translation/replication assays. The data indicated that 3AB sequences from PV1 and HRV14 are interchangeable; however, the substitutions cause a range of significant RNA replication defects, and in some cases, protein processing defects. Following transfection of RNAs encoding the domain substitutions into HeLa cell monolayers, virus isolates were harvested, and the corresponding viral RNAs were sequenced. The sequence data revealed that for the carboxy-terminal domain substitutions (domain II), multiple nucleotide changes were identified in the first, second, and third positions of different codons. In addition, the data indicated that for one of the PV1/HRV14 chimeras to replicate, compensatory mutations within poliovirus protein 2B may be required

  3. Human alpha2-macroglobulin is composed of multiple domains, as predicted by homology with complement component C3.

    Science.gov (United States)

    Doan, Ninh; Gettins, Peter G W

    2007-10-01

    Human alpha2M (alpha2-macroglobulin) and the complement components C3 and C4 are thiol ester-containing proteins that evolved from the same ancestral gene. The recent structure determination of human C3 has allowed a detailed prediction of the location of domains within human alpha2M to be made. We describe here the expression and characterization of three alpha(2)M domains predicted to be involved in the stabilization of the thiol ester in native alpha2M and in its activation upon bait region proteolysis. The three newly expressed domains are MG2 (macroglobulin domain 2), TED (thiol ester-containing domain) and CUB (complement protein subcomponents C1r/C1s, urchin embryonic growth factor and bone morphogenetic protein 1) domain. Together with the previously characterized RBD (receptor-binding domain), they represent approx. 42% of the alpha2M polypeptide. Their expression as folded domains strongly supports the predicted domain organization of alpha2M. An X-ray crystal structure of MG2 shows it to have a fibronectin type-3 fold analogous to MG1-MG8 of C3. TED is, as predicted, an alpha-helical domain. CUB is a spliced domain composed of two stretches of polypeptide that flank TED in the primary structure. In intact C3 TED interacts with RBD, where it is in direct contact with the thiol ester, and with MG2 and CUB on opposite, flanking sides. In contrast, these alpha2M domains, as isolated species, show negligible interaction with one another, suggesting that the native conformation of alpha2M, and the consequent thiol ester-stabilizing domain-domain interactions, result from additional restraints imposed by the physical linkage of these domains or by additional domains in the protein.

  4. Crystallization and preliminary X-ray analysis of the V domain of human nectin-2

    International Nuclear Information System (INIS)

    Qian, Xiaomin; Qi, Jianxun; Chu, Fuliang; Liu, Jun; Li, Qing; Yan, Jinghua

    2009-01-01

    Crystals of the V domain of human nectin-2 diffracted to 1.85 Å resolution and were monoclinic, belonging to space group P2 1 , with unit-cell parameters a = 52.3, b = 43.9, c = 56.1 Å, β = 118.2°. Nectin-2 belongs to a family of immunoglobulin-like cell adhesion molecules that are characterized by the presence of three immunoglobulin-like domains (V, C2 and C2) in the extracellular region. The V domain plays important roles in cell adhesion, NK cell activation and the entry of some herpesvirus. In this study, the V domain of human nectin-2 was expressed in Escherichia coli in the form of inclusion bodies, which were subsequently denatured and refolded. The soluble protein was crystallized using the hanging-drop vapour-diffusion method. The crystals diffracted to 1.85 Å resolution and belonged to space group P2 1 , with unit-cell parameters a = 52.3, b = 43.9, c = 56.1 Å, β = 118.2°

  5. Selective radiolabeling and isolation of the hydrophobic membrane-binding domain of human erythrocyte acetylcholinesterase

    International Nuclear Information System (INIS)

    Roberts, W.L.; Rosenberry, T.L.

    1986-01-01

    The hydrophobic, membrane-binding domain of purified human erythrocyte acetylcholinesterase was labeled with the photoactivated reagent 3-(trifluoromethyl)-3-(m-[ 125 I]iodophenyl)diazirine. The radiolabel was incorporated when the enzyme was prepared in detergent-free aggregates, in detergent micelles, or in phospholipid liposomes, but the highest percentage of labeling occurred in the detergent-free aggregates. Papain digestion of the enzyme released the hydrophobic domain, and polyacrylamide gel electrophoresis in sodium dodecyl sulfate or gel exclusion chromatography demonstrated that the label was localized exclusively in the cleaved hydrophobic domain fragment. This fragment was purified in a three-step procedure. Digestion was conducted with papain attached to Sepharose CL-4B, and the supernatant was adsorbed to acridinium affinity resin to remove the hydrophilic enzyme fragment. The nonretained fragment associated with Triton X-100 micelles was then chromatographed on Sepharose CL-6B, and finally detergent was removed by chromatography on Sephadex LH-60 in an ethanol-formic acid solvent. The fragment exhibited an apparent molecular weight of 3100 on the Sephadex LH-60 column when compared with peptide standards. However, amino acid analysis of the purified fragment revealed only 1 mol each of histidine and glycine per mole of fragment in contrast to the 25-30 mole of amino acids expected on the basis of the molecular weight estimate. This result suggests a novel non-amino acid structure for the hydrophobic domain of human erythrocyte acetylcholinesterase

  6. Methodologies for Development of Patient Specific Bone Models from Human Body CT Scans

    Science.gov (United States)

    Chougule, Vikas Narayan; Mulay, Arati Vinayak; Ahuja, Bharatkumar Bhagatraj

    2016-06-01

    This work deals with development of algorithm for physical replication of patient specific human bone and construction of corresponding implants/inserts RP models by using Reverse Engineering approach from non-invasive medical images for surgical purpose. In medical field, the volumetric data i.e. voxel and triangular facet based models are primarily used for bio-modelling and visualization, which requires huge memory space. On the other side, recent advances in Computer Aided Design (CAD) technology provides additional facilities/functions for design, prototyping and manufacturing of any object having freeform surfaces based on boundary representation techniques. This work presents a process to physical replication of 3D rapid prototyping (RP) physical models of human bone from various CAD modeling techniques developed by using 3D point cloud data which is obtained from non-invasive CT/MRI scans in DICOM 3.0 format. This point cloud data is used for construction of 3D CAD model by fitting B-spline curves through these points and then fitting surface between these curve networks by using swept blend techniques. This process also can be achieved by generating the triangular mesh directly from 3D point cloud data without developing any surface model using any commercial CAD software. The generated STL file from 3D point cloud data is used as a basic input for RP process. The Delaunay tetrahedralization approach is used to process the 3D point cloud data to obtain STL file. CT scan data of Metacarpus (human bone) is used as the case study for the generation of the 3D RP model. A 3D physical model of the human bone is generated on rapid prototyping machine and its virtual reality model is presented for visualization. The generated CAD model by different techniques is compared for the accuracy and reliability. The results of this research work are assessed for clinical reliability in replication of human bone in medical field.

  7. Micro-CT characterization of human trabecular bone in osteogenesis imperfecta

    Science.gov (United States)

    Jameson, John; Albert, Carolyne; Smith, Peter; Molthen, Robert; Harris, Gerald

    2011-03-01

    Osteogenesis imperfecta (OI) is a genetic syndrome affecting collagen synthesis and assembly. Its symptoms vary widely but commonly include bone fragility, reduced stature, and bone deformity. Because of the small size and paucity of human specimens, there is a lack of biomechanical data for OI bone. Most literature has focused on histomorphometric analyses, which rely on assumptions to extrapolate 3-D properties. In this study, a micro-computed tomography (μCT) system was used to directly measure structural and mineral properties in pediatric OI bone collected during routine surgical procedures. Surface renderings suggested a poorly organized, plate-like orientation. Patients with a history of bone-augmenting drugs exhibited increased bone volume fraction (BV/TV), trabecular number (Tb.N), and connectivity density (Eu.Conn.D). The latter two parameters appeared to be related to OI severity. Structural results were consistently higher than those reported in a previous histomorphometric study, but these differences can be attributed to factors such as specimen collection site, drug therapy, and assumptions associated with histomorphometry. Mineral testing revealed strong correlations with several structural parameters, highlighting the importance of a dual approach in trabecular bone testing. This study reports some of the first quantitative μCT data of human OI bone, and it suggests compelling possibilities for the future of OI bone assessment.

  8. Human α2-macroglobulin is composed of multiple domains, as predicted by homology with complement component C3

    Science.gov (United States)

    Doan, Ninh; Gettins, Peter G. W.

    2007-01-01

    Human α2M (α2-macroglobulin) and the complement components C3 and C4 are thiol ester-containing proteins that evolved from the same ancestral gene. The recent structure determination of human C3 has allowed a detailed prediction of the location of domains within human α2M to be made. We describe here the expression and characterization of three α2M domains predicted to be involved in the stabilization of the thiol ester in native α2M and in its activation upon bait region proteolysis. The three newly expressed domains are MG2 (macroglobulin domain 2), TED (thiol ester-containing domain) and CUB (complement protein subcomponents C1r/C1s, urchin embryonic growth factor and bone morphogenetic protein 1) domain. Together with the previously characterized RBD (receptor-binding domain), they represent approx. 42% of the α2M polypeptide. Their expression as folded domains strongly supports the predicted domain organization of α2M. An X-ray crystal structure of MG2 shows it to have a fibronectin type-3 fold analogous to MG1–MG8 of C3. TED is, as predicted, an α-helical domain. CUB is a spliced domain composed of two stretches of polypeptide that flank TED in the primary structure. In intact C3 TED interacts with RBD, where it is in direct contact with the thiol ester, and with MG2 and CUB on opposite, flanking sides. In contrast, these α2M domains, as isolated species, show negligible interaction with one another, suggesting that the native conformation of α2M, and the consequent thiol ester-stabilizing domain–domain interactions, result from additional restraints imposed by the physical linkage of these domains or by additional domains in the protein. PMID:17608619

  9. Nonlinear time-domain cochlear model for transient stimulation and human otoacoustic emission

    DEFF Research Database (Denmark)

    Verhulst, Sarah; Dau, Torsten; Shera, Christopher A.

    2012-01-01

    This paper describes the implementation and performance of a nonlinear time-domain model of the cochlea for transient stimulation and human otoacoustic emission generation. The nonlinearity simulates compressive growth of measured basilar-membrane impulse responses. The model accounts...... for reflection and distortion-source otoacoustic emissions (OAEs) and simulates spontaneous OAEs through manipulation of the middle-ear reflectance. The model was calibrated using human psychoacoustical and otoacoustic tuning parameters. It can be used to investigate time-dependent properties of cochlear...

  10. Clinical evaluation of lumbar CT assisted discography in comparison with human cadaver

    Energy Technology Data Exchange (ETDEWEB)

    Ito, Shigehiko

    1988-04-01

    In order to estimate the clinical value of lumbar CT assisted discography (CTD), results obtained by this method were compared with histological findings of the cross section of the spine in fresh human cadavera. Based on these findings, preoperative CTD of lumbar disc herniation was investigated. In the discs of human cadavera, the contrast medium mainly invaded the fissures of nucleus pulposus and the ruptures of annulus fibrosus and then diffused to the surrounding tissues. These ruptures were classified into two categories: radial and circumferential. This indentification was possible only in CTD and was obscure in the usual discogram. Not all the ruptures could be dyed in a severe degenerative disc, and a rupture which was not communicated with nucleus pulposus was not dyed in a mild degenerative disc. In the preoperative CTD of lumbar disc herniation, the posterior radial ruptures representing the route of herniated nuclei were characteristic and the circumferential ruptures were found complicated.

  11. Structures of BIR domains from human NAIP and cIAP2

    International Nuclear Information System (INIS)

    Herman, Maria Dolores; Moche, Martin; Flodin, Susanne; Welin, Martin; Trésaugues, Lionel; Johansson, Ida; Nilsson, Martina; Nordlund, Pär; Nyman, Tomas

    2009-01-01

    The crystal structures of the human NAIP BIR2 and cIAP2 BIR3 domains have been determined. Both BIR domains harbors an amino-terminal tetrapeptide in its peptide-binding groove. The inhibitor of apoptosis (IAP) family of proteins contains key modulators of apoptosis and inflammation that interact with caspases through baculovirus IAP-repeat (BIR) domains. Overexpression of IAP proteins frequently occurs in cancer cells, thus counteracting the activated apoptotic program. The IAP proteins have therefore emerged as promising targets for cancer therapy. In this work, X-ray crystallography was used to determine the first structures of BIR domains from human NAIP and cIAP2. Both structures harbour an N-terminal tetrapeptide in the conserved peptide-binding groove. The structures reveal that these two proteins bind the tetrapeptides in a similar mode as do other BIR domains. Detailed interactions are described for the P1′–P4′ side chains of the peptide, providing a structural basis for peptide-specific recognition. An arginine side chain in the P3′ position reveals favourable interactions with its hydrophobic moiety in the binding pocket, while hydrophobic residues in the P2′ and P4′ pockets make similar interactions to those seen in other BIR domain–peptide complexes. The structures also reveal how a serine in the P1′ position is accommodated in the binding pockets of NAIP and cIAP2. In addition to shedding light on the specificity determinants of these two proteins, the structures should now also provide a framework for future structure-based work targeting these proteins

  12. The effects of thyroid hormones on brown adipose tissue in humans: a PET-CT study.

    Science.gov (United States)

    Zhang, Qiongyue; Miao, Qing; Ye, Hongying; Zhang, Zhaoyun; Zuo, Chuantao; Hua, Fengchun; Guan, Yihui; Li, Yiming

    2014-09-01

    Brown adipose tissue (BAT) is important for energy expenditure through thermogenesis, although its regulatory factors are not well known in humans. There is evidence suggesting that thyroid hormones affect BAT functions in some mammals, but the effects of thyroid hormones on BAT activity in humans are still unclear. The aim of this study was to investigate the effects of thyroid hormones on glucose metabolism of BAT and other organs in humans. Nine Graves' disease-caused hyperthyroid patients who were newly diagnosed and untreated were studied. Putative brown adipose tissue activity was determined by the integrated ¹⁸F-fluorodeoxyglucose (¹⁸F-FDG) positron-emission tomography and computed tomography (PET-CT). All hyperthyroid patients were treated with methimazole and had been monitored until their symptoms disappeared and thyroid hormone levels returned to normal. At the end, a second PET-CT scan was performed. The average follow-up period was 77 days. Meanwhile, compared with a group of seventy-five brown adipose tissue-negative controls, thyroid hormones of seventy-five BAT-positive healthy subjects were measured. Active brown adipose tissue was not present in any of the hyperthyroid patients. However, one patient with normalized thyroid function showed active BAT after therapy. The free T3 levels and free T4 levels were significantly lower in the 75 BAT-positive subjects than in the BAT-negative subjects. All hyperthyroid patients showed symmetrically increased uptake of fluorodeoxyglucose in skeletal muscles before treatment, whereas, the standardized uptake value was substantially decreased after treatment. Abnormally high circulating thyroid hormone levels may not increase brown adipose tissue activity, which may be limited by the increased obligatory thermogenesis of muscle in adult humans. Copyright © 2014 John Wiley & Sons, Ltd.

  13. How does domain replacement affect fibril formation of the rabbit/human prion proteins.

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    Xu Yan

    Full Text Available It is known that in vivo human prion protein (PrP have the tendency to form fibril deposits and are associated with infectious fatal prion diseases, while the rabbit PrP does not readily form fibrils and is unlikely to cause prion diseases. Although we have previously demonstrated that amyloid fibrils formed by the rabbit PrP and the human PrP have different secondary structures and macromolecular crowding has different effects on fibril formation of the rabbit/human PrPs, we do not know which domains of PrPs cause such differences. In this study, we have constructed two PrP chimeras, rabbit chimera and human chimera, and investigated how domain replacement affects fibril formation of the rabbit/human PrPs.As revealed by thioflavin T binding assays and Sarkosyl-soluble SDS-PAGE, the presence of a strong crowding agent dramatically promotes fibril formation of both chimeras. As evidenced by circular dichroism, Fourier transform infrared spectroscopy, and proteinase K digestion assays, amyloid fibrils formed by human chimera have secondary structures and proteinase K-resistant features similar to those formed by the human PrP. However, amyloid fibrils formed by rabbit chimera have proteinase K-resistant features and secondary structures in crowded physiological environments different from those formed by the rabbit PrP, and secondary structures in dilute solutions similar to the rabbit PrP. The results from transmission electron microscopy show that macromolecular crowding caused human chimera but not rabbit chimera to form short fibrils and non-fibrillar particles.We demonstrate for the first time that the domains beyond PrP-H2H3 (β-strand 1, α-helix 1, and β-strand 2 have a remarkable effect on fibrillization of the rabbit PrP but almost no effect on the human PrP. Our findings can help to explain why amyloid fibrils formed by the rabbit PrP and the human PrP have different secondary structures and why macromolecular crowding has different

  14. Structures of BIR domains from human NAIP and cIAP2.

    Science.gov (United States)

    Herman, Maria Dolores; Moche, Martin; Flodin, Susanne; Welin, Martin; Trésaugues, Lionel; Johansson, Ida; Nilsson, Martina; Nordlund, Pär; Nyman, Tomas

    2009-11-01

    The inhibitor of apoptosis (IAP) family of proteins contains key modulators of apoptosis and inflammation that interact with caspases through baculovirus IAP-repeat (BIR) domains. Overexpression of IAP proteins frequently occurs in cancer cells, thus counteracting the activated apoptotic program. The IAP proteins have therefore emerged as promising targets for cancer therapy. In this work, X-ray crystallography was used to determine the first structures of BIR domains from human NAIP and cIAP2. Both structures harbour an N-terminal tetrapeptide in the conserved peptide-binding groove. The structures reveal that these two proteins bind the tetrapeptides in a similar mode as do other BIR domains. Detailed interactions are described for the P1'-P4' side chains of the peptide, providing a structural basis for peptide-specific recognition. An arginine side chain in the P3' position reveals favourable interactions with its hydrophobic moiety in the binding pocket, while hydrophobic residues in the P2' and P4' pockets make similar interactions to those seen in other BIR domain-peptide complexes. The structures also reveal how a serine in the P1' position is accommodated in the binding pockets of NAIP and cIAP2. In addition to shedding light on the specificity determinants of these two proteins, the structures should now also provide a framework for future structure-based work targeting these proteins.

  15. Fast identification of folded human protein domains expressed in E. coli suitable for structural analysis

    Directory of Open Access Journals (Sweden)

    Schlegel Brigitte

    2004-03-01

    Full Text Available Abstract Background High-throughput protein structure analysis of individual protein domains requires analysis of large numbers of expression clones to identify suitable constructs for structure determination. For this purpose, methods need to be implemented for fast and reliable screening of the expressed proteins as early as possible in the overall process from cloning to structure determination. Results 88 different E. coli expression constructs for 17 human protein domains were analysed using high-throughput cloning, purification and folding analysis to obtain candidates suitable for structural analysis. After 96 deep-well microplate expression and automated protein purification, protein domains were directly analysed using 1D 1H-NMR spectroscopy. In addition, analytical hydrophobic interaction chromatography (HIC was used to detect natively folded protein. With these two analytical methods, six constructs (representing two domains were quickly identified as being well folded and suitable for structural analysis. Conclusion The described approach facilitates high-throughput structural analysis. Clones expressing natively folded proteins suitable for NMR structure determination were quickly identified upon small scale expression screening using 1D 1H-NMR and/or analytical HIC. This procedure is especially effective as a fast and inexpensive screen for the 'low hanging fruits' in structural genomics.

  16. Crystal Structure of the Human, FIC-Domain Containing Protein HYPE and Implications for Its Functions

    Science.gov (United States)

    Bunney, Tom D.; Cole, Ambrose R.; Broncel, Malgorzata; Esposito, Diego; Tate, Edward W.; Katan, Matilda

    2014-01-01

    Summary Protein AMPylation, the transfer of AMP from ATP to protein targets, has been recognized as a new mechanism of host-cell disruption by some bacterial effectors that typically contain a FIC-domain. Eukaryotic genomes also encode one FIC-domain protein, HYPE, which has remained poorly characterized. Here we describe the structure of human HYPE, solved by X-ray crystallography, representing the first structure of a eukaryotic FIC-domain protein. We demonstrate that HYPE forms stable dimers with structurally and functionally integrated FIC-domains and with TPR-motifs exposed for protein-protein interactions. As HYPE also uniquely possesses a transmembrane helix, dimerization is likely to affect its positioning and function in the membrane vicinity. The low rate of autoAMPylation of the wild-type HYPE could be due to autoinhibition, consistent with the mechanism proposed for a number of putative FIC AMPylators. Our findings also provide a basis to further consider possible alternative cofactors of HYPE and distinct modes of target-recognition. PMID:25435325

  17. Significant Suppression of CT Radiation-Induced DNA Damage in Normal Human Cells by the PrC-210 Radioprotector.

    Science.gov (United States)

    Jermusek, Frank; Benedict, Chelsea; Dreischmeier, Emma; Brand, Michael; Uder, Michael; Jeffery, Justin J; Ranallo, Frank N; Fahl, William E

    2018-05-21

    While computed tomography (CT) is now commonly used and considered to be clinically valuable, significant DNA double-strand breaks (γ-H2AX foci) in white blood cells from adult and pediatric CT patients have been frequently reported. In this study to determine whether γ-H2AX foci and X-ray-induced naked DNA damage are suppressed by administration of the PrC-210 radioprotector, human blood samples were irradiated in a CT scanner at 50-150 mGy with or without PrC-210, and γ-H2AX foci were scored. X-ray-induced naked DNA damage was also studied, and the DNA protective efficacy of PrC-210 was compared against 12 other common "antioxidants." PrC-210 reduced CT radiation-induced γ-H2AX foci in white blood cells to near background ( P 95% DNA damage. A systemic PrC-210 dose known to confer 100% survival in irradiated mice had no discernible effect on micro-CT image signal-to-noise ratio and CT image integrity. PrC-210 suppressed DNA damage to background or near background in each of these assay systems, thus supporting its development as a radioprotector for humans in multiple radiation exposure settings.

  18. Herniation pits in human mummies: a CT investigation in the Capuchin Catacombs of Palermo, Sicily.

    Directory of Open Access Journals (Sweden)

    Stephanie Panzer

    Full Text Available Herniation pits (HPs of the femoral neck were first described in a radiological publication in 1982 as round to oval radiolucencies in the proximal superior quadrant of the femoral neck on anteroposterior radiographs of adults. In following early clinical publications, HPs were generally recognized as an incidental finding. In contrast, in current clinical literature they are mentioned in the context of femoroacetabular impingement (FAI of the hip joint, which is known to cause osteoarthritis (OA. The significance of HPs in chronic skeletal disorders such as OA is still unclear, but they are discussed as a possible radiological indicator for FAI in a large part of clinical studies.In this paleoradiological study we examined a sample of mummies from the Capuchin Catacombs of Palermo, Sicily, by a mobile computed tomography (CT scanner. Evaluation of the CT examinations revealed HPs in six out of 16 (37.5% adult male mummies.The first aim of this study was to compare the characteristics of HPs shown in our mummy collection to the findings described in clinical literature. Thereby CT evaluation revealed that their osseous imaging characteristics are in accordance, consisting of round to oval subcortical lesions at the anterior femoral neck, clearly demarcated by a sclerotic margin.The second aim was to introduce HPs to the paleoradiological and paleopathological methodology as an entity that underwent a renaissance from an incidental finding to a possible radiological indicator of FAI in the clinical situation. As FAI plays an important role in the development of OA of the hip, which is a very common finding in human skeletal remains, HPs should always be considered in paleoradiological evaluation of hip joint diseases.

  19. Fully automatic algorithm for segmenting full human diaphragm in non-contrast CT Images

    Science.gov (United States)

    Karami, Elham; Gaede, Stewart; Lee, Ting-Yim; Samani, Abbas

    2015-03-01

    The diaphragm is a sheet of muscle which separates the thorax from the abdomen and it acts as the most important muscle of the respiratory system. As such, an accurate segmentation of the diaphragm, not only provides key information for functional analysis of the respiratory system, but also can be used for locating other abdominal organs such as the liver. However, diaphragm segmentation is extremely challenging in non-contrast CT images due to the diaphragm's similar appearance to other abdominal organs. In this paper, we present a fully automatic algorithm for diaphragm segmentation in non-contrast CT images. The method is mainly based on a priori knowledge about the human diaphragm anatomy. The diaphragm domes are in contact with the lungs and the heart while its circumference runs along the lumbar vertebrae of the spine as well as the inferior border of the ribs and sternum. As such, the diaphragm can be delineated by segmentation of these organs followed by connecting relevant parts of their outline properly. More specifically, the bottom surface of the lungs and heart, the spine borders and the ribs are delineated, leading to a set of scattered points which represent the diaphragm's geometry. Next, a B-spline filter is used to find the smoothest surface which pass through these points. This algorithm was tested on a noncontrast CT image of a lung cancer patient. The results indicate that there is an average Hausdorff distance of 2.96 mm between the automatic and manually segmented diaphragms which implies a favourable accuracy.

  20. Mapping cis-Regulatory Domains in the Human Genome UsingMulti-Species Conservation of Synteny

    Energy Technology Data Exchange (ETDEWEB)

    Ahituv, Nadav; Prabhakar, Shyam; Poulin, Francis; Rubin, EdwardM.; Couronne, Olivier

    2005-06-13

    Our inability to associate distant regulatory elements with the genes that they regulate has largely precluded their examination for sequence alterations contributing to human disease. One major obstacle is the large genomic space surrounding targeted genes in which such elements could potentially reside. In order to delineate gene regulatory boundaries we used whole-genome human-mouse-chicken (HMC) and human-mouse-frog (HMF) multiple alignments to compile conserved blocks of synteny (CBS), under the hypothesis that these blocks have been kept intact throughout evolution at least in part by the requirement of regulatory elements to stay linked to the genes that they regulate. A total of 2,116 and 1,942 CBS>200 kb were assembled for HMC and HMF respectively, encompassing 1.53 and 0.86 Gb of human sequence. To support the existence of complex long-range regulatory domains within these CBS we analyzed the prevalence and distribution of chromosomal aberrations leading to position effects (disruption of a genes regulatory environment), observing a clear bias not only for mapping onto CBS but also for longer CBS size. Our results provide a genome wide data set characterizing the regulatory domains of genes and the conserved regulatory elements within them.

  1. Swapping the N- and C-terminal domains of human apolipoprotein E3 and AI reveals insights into their structure/activity relationship.

    Directory of Open Access Journals (Sweden)

    Mark T Lek

    Full Text Available Apolipoprotein (apo E3 and apoAI are exchangeable apolipoproteins that play a dominant role in regulating plasma lipoprotein metabolism. ApoE3 (299 residues is composed of an N-terminal (NT domain bearing a 4-helix bundle and a C-terminal (CT domain bearing a series of amphipathic α-helices. ApoAI (243 residues also comprises a highly helical NT domain and a less structured CT tail. The objective of this study was to understand their structural and functional role by generating domain swapped chimeras: apoE3-NT/apoAI-CT and apoAI-NT/apoE-CT. The bacterially overexpressed chimeras were purified by affinity chromatography and their identity confirmed by immunoblotting and mass spectrometry. Their α-helical content was comparable to that of the parent proteins. ApoE3-NT/apoAI-CT retained the denaturation profile of apoE3 NT domain, with apoAI CT tail eliciting a relatively unstructured state; its lipid binding ability improved dramatically compared to apoE3 indicative of a significant role of apoAI CT tail in lipid binding interaction. The LDL receptor interaction and ability to promote ABCA1-mediated cholesterol efflux of apoE3-NT/apoAI-CT was comparable to that of apoE3. In contrast, apoAI-NT/apoE-CT elicited an unfolding pattern and lipid binding ability that were similar to that of apoAI. As expected, DMPC/apoAI-NT/apoE-CT discoidal particles did not elicit LDLr binding ability, and promoted SR-B1 mediated cellular uptake of lipids to a limited extent. However, apoAI-NT/apoE-CT displayed an enhanced ability to promote cholesterol efflux compared to apoAI, indicative of a significant role for apoE CT domain in mediating this function. Together, these results indicate that the functional attributes of apoAI and apoE3 can be conferred on each other and that NT-CT domain interactions significantly modulate their structure and function.

  2. The Ecology of Human-Machine Systems II: Mediating 'Direct Perception' in Complex Work Domains

    DEFF Research Database (Denmark)

    Vicente, Kim J.; Rasmussen, Jens

    1990-01-01

    Recently, a new class of artifacts has appeared in our environment: complex, high-technology work domains. An important characteristic of such systems is that their goal-relevant properties cannot be directly observed by the unaided eye. As a result, interface design is a ubiquitous problem in th...... in the design of these work environments. Nevertheless, the problem is one that has yet to be addressed in an adequate manner. An analogy to human perceptual mechanisms suggests that a smart instrument approach to interface design is needed to supplant the rote instrument (single......-sensor-single-indicator) approach that has dominated to this point. Ecological interface design (ED) is a theoretical framework in the smart instrument vein that postulates a set of general, prescriptive principles for design. The goal of E D is twofold: first, to reveal the affordances of the work domain through the interface...

  3. Wavelet-domain de-noising of OCT images of human brain malignant glioma

    Science.gov (United States)

    Dolganova, I. N.; Aleksandrova, P. V.; Beshplav, S.-I. T.; Chernomyrdin, N. V.; Dubyanskaya, E. N.; Goryaynov, S. A.; Kurlov, V. N.; Reshetov, I. V.; Potapov, A. A.; Tuchin, V. V.; Zaytsev, K. I.

    2018-04-01

    We have proposed a wavelet-domain de-noising technique for imaging of human brain malignant glioma by optical coherence tomography (OCT). It implies OCT image decomposition using the direct fast wavelet transform, thresholding of the obtained wavelet spectrum and further inverse fast wavelet transform for image reconstruction. By selecting both wavelet basis and thresholding procedure, we have found an optimal wavelet filter, which application improves differentiation of the considered brain tissue classes - i.e. malignant glioma and normal/intact tissue. Namely, it allows reducing the scattering noise in the OCT images and retaining signal decrement for each tissue class. Therefore, the observed results reveals the wavelet-domain de-noising as a prospective tool for improved characterization of biological tissue using the OCT.

  4. Overlapping positive and negative regulatory domains of the human β-interferon gene

    International Nuclear Information System (INIS)

    Goodbourn, S.; Maniatis, T.

    1988-01-01

    Virus of poly(I) x poly(C) induction of human β-interferon gene expression requires a 40-base-pair DNA sequence designated the interferon gene regulatory element (IRE). Previous studies have shown that the IRE contains both positive and negative regulatory DNA sequences. To localize these sequences and study their interactions, the authors have examined the effects of a large number of single-base mutations within the IRE on β-interferon gene regulation. They find that the IRE consists of two genetically separable positive regulatory domains and an overlapping negative control sequence. They propose that the β-interferon gene is switched off in uninduced cells by a repressor that blocks the interaction between one of the two positive regulatory sequences and a specific transcription factor. Induction would then lead to inactivation or displacement of the repressor and binding of transcription factors to both positive regulatory domains

  5. Transactivation domain of p53 regulates DNA repair and integrity in human iPS cells.

    Science.gov (United States)

    Kannappan, Ramaswamy; Mattapally, Saidulu; Wagle, Pooja A; Zhang, Jianyi

    2018-05-18

    The role of p53 transactivation domain (p53-TAD), a multifunctional and dynamic domain, on DNA repair and retaining DNA integrity in human iPS cells has never been studied. p53-TAD was knocked out in iPS cells using CRISPR/Cas9 and was confirmed by DNA sequencing. p53-TAD KO cells were characterized by: accelerated proliferation, decreased population doubling time, and unaltered Bcl2, BBC3, IGF1R, Bax and altered Mdm2, p21, and PIDD transcripts expression. In p53-TAD KO cells p53 regulated DNA repair proteins XPA, DNA polH and DDB2 expression were found to be reduced compared to p53-WT cells. Exposure to low dose of doxorubicin (Doxo) induced similar DNA damage and DNA damage response (DDR) measured by RAD50 and MRE11 expression, Checkpoint kinase 2 activation and γH2A.X recruitment at DNA strand breaks in both the cell groups indicating silencing p53-TAD do not affect DDR mechanism upstream of p53. Following removal of Doxo p53-WT hiPS cells underwent DNA repair, corrected their damaged DNA and restored DNA integrity. Conversely, p53-TAD KO hiPS cells did not undergo complete DNA repair and failed to restore DNA integrity. More importantly continuous culture of p53-TAD KO hiPS cells underwent G2/M cell cycle arrest and expressed cellular senescent marker p16 INK4a . Our data clearly shows that silencing transactivation domain of p53 did not affect DDR but affected the DNA repair process implying the crucial role of p53 transactivation domain in maintaining DNA integrity. Therefore, activating p53-TAD domain using small molecules may promote DNA repair and integrity of cells and prevent senescence.

  6. Comparison of human and automatic segmentations of kidneys from CT images

    International Nuclear Information System (INIS)

    Rao, Manjori; Stough, Joshua; Chi, Y.-Y.; Muller, Keith; Tracton, Gregg; Pizer, Stephen M.; Chaney, Edward L.

    2005-01-01

    Purpose: A controlled observer study was conducted to compare a method for automatic image segmentation with conventional user-guided segmentation of right and left kidneys from planning computerized tomographic (CT) images. Methods and materials: Deformable shape models called m-reps were used to automatically segment right and left kidneys from 12 target CT images, and the results were compared with careful manual segmentations performed by two human experts. M-rep models were trained based on manual segmentations from a collection of images that did not include the targets. Segmentation using m-reps began with interactive initialization to position the kidney model over the target kidney in the image data. Fully automatic segmentation proceeded through two stages at successively smaller spatial scales. At the first stage, a global similarity transformation of the kidney model was computed to position the model closer to the target kidney. The similarity transformation was followed by large-scale deformations based on principal geodesic analysis (PGA). During the second stage, the medial atoms comprising the m-rep model were deformed one by one. This procedure was iterated until no changes were observed. The transformations and deformations at both stages were driven by optimizing an objective function with two terms. One term penalized the currently deformed m-rep by an amount proportional to its deviation from the mean m-rep derived from PGA of the training segmentations. The second term computed a model-to-image match term based on the goodness of match of the trained intensity template for the currently deformed m-rep with the corresponding intensity data in the target image. Human and m-rep segmentations were compared using quantitative metrics provided in a toolset called Valmet. Metrics reported in this article include (1) percent volume overlap; (2) mean surface distance between two segmentations; and (3) maximum surface separation (Hausdorff distance

  7. Volumetric gain of the human pancreas after left partial pancreatic resection: A CT-scan based retrospective study.

    Science.gov (United States)

    Phillip, Veit; Zahel, Tina; Danninger, Assiye; Erkan, Mert; Dobritz, Martin; Steiner, Jörg M; Kleeff, Jörg; Schmid, Roland M; Algül, Hana

    2015-01-01

    Regeneration of the pancreas has been well characterized in animal models. However, there are conflicting data on the regenerative capacity of the human pancreas. The aim of the present study was to assess the regenerative capacity of the human pancreas. In a retrospective study, data from patients undergoing left partial pancreatic resection at a single center were eligible for inclusion (n = 185). Volumetry was performed based on 5 mm CT-scans acquired through a 256-slice CT-scanner using a semi-automated software. Data from 24 patients (15 males/9 females) were included. Mean ± SD age was 68 ± 11 years (range, 40-85 years). Median time between surgery and the 1st postoperative CT was 9 days (range, 0-27 days; IQR, 7-13), 55 days (range, 21-141 days; IQR, 34-105) until the 2nd CT, and 191 days (range, 62-1902; IQR, 156-347) until the 3rd CT. The pancreatic volumes differed significantly between the first and the second postoperative CT scans (median volume 25.6 mL and 30.6 mL, respectively; p = 0.008) and had significantly increased further by the 3rd CT scan (median volume 37.9 mL; p = 0.001 for comparison with 1st CT scan and p = 0.003 for comparison with 2nd CT scan). The human pancreas shows a measurable and considerable potential of volumetric gain after partial resection. Multidetector-CT based semi-automated volume analysis is a feasible method for follow-up of the volume of the remaining pancreatic parenchyma after partial pancreatectomy. Effects on exocrine and endocrine pancreatic function have to be evaluated in a prospective manner. Copyright © 2015 IAP and EPC. Published by Elsevier B.V. All rights reserved.

  8. Enhanced human somatic cell reprogramming efficiency by fusion of the MYC transactivation domain and OCT4

    Directory of Open Access Journals (Sweden)

    Ling Wang

    2017-12-01

    Full Text Available The development of human induced pluripotent stem cells (iPSCs holds great promise for regenerative medicine. However the iPSC induction efficiency is still very low and with lengthy reprogramming process. We utilized the highly potent transactivation domain (TAD of MYC protein to engineer the human OCT4 fusion proteins. Applying the MYC-TAD-OCT4 fusion proteins in mouse iPSC generation leads to shorter reprogramming dynamics, with earlier activation of pluripotent markers in reprogrammed cells than wild type OCT4 (wt-OCT4. Dramatic enhancement of iPSC colony induction efficiency and shortened reprogramming dynamics were observed when these MYC-TAD-OCT4 fusion proteins were used to reprogram primary human cells. The OCT4 fusion proteins induced human iPSCs are pluripotent. We further show that the MYC Box I (MBI is dispensable while both MBII and the linking region between MBI/II are essential for the enhanced reprogramming activity of MYC-TAD-OCT4 fusion protein. Consistent with an enhanced transcription activity, the engineered OCT4 significantly stimulated the expression of genes specifically targeted by OCT4-alone, OCT4/SOX2, and OCT4/SOX2/KLF4 during human iPSC induction, compared with the wt-OCT4. The MYC-TAD-OCT4 fusion proteins we generated will be valuable tools for studying the reprogramming mechanisms and for efficient iPSC generation for humans as well as for other species.

  9. Assembly of the membrane domain of ATP synthase in human mitochondria.

    Science.gov (United States)

    He, Jiuya; Ford, Holly C; Carroll, Joe; Douglas, Corsten; Gonzales, Evvia; Ding, Shujing; Fearnley, Ian M; Walker, John E

    2018-03-20

    The ATP synthase in human mitochondria is a membrane-bound assembly of 29 proteins of 18 kinds. All but two membrane components are encoded in nuclear genes, synthesized on cytoplasmic ribosomes, and imported into the matrix of the organelle, where they are assembled into the complex with ATP6 and ATP8, the products of overlapping genes in mitochondrial DNA. Disruption of individual human genes for the nuclear-encoded subunits in the membrane portion of the enzyme leads to the formation of intermediate vestigial ATPase complexes that provide a description of the pathway of assembly of the membrane domain. The key intermediate complex consists of the F 1 -c 8 complex inhibited by the ATPase inhibitor protein IF 1 and attached to the peripheral stalk, with subunits e, f, and g associated with the membrane domain of the peripheral stalk. This intermediate provides the template for insertion of ATP6 and ATP8, which are synthesized on mitochondrial ribosomes. Their association with the complex is stabilized by addition of the 6.8 proteolipid, and the complex is coupled to ATP synthesis at this point. A structure of the dimeric yeast F o membrane domain is consistent with this model of assembly. The human 6.8 proteolipid (yeast j subunit) locks ATP6 and ATP8 into the membrane assembly, and the monomeric complexes then dimerize via interactions between ATP6 subunits and between 6.8 proteolipids (j subunits). The dimers are linked together back-to-face by DAPIT (diabetes-associated protein in insulin-sensitive tissue; yeast subunit k), forming long oligomers along the edges of the cristae.

  10. Genome-Wide Prediction and Analysis of 3D-Domain Swapped Proteins in the Human Genome from Sequence Information.

    Science.gov (United States)

    Upadhyay, Atul Kumar; Sowdhamini, Ramanathan

    2016-01-01

    3D-domain swapping is one of the mechanisms of protein oligomerization and the proteins exhibiting this phenomenon have many biological functions. These proteins, which undergo domain swapping, have acquired much attention owing to their involvement in human diseases, such as conformational diseases, amyloidosis, serpinopathies, proteionopathies etc. Early realisation of proteins in the whole human genome that retain tendency to domain swap will enable many aspects of disease control management. Predictive models were developed by using machine learning approaches with an average accuracy of 78% (85.6% of sensitivity, 87.5% of specificity and an MCC value of 0.72) to predict putative domain swapping in protein sequences. These models were applied to many complete genomes with special emphasis on the human genome. Nearly 44% of the protein sequences in the human genome were predicted positive for domain swapping. Enrichment analysis was performed on the positively predicted sequences from human genome for their domain distribution, disease association and functional importance based on Gene Ontology (GO). Enrichment analysis was also performed to infer a better understanding of the functional importance of these sequences. Finally, we developed hinge region prediction, in the given putative domain swapped sequence, by using important physicochemical properties of amino acids.

  11. Gating of human ClC-2 chloride channels and regulation by carboxy-terminal domains.

    Science.gov (United States)

    Garcia-Olivares, Jennie; Alekov, Alexi; Boroumand, Mohammad Reza; Begemann, Birgit; Hidalgo, Patricia; Fahlke, Christoph

    2008-11-15

    Eukaryotic ClC channels are dimeric proteins with each subunit forming an individual protopore. Single protopores are gated by a fast gate, whereas the slow gate is assumed to control both protopores through a cooperative movement of the two carboxy-terminal domains. We here study the role of the carboxy-terminal domain in modulating fast and slow gating of human ClC-2 channels, a ubiquitously expressed ClC-type chloride channel involved in transepithelial solute transport and in neuronal chloride homeostasis. Partial truncation of the carboxy-terminus abolishes function of ClC-2 by locking the channel in a closed position. However, unlike other isoforms, its complete removal preserves function of ClC-2. ClC-2 channels without the carboxy-terminus exhibit fast and slow gates that activate and deactivate significantly faster than in WT channels. In contrast to the prevalent view, a single carboxy-terminus suffices for normal slow gating, whereas both domains regulate fast gating of individual protopores. Our findings demonstrate that the carboxy-terminus is not strictly required for slow gating and that the cooperative gating resides in other regions of the channel protein. ClC-2 is expressed in neurons and believed to open at negative potentials and increased internal chloride concentrations after intense synaptic activity. We propose that the function of the ClC-2 carboxy-terminus is to slow down the time course of channel activation in order to stabilize neuronal excitability.

  12. Human diploid fibroblasts have receptors for the globular domain of C1Q

    International Nuclear Information System (INIS)

    Bordin, S.; Page, R.C.

    1986-01-01

    The authors showed that mass cultures of fibroblasts grown from gingival explants in DB medium with 10% human serum are enriched in a phenotype that binds C1q with an affinity much higher than the rest of the population. Because of potential biologic importance of C1q receptors, the authors studied whether the interaction between C1q and this phenotype was mediated by the globular or collagenous domains of the molecule. Globular fragments were prepared by digesting C1q with collagenase, and collagenous fragments obtained after pepsin treatment. C1q binding on cells in suspension was determined by reaction with 125 I-C1q as reported. Competition experiments were performed under conditions in which intact 125 I-C1q binding saturated all available receptors. The results showed that collagenous fragments inhibited 20% of the 125 I-C1q binding to high affinity receptors, whereas inhibition by globular fragments was 70%. Unlabeled intact C1q and collagen type 1 were used as controls, and inhibited 92% and 17% of C1q binding, respectively. These studies show that C1q interacts with the fibroblast phenotype expressing high affinity receptors through its globular domain. The authors suggest that at sites of trauma, native C1 may bind to the surface of these cells via the globular domain of C1q, and that this unique phenotype may play an important role in tissue repair

  13. Zn-binding AZUL domain of human ubiquitin protein ligase Ube3A

    Energy Technology Data Exchange (ETDEWEB)

    Lemak, Alexander; Yee, Adelinda [University of Toronto, and Northeast Structural Genomics Consortium, Ontario Cancer Institute, Campbell Family Cancer Research Institute and Department of Medical Biophysics (Canada); Bezsonova, Irina, E-mail: bezsonova@uchc.edu [University of Connecticut Health Center, Department of Molecular Microbial and Structural Biology (United States); Dhe-Paganon, Sirano, E-mail: sirano.dhepaganon@utoronto.ca [University of Toronto, Structural Genomics Consortium (Canada); Arrowsmith, Cheryl H., E-mail: carrow@uhnresearch.ca [University of Toronto, and Northeast Structural Genomics Consortium, Ontario Cancer Institute, Campbell Family Cancer Research Institute and Department of Medical Biophysics (Canada)

    2011-09-15

    Ube3A (also referred to as E6AP for E6 Associated Protein) is a E3 ubiquitin-protein ligase implicated in the development of Angelman syndrome by controlling degradation of synaptic protein Arc and oncogenic papilloma virus infection by controlling degradation of p53. This article describe the solution NMR structure of the conserved N-terminal domain of human Ube3A (residues 24-87) that contains two residues (Cys44 and Arg62) found to be mutated in patients with Angelman syndrome. The structure of this domain adopts a novel Zn-binding fold we called AZUL (Amino-terminal Zn-finger of Ube3a Ligase). The AZUL domain has a helix-loop-helix architecture with a Zn ion coordinated by four Cys residues arranged in Cys-X{sub 4}-Cys-X{sub 4}-Cys-X{sub 28}-Cys motif. Three of the Zn-bound residues are located in a 23-residue long and well structured loop that connects two {alpha}-helicies.

  14. Zn-binding AZUL domain of human ubiquitin protein ligase Ube3A

    International Nuclear Information System (INIS)

    Lemak, Alexander; Yee, Adelinda; Bezsonova, Irina; Dhe-Paganon, Sirano; Arrowsmith, Cheryl H.

    2011-01-01

    Ube3A (also referred to as E6AP for E6 Associated Protein) is a E3 ubiquitin-protein ligase implicated in the development of Angelman syndrome by controlling degradation of synaptic protein Arc and oncogenic papilloma virus infection by controlling degradation of p53. This article describe the solution NMR structure of the conserved N-terminal domain of human Ube3A (residues 24-87) that contains two residues (Cys44 and Arg62) found to be mutated in patients with Angelman syndrome. The structure of this domain adopts a novel Zn-binding fold we called AZUL (Amino-terminal Zn-finger of Ube3a Ligase). The AZUL domain has a helix-loop-helix architecture with a Zn ion coordinated by four Cys residues arranged in Cys-X 4 -Cys-X 4 -Cys-X 28 -Cys motif. Three of the Zn-bound residues are located in a 23-residue long and well structured loop that connects two α-helicies.

  15. Structural stability of human protein tyrosine phosphatase ρ catalytic domain: effect of point mutations.

    Directory of Open Access Journals (Sweden)

    Alessandra Pasquo

    Full Text Available Protein tyrosine phosphatase ρ (PTPρ belongs to the classical receptor type IIB family of protein tyrosine phosphatase, the most frequently mutated tyrosine phosphatase in human cancer. There are evidences to suggest that PTPρ may act as a tumor suppressor gene and dysregulation of Tyr phosphorylation can be observed in diverse diseases, such as diabetes, immune deficiencies and cancer. PTPρ variants in the catalytic domain have been identified in cancer tissues. These natural variants are nonsynonymous single nucleotide polymorphisms, variations of a single nucleotide occurring in the coding region and leading to amino acid substitutions. In this study we investigated the effect of amino acid substitution on the structural stability and on the activity of the membrane-proximal catalytic domain of PTPρ. We expressed and purified as soluble recombinant proteins some of the mutants of the membrane-proximal catalytic domain of PTPρ identified in colorectal cancer and in the single nucleotide polymorphisms database. The mutants show a decreased thermal and thermodynamic stability and decreased activation energy relative to phosphatase activity, when compared to wild- type. All the variants show three-state equilibrium unfolding transitions similar to that of the wild- type, with the accumulation of a folding intermediate populated at ~4.0 M urea.

  16. Structure of the SH3 domain of human osteoclast-stimulating factor at atomic resolution

    International Nuclear Information System (INIS)

    Chen, Liqing; Wang, Yujun; Wells, David; Toh, Diana; Harold, Hunt; Zhou, Jing; DiGiammarino, Enrico; Meehan, Edward J.

    2006-01-01

    The crystal structure of the SH3 domain of human osteoclast-stimulating factor has been determined and refined to the ultrahigh resolution of 1.07 Å. The structure at atomic resolution provides an accurate framework for structure-based design of its inhibitors. Osteoclast-stimulating factor (OSF) is an intracellular signaling protein, produced by osteoclasts themselves, that enhances osteoclast formation and bone resorption. It is thought to act via an Src-related signaling pathway and contains SH3 and ankyrin-repeat domains which are involved in protein–protein interactions. As part of a structure-based anti-bone-loss drug-design program, the atomic resolution X-ray structure of the recombinant human OSF SH3 domain (hOSF-SH3) has been determined. The domain, residues 12–72, yielded crystals that diffracted to the ultrahigh resolution of 1.07 Å. The overall structure shows a characteristic SH3 fold consisting of two perpendicular β-sheets that form a β-barrel. Structure-based sequence alignment reveals that the putative proline-rich peptide-binding site of hOSF-SH3 consists of (i) residues that are highly conserved in the SH3-domain family, including residues Tyr21, Phe23, Trp49, Pro62, Asn64 and Tyr65, and (ii) residues that are less conserved and/or even specific to hOSF, including Thr22, Arg26, Thr27, Glu30, Asp46, Thr47, Asn48 and Leu60, which might be key to designing specific inhibitors for hOSF to fight osteoporosis and related bone-loss diseases. There are a total of 13 well defined water molecules forming hydrogen bonds with the above residues in and around the peptide-binding pocket. Some of those water molecules might be important for drug-design approaches. The hOSF-SH3 structure at atomic resolution provides an accurate framework for structure-based design of its inhibitors

  17. Comparison of computational to human observer detection for evaluation of CT low dose iterative reconstruction

    Science.gov (United States)

    Eck, Brendan; Fahmi, Rachid; Brown, Kevin M.; Raihani, Nilgoun; Wilson, David L.

    2014-03-01

    Model observers were created and compared to human observers for the detection of low contrast targets in computed tomography (CT) images reconstructed with an advanced, knowledge-based, iterative image reconstruction method for low x-ray dose imaging. A 5-channel Laguerre-Gauss Hotelling Observer (CHO) was used with internal noise added to the decision variable (DV) and/or channel outputs (CO). Models were defined by parameters: (k1) DV-noise with standard deviation (std) proportional to DV std; (k2) DV-noise with constant std; (k3) CO-noise with constant std across channels; and (k4) CO-noise in each channel with std proportional to CO variance. Four-alternative forced choice (4AFC) human observer studies were performed on sub-images extracted from phantom images with and without a "pin" target. Model parameters were estimated using maximum likelihood comparison to human probability correct (PC) data. PC in human and all model observers increased with dose, contrast, and size, and was much higher for advanced iterative reconstruction (IMR) as compared to filtered back projection (FBP). Detection in IMR was better than FPB at 1/3 dose, suggesting significant dose savings. Model(k1,k2,k3,k4) gave the best overall fit to humans across independent variables (dose, size, contrast, and reconstruction) at fixed display window. However Model(k1) performed better when considering model complexity using the Akaike information criterion. Model(k1) fit the extraordinary detectability difference between IMR and FBP, despite the different noise quality. It is anticipated that the model observer will predict results from iterative reconstruction methods having similar noise characteristics, enabling rapid comparison of methods.

  18. Free-form image registration of human cochlear μCT data using skeleton similarity as anatomical prior

    DEFF Research Database (Denmark)

    Kjer, Hans Martin; Fagertun, Jens; Vera, Sergio

    2016-01-01

    Better understanding of the anatomical variability of the human cochlear is important for the design and function of Cochlear Implants. Proper non-rigid alignment of high-resolution cochlear μCT data is a challenge for the typical cubic B-spline registration model. In this paper we study one way ...

  19. Rearrangement of a common cellular DNA domain on chromosome 4 in human primary liver tumors

    International Nuclear Information System (INIS)

    Pasquinelli, C.; Garreau, F.; Bougueleret, L.; Cariani, E.; Thiers, V.; Croissant, O.; Hadchouel, M.; Tiollais, P.; Brechot, C.; Grzeschik, K.H.

    1988-01-01

    Hepatitis B virus (HBV) DNA integration has been shown to occur frequently in human hepatocellular carcinomas. The authors have investigated whether common cellular DNA domains might be rearranged, possibly by HBV integration, in human primary liver tumors. Unique cellular DNA sequences adjacent to an HBV integration site were isolated from a patient with hepatitis B surface antigen-positive hepatocellular carcinoma. These probes detected rearrangement of this cellular region of chromosomal DNA in 3 of 50 additional primary liver tumors studied. Of these three tumor samples, two contained HBV DNA, without an apparent link between the viral DNA and the rearranged allele; HBV DNA sequences were not detected in the third tumor sample. By use of a panel of somatic cell hybrids, these unique cellular DNA sequences were shown to be located on chromosome 4. Therefore, this region of chromosomal DNA might be implicated in the formation of different tumors at one step of liver cell transformation, possible related to HBV integration

  20. Nucleic acid sequences encoding D1 and D1/D2 domains of human coxsackievirus and adenovirus receptor (CAR)

    Science.gov (United States)

    Freimuth, Paul I.

    2010-04-06

    The invention provides recombinant human CAR (coxsackievirus and adenovirus receptor) polypeptides which bind adenovirus. Specifically, polypeptides corresponding to adenovirus binding domain D1 and the entire extracellular domain of human CAR protein comprising D1 and D2 are provided. In another aspect, the invention provides nucleic acid sequences encoding these domains and expression vectors for producing the domains and bacterial cells containing such vectors. The invention also includes an isolated fusion protein comprised of the D1 polypeptide fused to a polypeptide which facilitates folding of D1 when expressed in bacteria. The functional D1 domain finds application in a therapeutic method for treating a patient infected with a CAR D1-binding virus, and also in a method for identifying an antiviral compound which interferes with viral attachment. The invention also provides a method for specifically targeting a cell for infection by a virus which binds to D1.

  1. Guanidine hydrochloride denaturation of human serum albumin originates by local unfolding of some stable loops in domain III.

    Science.gov (United States)

    Ahmad, Basir; Ahmed, Md Zulfazal; Haq, Soghra Khatun; Khan, Rizwan Hasan

    2005-06-15

    The effect of guanidine hydrochloride (GnHCl) on the global stability of human serum albumin (HSA) has been studied by fluorescence and circular dichroism spectroscopic measurements. The differential stability of native conformation of three HSA domains were explored by using domain-specific ligands, hemin (domain I), chloroform (domain II), bilirubin (at domain I/domain II interface) and diazepam (domain III). GnHCl induced unfolding transition curves as monitored by probes for secondary and tertiary structures were cooperative but noncoincidental. A strong ANS binding to the protein was observed around 1.8 M GnHCl, suggesting existence of intermediate states in the unfolding pathway of HSA. A gradual decrease (in the GnHCl concentration range 0.0-1.8 M) in the binding of diazepam indicates that domain III is the most labile to GnHCl denaturation. A significant increase in the binding of bilirubin up to 1.4 M GnHCl and decrease thereafter leading to complete abolishment of bilirubin binding at around 2.0 M GnHCl suggest favorable rearrangement and separation of domains I and II at 1.4 and 2.0 M GnHCl concentration, respectively. Above 1.6 M GnHCl, decrease of the binding of hemin, a ligand for domain I, chloroform, which binds in domain II and lone tryptophanyl fluorescence (Trp-214 located in domain II) indicate that at higher concentration of GnHCl domains I and II start unfolding simultaneously but the stability of domain I (7.4 Kcal/mol) is much more than domain II (4.3 Kcal/mol). A pictorial model for the unfolding of HSA domains, consistent with all these results, has been formulated, suggesting that domain III is the most labile followed by domain II while domain I is the most stable. A molten globule like state of domain III around 1.8 M GnHCl has also been identified and characterized.

  2. Crystal Structure of a Complex of the Intracellular Domain of Interferon λ Receptor 1 (IFNLR1) and the FERM/SH2 Domains of Human JAK1.

    Science.gov (United States)

    Zhang, Di; Wlodawer, Alexander; Lubkowski, Jacek

    2016-11-20

    The crystal structure of a construct consisting of the FERM and SH2-like domains of the human Janus kinase 1 (JAK1) bound to a fragment of the intracellular domain of the interferon-λ receptor 1 (IFNLR1) has been determined at the nominal resolution of 2.1Å. In this structure, the receptor peptide forms an 85-Å-long extended chain, in which both the previously identified box1 and box2 regions bind simultaneously to the FERM and SH2-like domains of JAK1. Both domains of JAK1 are generally well ordered, with regions not seen in the crystal structure limited to loops located away from the receptor-binding regions. The structure provides a much more complete and accurate picture of the interactions between JAK1 and IFNLR1 than those given in earlier reports, illuminating the molecular basis of the JAK-cytokine receptor association. A glutamate residue adjacent to the box2 region in IFNLR1 mimics the mode of binding of a phosphotyrosine in classical SH2 domains. It was shown here that a deletion of residues within the box1 region of the receptor abolishes stable interactions with JAK1, although it was previously shown that box2 alone is sufficient to stabilize a similar complex of the interferon-α receptor and TYK2. Published by Elsevier Ltd.

  3. Crystal structure of the C-terminal domain of the RAP74 subunit of human transcription factor IIF

    Energy Technology Data Exchange (ETDEWEB)

    Kamada, Katsuhiko; De Angelis, Jacqueline; Roeder, Robert G.; Burley, Stephen K. (Rockefeller)

    2012-12-13

    The x-ray structure of a C-terminal fragment of the RAP74 subunit of human transcription factor (TF) IIF has been determined at 1.02-{angstrom} resolution. The {alpha}/{beta} structure is strikingly similar to the globular domain of linker histone H5 and the DNA-binding domain of hepatocyte nuclear factor 3{gamma} (HNF-3{gamma}), making it a winged-helix protein. The surface electrostatic properties of this compact domain differ significantly from those of bona fide winged-helix transcription factors (HNF-3{gamma} and RFX1) and from the winged-helix domains found within the RAP30 subunit of TFIIF and the {beta} subunit of TFIIE. RAP74 has been shown to interact with the TFIIF-associated C-terminal domain phosphatase FCP1, and a putative phosphatase binding site has been identified within the RAP74 winged-helix domain.

  4. Crystal structure of the extracellular domain of human myelin protein zero

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Zhigang; Wang, Yong; Yedidi, Ravikiran S.; Brunzelle, Joseph S.; Kovari, Iulia A.; Sohi, Jasloveleen; Kamholz, John; Kovari, Ladislau C. (WSU-MED); (NWU)

    2012-03-27

    Charcot-Marie-Tooth disease (CMT), a hereditary motor and sensory neuropathy, is the most common genetic neuropathy with an incidence of 1 in 2600. Several forms of CMT have been identified arising from different genomic abnormalities such as CMT1 including CMT1A, CMT1B, and CMTX. CMT1 with associated peripheral nervous system (PNS) demyelination, the most frequent diagnosis, demonstrates slowed nerve conduction velocities and segmental demyelination upon nerve biopsy. One of its subtypes, CMT1A, presents a 1.5-Mb duplication in the p11-p12 region of the human chromosome 17 which encodes peripheral myelin protein 22 (PMP22). CMT1B, a less common form, arises from the mutations in the myelin protein zero (MPZ) gene on chromosome 1, region q22-q23, which encodes the major structural component of the peripheral myelin. A rare type of CMT1 has been found recently and is caused by point mutations in early growth response gene 2 (EGR2), encoding a zinc finger transcription factor in Schwann cells. In addition, CMTX, an X-linked form of CMT, arises from a mutation in the connexin-32 gene. Myelin protein zero, associated with CMT1B, is a transmembrane protein of 219 amino acid residues. Human MPZ consists of three domains: 125 residues constitute the glycosylated immunoglobulin-like extracellular domain; 27 residues span the membrane; and 67 residues comprise the highly basic intracellular domain. MPZ makes up approximately 50% of the protein content of myelin, and is expressed predominantly in Schwann cells, the myelinating cell of the PNS. Myelin protein zero, a homophilic adhesion molecule, is a member of the immunoglobulin super-family and is essential for normal myelin structure and function. In addition, MPZ knockout mice displayed abnormal myelin that severely affects the myelination pathway, and overexpression of MPZ causes congenital hypomyelination of peripheral nerves. Myelin protein zero mutations account for {approx}5% of patients with CMT. To date, over 125

  5. Usefulness of 2D fusion of postmortem CT and antemortem chest radiography studies for human identification.

    Science.gov (United States)

    Shinkawa, Norihiro; Hirai, Toshinori; Nishii, Ryuichi; Yukawa, Nobuhiro

    2017-06-01

    To determine the feasibility of human identification through the two-dimensional (2D) fusion of postmortem computed tomography (PMCT) and antemortem chest radiography. The study population consisted of 15 subjects who had undergone chest radiography studies more than 12 months before death. Fused images in which a chest radiograph was fused with a PMCT image were obtained for those subjects using a workstation, and the minimum distance gaps between corresponding anatomical landmarks (located at soft tissue and bone sites) in the images obtained with the two modalities were calculated. For each fused image, the mean of all these minimum distance gaps was recorded as the mean distance gap (MDG). For each subject, the MDG obtained for the same-subject fused image (i.e., where both of the images that were fused derived from that subject) was compared with the MDGs for different-subject fused images (i.e., where only one of the images that were fused derived from that subject; the other image derived from a different subject) in order to determine whether same-subject fused images can be reliably distinguished from different-subject fused images. The MDGs of the same-subject fused images were found to be significantly smaller than the MDGs of the different-subject fused images (p chest radiography and postmortem CT images may assist in human identification.

  6. Morphometric studies on the facial skeleton of humans and pongids based on CT-scans.

    Science.gov (United States)

    Schumacher, K U; Koppe, T; Fanghänel, J; Schumacher, G H; Nagai, H

    1994-10-01

    The changes of the skull, which we can observe during the anthropogenesis, are reflected especially in the different skull proportions. We carried out metric measurements at the median level on 10 adult skulls each of humans, chimpanzees and gorillas as well as 11 skulls of orangutans. All skulls were scanned with a CT at the median level. We measured the lines and angles of the scans and the means and the standard deviations were calculated. We carried out a correlation analysis to observe the relation of their characteristics. We showed that there is a relation between the length of the skull base and the facial length in all species. From the results of the correlation analysis, we can also conclude that a relation exists between the degree of prognathism and the different length measurements of the facial skeleton. We also found a bending of the facial skeleton in relation to the cranial base towards the ventral side, also known as klinorhynchy, in all observed species. The highest degree of klinorhynchy was found in humans and the lowest in orangutans. We will discuss the different definition of the term klinorhynchy and its importance in the evolution of the hominoids.

  7. Extending NGOMSL Model for Human-Humanoid Robot Interaction in the Soccer Robotics Domain

    Directory of Open Access Journals (Sweden)

    Rajesh Elara Mohan

    2008-01-01

    Full Text Available In the field of human-computer interaction, the Natural Goals, Operators, Methods, and Selection rules Language (NGOMSL model is one of the most popular methods for modelling knowledge and cognitive processes for rapid usability evaluation. The NGOMSL model is a description of the knowledge that a user must possess to operate the system represented as elementary actions for effective usability evaluations. In the last few years, mobile robots have been exhibiting a stronger presence in commercial markets and very little work has been done with NGOMSL modelling for usability evaluations in the human-robot interaction discipline. This paper focuses on extending the NGOMSL model for usability evaluation of human-humanoid robot interaction in the soccer robotics domain. The NGOMSL modelled human-humanoid interaction design of Robo-Erectus Junior was evaluated and the results of the experiments showed that the interaction design was able to find faults in an average time of 23.84 s. Also, the interaction design was able to detect the fault within the 60 s in 100% of the cases. The Evaluated Interaction design was adopted by our Robo-Erectus Junior version of humanoid robots in the RoboCup 2007 humanoid soccer league.

  8. Characterization of Human and Murine T-Cell Immunoglobulin Mucin Domain 4 (TIM-4) IgV Domain Residues Critical for Ebola Virus Entry.

    Science.gov (United States)

    Rhein, Bethany A; Brouillette, Rachel B; Schaack, Grace A; Chiorini, John A; Maury, Wendy

    2016-07-01

    Phosphatidylserine (PtdSer) receptors that are responsible for the clearance of dying cells have recently been found to mediate enveloped virus entry. Ebola virus (EBOV), a member of the Filoviridae family of viruses, utilizes PtdSer receptors for entry into target cells. The PtdSer receptors human and murine T-cell immunoglobulin mucin (TIM) domain proteins TIM-1 and TIM-4 mediate filovirus entry by binding to PtdSer on the virion surface via a conserved PtdSer binding pocket within the amino-terminal IgV domain. While the residues within the TIM-1 IgV domain that are important for EBOV entry are characterized, the molecular details of virion-TIM-4 interactions have yet to be investigated. As sequences and structural alignments of the TIM proteins suggest distinct differences in the TIM-1 and TIM-4 IgV domain structures, we sought to characterize TIM-4 IgV domain residues required for EBOV entry. Using vesicular stomatitis virus pseudovirions bearing EBOV glycoprotein (EBOV GP/VSVΔG), we evaluated virus binding and entry into cells expressing TIM-4 molecules mutated within the IgV domain, allowing us to identify residues important for entry. Similar to TIM-1, residues in the PtdSer binding pocket of murine and human TIM-4 (mTIM-4 and hTIM-4) were found to be important for EBOV entry. However, additional TIM-4-specific residues were also found to impact EBOV entry, with a total of 8 mTIM-4 and 14 hTIM-4 IgV domain residues being critical for virion binding and internalization. Together, these findings provide a greater understanding of the interaction of TIM-4 with EBOV virions. With more than 28,000 cases and over 11,000 deaths during the largest and most recent Ebola virus (EBOV) outbreak, there has been increased emphasis on the development of therapeutics against filoviruses. Many therapies under investigation target EBOV cell entry. T-cell immunoglobulin mucin (TIM) domain proteins are cell surface factors important for the entry of many enveloped viruses

  9. [Peptide fragments of chemokine domain of fractalkine: effect on human monocyte migration].

    Science.gov (United States)

    Kukhtina, N B; Aref'eva, T I; Ruleva, N Iu; Sidorova, M V; Az'muko, A A; Bespalova, Zh D; Krasnikova, T L

    2012-01-01

    Leukocyte chemotaxis to the area of tissue damage is mediated by chemokines. According to the primary structure, chemokines are divided into four families, fractalkine (CX3CL1) is the only one member of CX3C family and the only membrane-bound chemokine. Fractalkine molecule includes the extracellular N-terminal chemokine domain, mucin-like rod, the transmembrane and the intracellular domains. In membrane-bound state fractalkine has the properties of an adhesion molecule. Chemokine domain of fractalkine (CDF) is released from cell membrane by proteolysis, and this soluble form acts as a chemoattractant for leukocytes expressing fractalkine receptor CX3CR1. Fractalkine is involved in development of a number of pathological processes caused by inflammation, and therefore a search for fractalkine inhibitors is very important. For this purpose we identified several antigenic determinants--the fragments of CDF, and the following peptides were synthesized--P41-52 H-Leu-Glu-Thr-Arg-Gln-His-Arg-Leu-Phe-Cys-Ala-Asp-NH2, P53-60 H-Pro-Lys-Glu-Gln-Trp-Val-Lys-Asp-NH2 and P60-71 H-Asp-Ala-Met-Gln-His-Leu-Asp-Arg-Gln-Ala-Ala-Ala-NH2. The peptide effects on adhesion and migration of human peripheral blood monocytes expressing fractalkine receptors were investigated. In the presence of CDF and P41-52 we observed the increased adhesion and migration of monocytes compared with spontaneous values. Peptides P53-60 and P60-71 significantly inhibited monocyte adhesion and migration stimulated by CDF. Since the chemotactic activity of chemokines was shown to be dependent on their binding to glycosaminoglycans of the cell surface and extracellular matrix, the effect ofpeptides on the interaction of CDF with heparin was analyzed by ELISA. Peptide P41-52 competed with CDF for heparin binding, while peptides P53-60 and P60-71 had no significant activity.

  10. Structure of FGFR3 transmembrane domain dimer: implications for signaling and human pathologies.

    Science.gov (United States)

    Bocharov, Eduard V; Lesovoy, Dmitry M; Goncharuk, Sergey A; Goncharuk, Marina V; Hristova, Kalina; Arseniev, Alexander S

    2013-11-05

    Fibroblast growth factor receptor 3 (FGFR3) transduces biochemical signals via lateral dimerization in the plasma membrane, and plays an important role in human development and disease. Eight different pathogenic mutations, implicated in cancers and growth disorders, have been identified in the FGFR3 transmembrane segment. Here, we describe the dimerization of the FGFR3 transmembrane domain in membrane-mimicking DPC/SDS (9/1) micelles. In the solved NMR structure, the two transmembrane helices pack into a symmetric left-handed dimer, with intermolecular stacking interactions occurring in the dimer central region. Some pathogenic mutations fall within the helix-helix interface, whereas others are located within a putative alternative interface. This implies that although the observed dimer structure is important for FGFR3 signaling, the mechanism of FGFR3-mediated transduction across the membrane is complex. We propose an FGFR3 signaling mechanism that is based on the solved structure, available structures of isolated soluble FGFR domains, and published biochemical and biophysical data. Copyright © 2013 Elsevier Ltd. All rights reserved.

  11. Disruption of PH–kinase domain interactions leads to oncogenic activation of AKT in human cancers

    Science.gov (United States)

    Parikh, Chaitali; Janakiraman, Vasantharajan; Wu, Wen-I; Foo, Catherine K.; Kljavin, Noelyn M.; Chaudhuri, Subhra; Stawiski, Eric; Lee, Brian; Lin, Jie; Li, Hong; Lorenzo, Maria N.; Yuan, Wenlin; Guillory, Joseph; Jackson, Marlena; Rondon, Jesus; Franke, Yvonne; Bowman, Krista K.; Sagolla, Meredith; Stinson, Jeremy; Wu, Thomas D.; Wu, Jiansheng; Stokoe, David; Stern, Howard M.; Brandhuber, Barbara J.; Lin, Kui; Skelton, Nicholas J.; Seshagiri, Somasekar

    2012-01-01

    The protein kinase v-akt murine thymoma viral oncogene homolog (AKT), a key regulator of cell survival and proliferation, is frequently hyperactivated in human cancers. Intramolecular pleckstrin homology (PH) domain–kinase domain (KD) interactions are important in maintaining AKT in an inactive state. AKT activation proceeds after a conformational change that dislodges the PH from the KD. To understand these autoinhibitory interactions, we generated mutations at the PH–KD interface and found that most of them lead to constitutive activation of AKT. Such mutations are likely another mechanism by which activation may occur in human cancers and other diseases. In support of this likelihood, we found somatic mutations in AKT1 at the PH–KD interface that have not been previously described in human cancers. Furthermore, we show that the AKT1 somatic mutants are constitutively active, leading to oncogenic signaling. Additionally, our studies show that the AKT1 mutants are not effectively inhibited by allosteric AKT inhibitors, consistent with the requirement for an intact PH–KD interface for allosteric inhibition. These results have important implications for therapeutic intervention in patients with AKT mutations at the PH–KD interface. PMID:23134728

  12. TGFβ1-mediated expression and alternative splicing of Fibronectin Extra Domain A in human podocyte culture.

    Science.gov (United States)

    Madne, Tarunkumar Hemraj; Dockrell, Mark Edward Carl

    2018-02-28

    Alternative splicing is a fundamental phenomenon to build protein diversity in health and diseases. Extra Domain A+ Fibronectin (EDA+Fn) is an alternatively spliced form of fibronectin protein present in the extra cellular matrix (ECM) in renal fibrosis. Podocytes are spectacular cell type and play a key role in filtration and synthesise ECM proteins in renal physiology and pathology. TGFβ1 is a strong stimulator of ECM proteins in renal injury. In this study, we have investigated alternative splicing of EDA+ Fn in human podocytes in response to TGFβ1. We have performed western blotting and immunofluorescence to characterise the expression of the EDA+Fn protein, real-time PCR for RNA expression and RT-PCR to look for alternative splicing of EDA+Fn in conditionally immortalised human podocytes culture.We used TGFβ1 as a stimulator and SB431542 and SRPIN340 for inhibitory studies. In this work, for the first time we have demonstrated in human podocytes culture EDA+Fn is expressed in the basal condition and TGFβ1 2.5ng/ml induced the Fn mRNA and EDA+Fn protein expression demonstrated by real-time PCR, western blotting and immunofluorescence. TGFβ1 2.5ng/ml induced the alternative splicing of EDA+Fn shown by conventional RT-PCR. Studies with ALK5 inhibitor SB431542 and SRPIN340 show that TGFβ1 induced alternative splicing of EDA+Fn was by the ALK5 receptor and the SR proteins.  In human podocytes culture, alternative splicing of EDA+Fn occurs at basal conditions and TGFβ1 further induced the alternative splicing of EDA+Fn via ALK5 receptor activation and SR proteins. This is the first evidence of basal and TGFβ1 mediated alternative splicing of EDA+Fn in human podocytes culture.

  13. Structure of a new crystal form of human Hsp70 ATPase domain.

    Science.gov (United States)

    Osipiuk, J; Walsh, M A; Freeman, B C; Morimoto, R I; Joachimiak, A

    1999-05-01

    Hsp70 proteins are highly conserved proteins induced by heat shock and other stress conditions. An ATP-binding domain of human Hsp70 protein has been crystallized in two major morphological forms at pH 7.0 in the presence of PEG 8000 and CaCl2. Both crystal forms belong to the orthorhombic space group P212121, but show no resemblance in unit-cell parameters. Analysis of the crystal structures for both forms shows a 1-2 A shift of one of the subdomains of the protein. This conformational change could reflect a 'natural' flexibility of the protein which might be relevant to ATP binding and may facilitate the interaction of other proteins with Hsp70 protein.

  14. Structural Basis for Substrate Recognition by the Ankyrin Repeat Domain of Human DHHC17 Palmitoyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    Verardi, Raffaello; Kim, Jin-Sik; Ghirlando, Rodolfo; Banerjee, Anirban

    2017-09-01

    DHHC enzymes catalyze palmitoylation, a major post-translational modification that regulates a number of key cellular processes. There are up to 24 DHHCs in mammals and hundreds of substrate proteins that get palmitoylated. However, how DHHC enzymes engage with their substrates is still poorly understood. There is currently no structural information about the interaction between any DHHC enzyme and protein substrates. In this study we have investigated the structural and thermodynamic bases of interaction between the ankyrin repeat domain of human DHHC17 (ANK17) and Snap25b. We solved a high-resolution crystal structure of the complex between ANK17 and a peptide fragment of Snap25b. Through structure-guided mutagenesis, we discovered key residues in DHHC17 that are critically important for interaction with Snap25b. We further extended our finding by showing that the same residues are also crucial for the interaction of DHHC17 with Huntingtin, one of its most physiologically relevant substrates.

  15. Human mate-choice copying is domain-general social learning.

    Science.gov (United States)

    Street, Sally E; Morgan, Thomas J H; Thornton, Alex; Brown, Gillian R; Laland, Kevin N; Cross, Catharine P

    2018-01-29

    Women appear to copy other women's preferences for men's faces. This 'mate-choice copying' is often taken as evidence of psychological adaptations for processing social information related to mate choice, for which facial information is assumed to be particularly salient. No experiment, however, has directly investigated whether women preferentially copy each other's face preferences more than other preferences. Further, because prior experimental studies used artificial social information, the effect of real social information on attractiveness preferences is unknown. We collected attractiveness ratings of pictures of men's faces, men's hands, and abstract art given by heterosexual women, before and after they saw genuine social information gathered in real time from their peers. Ratings of faces were influenced by social information, but no more or less than were images of hands and abstract art. Our results suggest that evidence for domain-specific social learning mechanisms in humans is weaker than previously suggested.

  16. Similarity of recombinant human perlecan domain 1 by alternative expression systems bioactive heterogenous recombinant human perlecan D1

    DEFF Research Database (Denmark)

    Ellis, April L; Pan, Wensheng; Yang, Guang

    2010-01-01

    BACKGROUND: Heparan sulfate glycosaminoglycans are diverse components of certain proteoglycans and are known to interact with growth factors as a co-receptor necessary to induce signalling and growth factor activity. In this report we characterize heterogeneously glycosylated recombinant human...... perlecan domain 1 (HSPG2 abbreviated as rhPln.D1) synthesized in either HEK 293 cells or HUVECs by transient gene delivery using either adenoviral or expression plasmid technology. RESULTS: By SDS-PAGE analysis following anion exchange chromatography, the recombinant proteoglycans appeared to possess...... glycosaminoglycan chains ranging, in total, from 6 kDa to >90 kDa per recombinant. Immunoblot analysis of enzyme-digested high Mr rhPln.D1 demonstrated that the rhPln.D1 was synthesized as either a chondroitin sulfate or heparan sulfate proteoglycan, in an approximately 2:1 ratio, with negligible hybrids. Secondary...

  17. Systematic analysis of FKBP inducible degradation domain tagging strategies for the human malaria parasite Plasmodium falciparum.

    Directory of Open Access Journals (Sweden)

    Mauro Ferreira de Azevedo

    Full Text Available Targeted regulation of protein levels is an important tool to gain insights into the role of proteins essential to cell function and development. In recent years, a method based on mutated forms of the human FKBP12 has been established and used to great effect in various cell types to explore protein function. The mutated FKBP protein, referred to as destabilization domain (DD tag when fused with a native protein at the N- or C-terminus targets the protein for proteosomal degradation. Regulated expression is achieved via addition of a compound, Shld-1, that stabilizes the protein and prevents degradation. A limited number of studies have used this system to provide powerful insight into protein function in the human malaria parasite Plasmodium falciparum. In order to better understand the DD inducible system in P. falciparum, we studied the effect of Shld-1 on parasite growth, demonstrating that although development is not impaired, it is delayed, requiring the appropriate controls for phenotype interpretation. We explored the quantified regulation of reporter Green Fluorescent Protein (GFP and luciferase constructs fused to three DD variants in parasite cells either via transient or stable transfection. The regulation obtained with the original FKBP derived DD domain was compared to two triple mutants DD24 and DD29, which had been described to provide better regulation for C-terminal tagging in other cell types. When cloned to the C-terminal of reporter proteins, DD24 provided the strongest regulation allowing reporter activity to be reduced to lower levels than DD and to restore the activity of stabilised proteins to higher levels than DD29. Importantly, DD24 has not previously been applied to regulate proteins in P. falciparum. The possibility of regulating an exported protein was addressed by targeting the Ring-Infected Erythrocyte Surface Antigen (RESA at its C-terminus. The tagged protein demonstrated an important modulation of its

  18. Applications of human hepatitis B virus preS domain in bio- and nanotechnology.

    Science.gov (United States)

    Toita, Riki; Kawano, Takahito; Kang, Jeong-Hun; Murata, Masaharu

    2015-06-28

    Human hepatitis B virus (HBV) is a member of the family Hepadnaviridae, and causes acute and chronic infections of the liver. The hepatitis B surface antigen (HBsAg) contains the large (L), middle (M), and small (S) surface proteins. The L protein consists of the S protein, preS1, and preS2. In HBsAg, the preS domain (preS1 + preS2) plays a key role in the infection of hepatocytic cells by HBV and has several immunogenic epitopes. Based on these characteristics of preS, several preS-based diagnostic and therapeutic materials and systems have been developed. PreS1-specific monoclonal antibodies (e.g., MA18/7 and KR127) can be used to inhibit HBV infection. A myristoylated preS1 peptide (amino acids 2-48) also inhibits the attachment of HBV to HepaRG cells, primary human hepatocytes, and primary tupaia hepatocytes. Antibodies and antigens related to the components of HBsAg, preS (preS1 + preS2), or preS1 can be available as diagnostic markers of acute and chronic HBV infections. Hepatocyte-targeting delivery systems for therapeutic molecules (drugs, genes, or proteins) are very important for increasing the clinical efficacy of these molecules and in reducing their adverse effects on other organs. The selective delivery of diagnostic molecules to target hepatocytic cells can also improve the efficiency of diagnosis. In addition to the full-length HBV vector, preS (preS1 + preS2), preS1, and preS1-derived fragments can be useful in hepatocyte-specific targeting. In this review, we discuss the literature concerning the applications of the HBV preS domain in bio- and nanotechnology.

  19. Purification, crystallization and preliminary X-ray analysis of the IgV domain of human nectin-4

    OpenAIRE

    Xu, Xiang; Zhang, Xiaoai; Lu, Guangwen; Cai, Yongping

    2012-01-01

    Nectin-4 belongs to a family of immunoglobulin-like cell adhesion molecules and is highly expressed in cancer cells. Recently, nectin-4 was found to be a receptor of measles virus and the IgV domain sustains strong binding to measles virus H protein. In this study, the successful expression and purification of human nectin-4 V domain (nectin-4v) is reported

  20. MO-FG-204-03: Using Edge-Preserving Algorithm for Significantly Improved Image-Domain Material Decomposition in Dual Energy CT

    International Nuclear Information System (INIS)

    Zhao, W; Niu, T; Xing, L; Xiong, G; Elmore, K; Min, J; Zhu, J; Wang, L

    2015-01-01

    Purpose: To significantly improve dual energy CT (DECT) imaging by establishing a new theoretical framework of image-domain material decomposition with incorporation of edge-preserving techniques. Methods: The proposed algorithm, HYPR-NLM, combines the edge-preserving non-local mean filter (NLM) with the HYPR-LR (Local HighlY constrained backPRojection Reconstruction) framework. Image denoising using HYPR-LR framework depends on the noise level of the composite image which is the average of the different energy images. For DECT, the composite image is the average of high- and low-energy images. To further reduce noise, one may want to increase the window size of the filter of the HYPR-LR, leading resolution degradation. By incorporating the NLM filtering and the HYPR-LR framework, HYPR-NLM reduces the boost material decomposition noise using energy information redundancies as well as the non-local mean. We demonstrate the noise reduction and resolution preservation of the algorithm with both iodine concentration numerical phantom and clinical patient data by comparing the HYPR-NLM algorithm to the direct matrix inversion, HYPR-LR and iterative image-domain material decomposition (Iter-DECT). Results: The results show iterative material decomposition method reduces noise to the lowest level and provides improved DECT images. HYPR-NLM significantly reduces noise while preserving the accuracy of quantitative measurement and resolution. For the iodine concentration numerical phantom, the averaged noise levels are about 2.0, 0.7, 0.2 and 0.4 for direct inversion, HYPR-LR, Iter- DECT and HYPR-NLM, respectively. For the patient data, the noise levels of the water images are about 0.36, 0.16, 0.12 and 0.13 for direct inversion, HYPR-LR, Iter-DECT and HYPR-NLM, respectively. Difference images of both HYPR-LR and Iter-DECT show edge effect, while no significant edge effect is shown for HYPR-NLM, suggesting spatial resolution is well preserved for HYPR-NLM. Conclusion: HYPR

  1. Impact of the [delta]F508 Mutation in First Nucleotide-binding Domain of Human Cystic Fibrosis Transmembrane Conductance Regulator on Domain Folding and Structure

    Energy Technology Data Exchange (ETDEWEB)

    Lewis, Hal A.; Zhao, Xun; Wang, Chi; Sauder, J. Michael; Rooney, Isabelle; Noland, Brian W.; Lorimer, Don; Kearins, Margaret C.; Conners, Kris; Condon, Brad; Maloney, Peter C.; Guggino, William B.; Hunt, John F.; Emtage, Spencer (SG); (Columbia); (JHU)

    2010-07-19

    Cystic fibrosis is caused by defects in the cystic fibrosis transmembrane conductance regulator (CFTR), commonly the deletion of residue Phe-508 (DeltaF508) in the first nucleotide-binding domain (NBD1), which results in a severe reduction in the population of functional channels at the epithelial cell surface. Previous studies employing incomplete NBD1 domains have attributed this to aberrant folding of DeltaF508 NBD1. We report structural and biophysical studies on complete human NBD1 domains, which fail to demonstrate significant changes of in vitro stability or folding kinetics in the presence or absence of the DeltaF508 mutation. Crystal structures show minimal changes in protein conformation but substantial changes in local surface topography at the site of the mutation, which is located in the region of NBD1 believed to interact with the first membrane spanning domain of CFTR. These results raise the possibility that the primary effect of DeltaF508 is a disruption of proper interdomain interactions at this site in CFTR rather than interference with the folding of NBD1. Interestingly, increases in the stability of NBD1 constructs are observed upon introduction of second-site mutations that suppress the trafficking defect caused by the DeltaF508 mutation, suggesting that these suppressors might function indirectly by improving the folding efficiency of NBD1 in the context of the full-length protein. The human NBD1 structures also solidify the understanding of CFTR regulation by showing that its two protein segments that can be phosphorylated both adopt multiple conformations that modulate access to the ATPase active site and functional interdomain interfaces.

  2. A graph kernel approach for alignment-free domain-peptide interaction prediction with an application to human SH3 domains.

    Science.gov (United States)

    Kundu, Kousik; Costa, Fabrizio; Backofen, Rolf

    2013-07-01

    State-of-the-art experimental data for determining binding specificities of peptide recognition modules (PRMs) is obtained by high-throughput approaches like peptide arrays. Most prediction tools applicable to this kind of data are based on an initial multiple alignment of the peptide ligands. Building an initial alignment can be error-prone, especially in the case of the proline-rich peptides bound by the SH3 domains. Here, we present a machine-learning approach based on an efficient graph-kernel technique to predict the specificity of a large set of 70 human SH3 domains, which are an important class of PRMs. The graph-kernel strategy allows us to (i) integrate several types of physico-chemical information for each amino acid, (ii) consider high-order correlations between these features and (iii) eliminate the need for an initial peptide alignment. We build specialized models for each human SH3 domain and achieve competitive predictive performance of 0.73 area under precision-recall curve, compared with 0.27 area under precision-recall curve for state-of-the-art methods based on position weight matrices. We show that better models can be obtained when we use information on the noninteracting peptides (negative examples), which is currently not used by the state-of-the art approaches based on position weight matrices. To this end, we analyze two strategies to identify subsets of high confidence negative data. The techniques introduced here are more general and hence can also be used for any other protein domains, which interact with short peptides (i.e. other PRMs). The program with the predictive models can be found at http://www.bioinf.uni-freiburg.de/Software/SH3PepInt/SH3PepInt.tar.gz. We also provide a genome-wide prediction for all 70 human SH3 domains, which can be found under http://www.bioinf.uni-freiburg.de/Software/SH3PepInt/Genome-Wide-Predictions.tar.gz. Supplementary data are available at Bioinformatics online.

  3. Crystal structure of NL63 respiratory coronavirus receptor-binding domain complexed with its human receptor

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Kailang; Li, Weikai; Peng, Guiqing; Li, Fang; (Harvard-Med); (UMM-MED)

    2010-03-04

    NL63 coronavirus (NL63-CoV), a prevalent human respiratory virus, is the only group I coronavirus known to use angiotensin-converting enzyme 2 (ACE2) as its receptor. Incidentally, ACE2 is also used by group II SARS coronavirus (SARS-CoV). We investigated how different groups of coronaviruses recognize the same receptor, whereas homologous group I coronaviruses recognize different receptors. We determined the crystal structure of NL63-CoV spike protein receptor-binding domain (RBD) complexed with human ACE2. NL63-CoV RBD has a novel {beta}-sandwich core structure consisting of 2 layers of {beta}-sheets, presenting 3 discontinuous receptor-binding motifs (RBMs) to bind ACE2. NL63-CoV and SARS-CoV have no structural homology in RBD cores or RBMs; yet the 2 viruses recognize common ACE2 regions, largely because of a 'virus-binding hotspot' on ACE2. Among group I coronaviruses, RBD cores are conserved but RBMs are variable, explaining how these viruses recognize different receptors. These results provide a structural basis for understanding viral evolution and virus-receptor interactions.

  4. In Vitro Maturation of a Humanized Shark VNAR Domain to Improve Its Biophysical Properties to Facilitate Clinical Development

    Directory of Open Access Journals (Sweden)

    John Steven

    2017-10-01

    Full Text Available Molecular engineering to increase the percentage identity to common human immunoglobulin sequences of non-human therapeutic antibodies and scaffolds has become standard practice. This strategy is often used to reduce undesirable immunogenic responses, accelerating the clinical development of candidate domains. The first humanized shark variable domain (VNAR was reported by Kovalenko and colleagues and used the anti-human serum albumin (HSA domain, clone E06, as a model to construct a number of humanized versions including huE06v1.10. This study extends this work by using huE06v1.10 as a template to isolate domains with improved biophysical properties and reduced antigenicity. Random mutagenesis was conducted on huE06v1.10 followed by refinement of clones through an off-rate ranking-based selection on target antigen. Many of these next-generation binders retained high affinity for target, together with good species cross-reactivity. Lead domains were assessed for any tendency to dimerize, tolerance to N- and C-terminal fusions, affinity, stability, and relative antigenicity in human dendritic cell assays. Functionality of candidate clones was verified in vivo through the extension of serum half-life in a typical drug format. From these analyses the domain, BA11, exhibited negligible antigenicity, high stability and high affinity for mouse, rat, and HSA. When these attributes were combined with demonstrable functionality in a rat model of PK, the BA11 clone was established as our clinical candidate.

  5. Characterization of Human and Murine T-Cell Immunoglobulin Mucin Domain 4 (TIM-4) IgV Domain Residues Critical for Ebola Virus Entry

    OpenAIRE

    Rhein, Bethany A.; Brouillette, Rachel B.; Schaack, Grace A.; Chiorini, John A.; Maury, Wendy

    2016-01-01

    Phosphatidylserine (PtdSer) receptors that are responsible for the clearance of dying cells have recently been found to mediate enveloped virus entry. Ebola virus (EBOV), a member of the Filoviridae family of viruses, utilizes PtdSer receptors for entry into target cells. The PtdSer receptors human and murine T-cell immunoglobulin mucin (TIM) domain proteins TIM-1 and TIM-4 mediate filovirus entry by binding to PtdSer on the virion surface via a conserved PtdSer binding pocket within the amin...

  6. The structure function of the death domain of human IRAK-M.

    Science.gov (United States)

    Du, Jiangfeng; Nicolaes, Gerry Af; Kruijswijk, Danielle; Versloot, Miranda; van der Poll, Tom; van 't Veer, Cornelis

    2014-12-07

    IRAK-M is an inhibitor of Toll-like receptor signaling that acts by re-directing IRAK-4 activity to TAK1 independent NF-κB activation and by inhibition of IRAK-1/IRAK-2 activity. IRAK-M is expressed in monocytes/macrophages and lung epithelial cells. Lack of IRAK-M in mice greatly improves the resistance to nosocomial pneumonia and lung tumors, which entices IRAK-M as a potential therapeutic target. IRAK-M consists of an N-terminal death domain (DD), a dysfunctional kinase domain and unstructured C-terminal domain. Little is known however on IRAK-M's structure-function relationships. Since death domains provide the important interactions of IRAK-1, IRAK-2 and IRAK-4 molecules, we generated a 3D structure model of the human IRAK-M-DD (residues C5-G119) to guide mutagenesis studies and predict protein-protein interaction points. First we identified the DD residues involved in the endogenous capacity of IRAK-M to activate NF-κB that is displayed upon overexpression in 293T cells. W74 and R97, at distinct interfaces of the IRAK-M-DD, were crucial for this endogenous NF-κB activating capacity, as well as the C-terminal domain (S445-E596) of IRAK-M. Resulting anti-inflammatory A20 and pro-inflammatory IL-8 transcription in 293T cells was W74 dependent, while IL-8 protein expression was dependent on R97 and the TRAF6 binding motif at P478. The IRAK-M-DD W74 and R97 binding interfaces are predicted to interact with opposite sides of IRAK-4-DD's. Secondly we identified DD residues important for the inhibitory action of IRAK-M by stable overexpression of mutants in THP-1 macrophages and H292 lung epithelial cells. IRAK-M inhibited TLR2/4-mediated cytokine production in macrophages in a manner that is largely dependent on W74. R97 was not involved in inhibition of TNF production but was engaged in IL-6 down-regulation by IRAK-M. Protein-interactive residues D19-A23, located in between W74 and R97, were also observed to be crucial for inhibition of TLR2/4 mediated cytokine

  7. Spiral CT scanning plan to generate accurate Fe models of the human femur

    International Nuclear Information System (INIS)

    Zannoni, C.; Testi, D.; Capello, A.

    1999-01-01

    In spiral computed tomography (CT), source rotation, patient translation, and data acquisition are continuously conducted. Settings of the detector collimation and the table increment affect the image quality in terms of spatial and contrast resolution. This study assessed and measured the efficacy of spiral CT in those applications where the accurate reconstruction of bone morphology is critical: custom made prosthesis design or three dimensional modelling of the mechanical behaviour of long bones. Results show that conventional CT grants the highest accuracy. Spiral CT with D=5 mm and P=1,5 in the regions where the morphology is more regular, slightly degrades the image quality but allows to acquire at comparable cost an higher number of images increasing the longitudinal resolution of the acquired data set. (author)

  8. From the chromatin interaction network to the organization of the human genome into replication N/U-domains

    International Nuclear Information System (INIS)

    Boulos, Rasha E; Julienne, Hanna; Baker, Antoine; Jensen, Pablo; Arneodo, Alain; Audit, Benjamin; Chen, Chun-Long; D'Aubenton-Carafa, Yves; Thermes, Claude; Petryk, Nataliya; Kahli, Malik; Hyrien, Olivier; Goldar, Arach

    2014-01-01

    The three-dimensional (3D) architecture of the mammalian nucleus is now being unraveled thanks to the recent development of chromatin conformation capture (3C) technologies. Here we report the results of a combined multiscale analysis of genome-wide mean replication timing and chromatin conformation data that reveal some intimate relationships between chromatin folding and human DNA replication. We previously described megabase replication N/U-domains as mammalian multiorigin replication units, and showed that their borders are ‘master’ replication initiation zones that likely initiate cascades of origin firing responsible for the stereotypic replication of these domains. Here, we demonstrate that replication N/U-domains correspond to the structural domains of self-interacting chromatin, and that their borders act as insulating regions both in high-throughput 3C (Hi-C) data and high-resolution 3C (4C) experiments. Further analyses of Hi-C data using a graph-theoretical approach reveal that N/U-domain borders are long-distance, interconnected hubs of the chromatin interaction network. Overall, these results and the observation that a well-defined ordering of chromatin states exists from N/U-domain borders to centers suggest that ‘master’ replication initiation zones are at the heart of a high-order, epigenetically controlled 3D organization of the human genome. (paper)

  9. Molecular Dynamics Simulations of the STAS Domains of Rat Prestin and Human Pendrin Reveal Conformational Motions in Conserved Flexible Regions

    Directory of Open Access Journals (Sweden)

    Alok K. Sharma

    2014-02-01

    Full Text Available Background: Molecular dynamics (MD simulations provide valuable information on the conformational changes that accompany time-dependent motions in proteins. The reported crystal structure of rat prestin (PDB 3LLO is remarkable for an α1-α2 inter-helical angle that differs substantially from those observed in bacterial STAS domains of SulP anion transporters and anti-sigma factor antagonists. However, NMR data on the rat prestin STAS domain in solution suggests dynamic features at or near the α1-α2 helical region (Pasqualetto et al JMB, 2010. We therefore performed a 100 ns 300K MD simulation study comparing the STAS domains of rat prestin and (modeled human pendrin, to explore possible conformational flexibility in the region of the α1 and α2 helices. Methods: The conformation of the loop missing in the crystal structure of rat prestin STAS (11 amino acids between helix α1 and strand β3 was built using Modeller. MD simulations were performed with GROMACSv4.6 using GROMOS96 53a6 all-atom force field. Results: A subset of secondary structured elements of the STAS domains exhibits significant conformational changes during the simulation time course. The conformationally perturbed segments include the majority of loop regions, as well as the α1 and α2 helices. A significant decrease in the α1-α2 inter-helical angle observed across the simulation trajectory leads to closer helical packing at their C-termini. The end-simulation conformations of the prestin and pendrin STAS domains, including their decreased α1-α2 inter-helical angles, resemble more closely the packing of corresponding helices in the STAS structures of bacterial SulP transporters Rv1739c and ychM, as well as those of the anti-sigma factor antagonists. Several structural segments of the modeled human pendrin STAS domain exhibit larger atomic motions and greater conformational deviations than the corresponding regions of rat prestin, predicting that the human pendrin STAS

  10. The Human Escort Protein Hep Binds to the ATPase Domain of Mitochondrial Hsp70 and Regulates ATP Hydrolysis*

    Science.gov (United States)

    Zhai, Peng; Stanworth, Crystal; Liu, Shirley; Silberg, Jonathan J.

    2008-01-01

    Hsp70 escort proteins (Hep) have been implicated as essential for maintaining the function of yeast mitochondrial hsp70 molecular chaperones (mtHsp70), but the role that escort proteins play in regulating mammalian chaperone folding and function has not been established. We present evidence that human mtHsp70 exhibits limited solubility due to aggregation mediated by its ATPase domain and show that human Hep directly enhances chaperone solubility through interactions with this domain. In the absence of Hep, mtHsp70 was insoluble when expressed in Escherichia coli, as was its isolated ATPase domain and a chimera having this domain fused to the peptide-binding domain of HscA, a soluble monomeric chaperone. In contrast, these proteins all exhibited increased solubility when expressed in the presence of Hep. In vitro studies further revealed that purified Hep regulates the interaction of mtHsp70 with nucleotides. Full-length mtHsp70 exhibited slow intrinsic ATP hydrolysis activity (6.8 ± 0.2 × 10-4 s-1) at 25 °C, which was stimulated up to 49-fold by Hep. Hep also stimulated the activity of the isolated ATPase domain, albeit to a lower maximal extent (11.5-fold). In addition, gel-filtration studies showed that formation of chaperone-escort protein complexes inhibited mtHsp70 self-association, and they revealed that Hep binding to full-length mtHsp70 and its isolated ATPase domain is strongest in the absence of nucleotides. These findings provide evidence that metazoan escort proteins regulate the catalytic activity and solubility of their cognate chaperones, and they indicate that both forms of regulation arise from interactions with the mtHsp70 ATPase domain. PMID:18632665

  11. The human escort protein Hep binds to the ATPase domain of mitochondrial hsp70 and regulates ATP hydrolysis.

    Science.gov (United States)

    Zhai, Peng; Stanworth, Crystal; Liu, Shirley; Silberg, Jonathan J

    2008-09-19

    Hsp70 escort proteins (Hep) have been implicated as essential for maintaining the function of yeast mitochondrial hsp70 molecular chaperones (mtHsp70), but the role that escort proteins play in regulating mammalian chaperone folding and function has not been established. We present evidence that human mtHsp70 exhibits limited solubility due to aggregation mediated by its ATPase domain and show that human Hep directly enhances chaperone solubility through interactions with this domain. In the absence of Hep, mtHsp70 was insoluble when expressed in Escherichia coli, as was its isolated ATPase domain and a chimera having this domain fused to the peptide-binding domain of HscA, a soluble monomeric chaperone. In contrast, these proteins all exhibited increased solubility when expressed in the presence of Hep. In vitro studies further revealed that purified Hep regulates the interaction of mtHsp70 with nucleotides. Full-length mtHsp70 exhibited slow intrinsic ATP hydrolysis activity (6.8+/-0.2 x 10(-4) s(-1)) at 25 degrees C, which was stimulated up to 49-fold by Hep. Hep also stimulated the activity of the isolated ATPase domain, albeit to a lower maximal extent (11.5-fold). In addition, gel-filtration studies showed that formation of chaperone-escort protein complexes inhibited mtHsp70 self-association, and they revealed that Hep binding to full-length mtHsp70 and its isolated ATPase domain is strongest in the absence of nucleotides. These findings provide evidence that metazoan escort proteins regulate the catalytic activity and solubility of their cognate chaperones, and they indicate that both forms of regulation arise from interactions with the mtHsp70 ATPase domain.

  12. MicroCT evaluation of bone mineral density loss in human bones

    International Nuclear Information System (INIS)

    Nogueira, Liebert P.; Braz, Delson; Lopes, Ricardo T.; Barroso, Regina C.; Oliveira, Luis F.

    2007-01-01

    Bone is a connective tissue largely composed of an organic protein, collagen and the inorganic mineral hydroxyapatite [Ca 10 (PO 4 ) 6 OH 2 ], which combine to provide a mechanical and supportive role in the body. Depending on the orientation of collagen fibers, two types of bone can be distinguished: trabecular and cortical bone. Degree of mineralization is considered an important feature of bone quality. Changes in the degree of mineralization is generally due to osteoporosis, but many recent studies have already shown that alterations in degree of mineralization can occur due to a large variety of factors. The transmission X-ray microtomography is one of the most popular methods, which provides the spatial distribution of the total absorption coefficient inside the sample. The aim of this study was to investigate the suitability of using microCT as a supplementary tool for the diagnosis of the health status of human bones. Eleven samples were constructed simulating the physiological range of bone mineral density (BMD) found in cortical human bone. The samples represent healthy mixtures of swine compact bone dried at room temperature, powdered and mixed with fat (0 - 100 % by mass). The samples were imaged by a microfocus tube (Fein-Focus) with focal size of about 60 μm (±5%), and a CCD camera (0.143 mm pixel size) coupled with an intensifier tube with fluoroscope screen at the Nuclear Instrumentation Laboratory (COPPE/UFRJ), Brazil. The images were reconstructed and treated with suitable software developed at the Nuclear Instrumentation Laboratory. The mineral content in cortical bone is defined by the volume of dry, fat-free bone per unit bulk volume of the bone. The volumes were calculated from the bone density using the relationship between volume and density. The densities of fat and bone were taken to be 0.95 g.cm -3 and 1.92 g.cm -3 respectively. The correlation of the measured absorption coefficient with the mineral content in the samples was then

  13. MicroCT evaluation of bone mineral density loss in human bones

    Energy Technology Data Exchange (ETDEWEB)

    Nogueira, Liebert P.; Braz, Delson; Lopes, Ricardo T. [Universidade Federal do Rio de Janeiro (UFRJ), RJ (Brazil). Coordenacao dos Programas de Pos-graduacao de Engenharia (COPPE). Lab. de Instrumentacao Nuclear]. E-mails: lnogueira@con.ufrj.br; Barroso, Regina C.; Oliveira, Luis F. [Universidade do Estado do Rio de Janeiro (UERJ), Rio de Janeiro, RJ (Brazil). Inst. de Fisica]. E-mail: cely@uerj.br

    2007-07-01

    Bone is a connective tissue largely composed of an organic protein, collagen and the inorganic mineral hydroxyapatite [Ca{sub 10}(PO{sub 4}){sub 6}OH{sub 2}], which combine to provide a mechanical and supportive role in the body. Depending on the orientation of collagen fibers, two types of bone can be distinguished: trabecular and cortical bone. Degree of mineralization is considered an important feature of bone quality. Changes in the degree of mineralization is generally due to osteoporosis, but many recent studies have already shown that alterations in degree of mineralization can occur due to a large variety of factors. The transmission X-ray microtomography is one of the most popular methods, which provides the spatial distribution of the total absorption coefficient inside the sample. The aim of this study was to investigate the suitability of using microCT as a supplementary tool for the diagnosis of the health status of human bones. Eleven samples were constructed simulating the physiological range of bone mineral density (BMD) found in cortical human bone. The samples represent healthy mixtures of swine compact bone dried at room temperature, powdered and mixed with fat (0 - 100 % by mass). The samples were imaged by a microfocus tube (Fein-Focus) with focal size of about 60 {mu}m ({+-}5%), and a CCD camera (0.143 mm pixel size) coupled with an intensifier tube with fluoroscope screen at the Nuclear Instrumentation Laboratory (COPPE/UFRJ), Brazil. The images were reconstructed and treated with suitable software developed at the Nuclear Instrumentation Laboratory. The mineral content in cortical bone is defined by the volume of dry, fat-free bone per unit bulk volume of the bone. The volumes were calculated from the bone density using the relationship between volume and density. The densities of fat and bone were taken to be 0.95 g.cm{sup -3} and 1.92 g.cm{sup -3} respectively. The correlation of the measured absorption coefficient with the mineral content

  14. PET/CT imaging of human somatostatin receptor 2 (hsstr2) as reporter gene for gene therapy

    International Nuclear Information System (INIS)

    Hofmann, M.; Gazdhar, A.; Weitzel, T.; Schmid, R.; Krause, T.

    2006-01-01

    Localized information on region-selective gene expression in small animals is widely obtained by use of reporter genes inducing light emission. Using these reporter genes for imaging deep inside the human body fluorescent probes are hindered by attenuation, scattering and possible fluorescence quenching. This can be overcome by use of radio-peptide receptors as reporter genes. Therefore, the feasibility of the somatostatin receptor 2 expression vector system for expression imaging was checked against a control vector containing luciferase gene. For in vivo transduction of vector DNA into the rat forelimb muscles the in vivo electroporation technique was chosen because of its high regio-selectivity. The gene expression was imaged by high-sensitive CCD camera (luciferase activity) and by PET/CT using a Ga-68-DOTATOC as radio peptide probe. The relative sstr2 expression was enhanced by gene transduction at maximum to a factor of 15. The PET/CT images could be fully quantified. The above demonstrated feasibility of radio-peptide PET/CT reporter gene imaging may serve in the future as a tool for full quantitative understanding of regional gene expression, especially in large animals and humans

  15. PET/CT imaging of human somatostatin receptor 2 (hsstr2) as reporter gene for gene therapy

    Energy Technology Data Exchange (ETDEWEB)

    Hofmann, M. [Molecular Imaging and Therapy Group (MIT-Bern), Clinic of Nuclear Medicine, Inselspital, Medical School Bern (Switzerland)]. E-mail: Michael.Hofmann@insel.ch; Gazdhar, A. [Division of Pulmonary Medicine, University Hospital Bern (Switzerland); Weitzel, T. [Molecular Imaging and Therapy Group (MIT-Bern), Clinic of Nuclear Medicine, Inselspital, Medical School Bern (Switzerland); Schmid, R. [Division of Thoracic Surgery, University Hospital Bern (Switzerland); Krause, T. [Molecular Imaging and Therapy Group (MIT-Bern), Clinic of Nuclear Medicine, Inselspital, Medical School Bern (Switzerland)

    2006-12-20

    Localized information on region-selective gene expression in small animals is widely obtained by use of reporter genes inducing light emission. Using these reporter genes for imaging deep inside the human body fluorescent probes are hindered by attenuation, scattering and possible fluorescence quenching. This can be overcome by use of radio-peptide receptors as reporter genes. Therefore, the feasibility of the somatostatin receptor 2 expression vector system for expression imaging was checked against a control vector containing luciferase gene. For in vivo transduction of vector DNA into the rat forelimb muscles the in vivo electroporation technique was chosen because of its high regio-selectivity. The gene expression was imaged by high-sensitive CCD camera (luciferase activity) and by PET/CT using a Ga-68-DOTATOC as radio peptide probe. The relative sstr2 expression was enhanced by gene transduction at maximum to a factor of 15. The PET/CT images could be fully quantified. The above demonstrated feasibility of radio-peptide PET/CT reporter gene imaging may serve in the future as a tool for full quantitative understanding of regional gene expression, especially in large animals and human000.

  16. First-in-human pilot study of a spatial frequency domain oxygenation imaging system

    Science.gov (United States)

    Gioux, Sylvain; Mazhar, Amaan; Lee, Bernard T.; Lin, Samuel J.; Tobias, Adam M.; Cuccia, David J.; Stockdale, Alan; Oketokoun, Rafiou; Ashitate, Yoshitomo; Kelly, Edward; Weinmann, Maxwell; Durr, Nicholas J.; Moffitt, Lorissa A.; Durkin, Anthony J.; Tromberg, Bruce J.; Frangioni, John V.

    2011-08-01

    Oxygenation measurements are widely used in patient care. However, most clinically available instruments currently consist of contact probes that only provide global monitoring of the patient (e.g., pulse oximetry probes) or local monitoring of small areas (e.g., spectroscopy-based probes). Visualization of oxygenation over large areas of tissue, without a priori knowledge of the location of defects, has the potential to improve patient management in many surgical and critical care applications. In this study, we present a clinically compatible multispectral spatial frequency domain imaging (SFDI) system optimized for surgical oxygenation imaging. This system was used to image tissue oxygenation over a large area (16×12 cm) and was validated during preclinical studies by comparing results obtained with an FDA-approved clinical oxygenation probe. Skin flap, bowel, and liver vascular occlusion experiments were performed on Yorkshire pigs and demonstrated that over the course of the experiment, relative changes in oxygen saturation measured using SFDI had an accuracy within 10% of those made using the FDA-approved device. Finally, the new SFDI system was translated to the clinic in a first-in-human pilot study that imaged skin flap oxygenation during reconstructive breast surgery. Overall, this study lays the foundation for clinical translation of endogenous contrast imaging using SFDI.

  17. Reconstitution of the NF1 GAP-related domain in NF1-deficient human Schwann cells

    International Nuclear Information System (INIS)

    Thomas, Stacey L.; Deadwyler, Gail D.; Tang, Jun; Stubbs, Evan B.; Muir, David; Hiatt, Kelly K.; Clapp, D. Wade; De Vries, George H.

    2006-01-01

    Schwann cells derived from peripheral nerve sheath tumors from individuals with Neurofibromatosis Type 1 (NF1) are deficient for the protein neurofibromin, which contains a GAP-related domain (NF1-GRD). Neurofibromin-deficient Schwann cells have increased Ras activation, increased proliferation in response to certain growth stimuli, increased angiogenic potential, and altered cell morphology. This study examined whether expression of functional NF1-GRD can reverse the transformed phenotype of neurofibromin-deficient Schwann cells from both benign and malignant peripheral nerve sheath tumors. We reconstituted the NF1-GRD using retroviral transduction and examined the effects on cell morphology, growth potential, and angiogenic potential. NF1-GRD reconstitution resulted in morphologic changes, a 16-33% reduction in Ras activation, and a 53% decrease in proliferation in neurofibromin-deficient Schwann cells. However, NF1-GRD reconstitution was not sufficient to decrease the in vitro angiogenic potential of the cells. This study demonstrates that reconstitution of the NF1-GRD can at least partially reverse the transformation of human NF1 tumor-derived Schwann cells

  18. ATRX ADD domain links an atypical histone methylation recognition mechanism to human mental-retardation syndrome

    Energy Technology Data Exchange (ETDEWEB)

    Iwase, Shigeki; Xiang, Bin; Ghosh, Sharmistha; Ren, Ting; Lewis, Peter W.; Cochrane, Jesse C.; Allis, C. David; Picketts, David J.; Patel, Dinshaw J.; Li, Haitao; Shi, Yang (Harvard-Med); (Ottawa Hosp.); (MSKCC); (Rockefeller); (CH-Boston); (Tsinghua); (Mass. Gen. Hosp.)

    2011-07-19

    ATR-X (alpha-thalassemia/mental retardation, X-linked) syndrome is a human congenital disorder that causes severe intellectual disabilities. Mutations in the ATRX gene, which encodes an ATP-dependent chromatin-remodeler, are responsible for the syndrome. Approximately 50% of the missense mutations in affected persons are clustered in a cysteine-rich domain termed ADD (ATRX-DNMT3-DNMT3L, ADD{sub ATRX}), whose function has remained elusive. Here we identify ADD{sub ATRX} as a previously unknown histone H3-binding module, whose binding is promoted by lysine 9 trimethylation (H3K9me3) but inhibited by lysine 4 trimethylation (H3K4me3). The cocrystal structure of ADD{sub ATRX} bound to H3{sub 1-15}K9me3 peptide reveals an atypical composite H3K9me3-binding pocket, which is distinct from the conventional trimethyllysine-binding aromatic cage. Notably, H3K9me3-pocket mutants and ATR-X syndrome mutants are defective in both H3K9me3 binding and localization at pericentromeric heterochromatin; thus, we have discovered a unique histone-recognition mechanism underlying the ATR-X etiology.

  19. ATRX ADD Domain Links an Atypical Histone Methylation Recognition Mechanism to Human Mental-Retardation Syndrome

    Energy Technology Data Exchange (ETDEWEB)

    S Iwase; B Xiang; S Ghosh; T Ren; P Lewis; J Cochrane; C Allis; D Picketts; D Patel; et al.

    2011-12-31

    ATR-X (alpha-thalassemia/mental retardation, X-linked) syndrome is a human congenital disorder that causes severe intellectual disabilities. Mutations in the ATRX gene, which encodes an ATP-dependent chromatin-remodeler, are responsible for the syndrome. Approximately 50% of the missense mutations in affected persons are clustered in a cysteine-rich domain termed ADD (ATRX-DNMT3-DNMT3L, ADD{sub ATRX}), whose function has remained elusive. Here we identify ADD{sub ATRX} as a previously unknown histone H3-binding module, whose binding is promoted by lysine 9 trimethylation (H3K9me3) but inhibited by lysine 4 trimethylation (H3K4me3). The cocrystal structure of ADD{sub ATRX} bound to H3{sub 1-15}K9me3 peptide reveals an atypical composite H3K9me3-binding pocket, which is distinct from the conventional trimethyllysine-binding aromatic cage. Notably, H3K9me3-pocket mutants and ATR-X syndrome mutants are defective in both H3K9me3 binding and localization at pericentromeric heterochromatin; thus, we have discovered a unique histone-recognition mechanism underlying the ATR-X etiology.

  20. Structure, functional characterization, and evolution of the dihydroorotase domain of human CAD.

    Science.gov (United States)

    Grande-García, Araceli; Lallous, Nada; Díaz-Tejada, Celsa; Ramón-Maiques, Santiago

    2014-02-04

    Upregulation of CAD, the multifunctional protein that initiates and controls the de novo biosynthesis of pyrimidines in animals, is essential for cell proliferation. Deciphering the architecture and functioning of CAD is of interest for its potential usage as an antitumoral target. However, there is no detailed structural information about CAD other than that it self-assembles into hexamers of ∼1.5 MDa. Here we report the crystal structure and functional characterization of the dihydroorotase domain of human CAD. Contradicting all assumptions, the structure reveals an active site enclosed by a flexible loop with two Zn²⁺ ions bridged by a carboxylated lysine and a third Zn coordinating a rare histidinate ion. Site-directed mutagenesis and functional assays prove the involvement of the Zn and flexible loop in catalysis. Comparison with homologous bacterial enzymes supports a reclassification of the DHOase family and provides strong evidence against current models of the architecture of CAD. Copyright © 2014 Elsevier Ltd. All rights reserved.

  1. Human phenotypically distinct TGFBI corneal dystrophies are linked to the stability of the fourth FAS1 domain of TGFBIp

    DEFF Research Database (Denmark)

    Runager, Kasper; Basaiawmoit, Rajiv Vaid; Deva, Taru

    2011-01-01

    Mutations in the human TGFBI gene encoding TGFBIp have been linked to protein deposits in the cornea leading to visual impairment. The protein consists of an N-terminal Cys-rich EMI domain and four consecutive fasciclin 1 (FAS1) domains. We have compared the stabilities of wild-type (WT) human...... TGFBIp and six mutants known to produce phenotypically distinct deposits in the cornea. Amino acid substitutions in the first FAS1 (FAS1-1) domain (R124H, R124L, and R124C) did not alter the stability. However, substitutions within the fourth FAS1 (FAS1-4) domain (A546T, R555Q, and R555W) affected...... the overall stability of intact TGFBIp revealing the following stability ranking R555W>WT>R555Q>A546T. Significantly, the stability ranking of the isolated FAS1-4 domains mirrored the behavior of the intact protein. In addition, it was linked to the aggregation propensity as the least stable mutant (A546T...

  2. A New Method for 3D Finite Element Modeling of Human Mandible Based on CT Data

    Institute of Scientific and Technical Information of China (English)

    于力牛; 叶铭; 王成焘

    2004-01-01

    This study presents a reliable method for the semi-automatic generation of an FE model, which determines both geometrical data and bone properties from patient CT scans.3D FE analysis is one of the best approaches to predict the stress and strain distribution in complex bone structures, but its accuracy strongly depends on the precision of input information. In geometric reconstruction, various methods of image processing, geometric modeling and finite element analysis are combined and extended. Emphasis is given to the assignment of the material properties based on the density values computed from CT data. Through this technique, the model with high geometric and material similarities were generated in an easy way. Consequently, the patient-specific FE model from mandible CT data is realized also.

  3. Inner images of the human body with a 3D CT scanner

    International Nuclear Information System (INIS)

    Kobayashi, Hisashi

    1994-01-01

    This article deals with not only CT-endoscopy (CTES) technique but also various imaging and processing techniques of 3D CT. CTES images, which were obtained from 137 patients with suspected cardiovascular disorder or disease of other tubular organs, were reconstructed using a newly developed volumetric scanner with a slip-ring system. Among the 137 patients, 107 (78%) were successfully diagnosed by CTES. For cardiovascular region, dissecting aneurysm was detected in 27/32, aortitis in 9/9, and intra-arterial thrombosis in 5/6. Various imaging and processing techniques, including CT number conversion technique, multi-threshold range imaging, 'open-window' and 'virtual operation', and long segmental arteriogram by intravenous contrast injection, are displayed in futures. In conclusion, CTES might become a safe and minimally invasive means for observing the inner surface of the tubular organs, particularly of the aorta, without the need of fiberscopic manipulation. (N.K.)

  4. Refolding and characterization of the functional ligand-binding domain of human lectin-like oxidized LDL receptor.

    Science.gov (United States)

    Xie, Qiuhong; Matsunaga, Shigeru; Shi, Xiaohua; Ogawa, Setsuko; Niimi, Setsuko; Wen, Zhesheng; Tokuyasu, Ken; Machida, Sachiko

    2003-11-01

    Lectin-like oxidized low-density lipoprotein receptor (LOX-1), a type II membrane protein that can recognize a variety of structurally unrelated macromolecules, plays an important role in host defense and is implicated in atherogenesis. To understand the interaction between human LOX-1 and its ligands, in this study the functional C-type lectin-like domain (CTLD) of LOX-1 was reconstituted at high efficiency from inactive aggregates in Escherichia coli using a refolding technique based on an artificial chaperone. The CD spectra of the purified domain suggested that the domain has alpha-helical structure and the blue shift of Trp residues was observed on refolding of the domain. Like wild-type hLOX-1, the refolded CTLD domain was able to bind modified LDL. Thus, even though CTLD contains six Cys residues that form disulfide bonds, it recovered its specific binding ability on refolding. This suggests that the correct disulfide bonds in CTLD were formed by the artificial chaperone technique. Although the domain lacked N-glycosylation, it showed high affinity for its ligand in surface plasmon resonance experiments. Thus, unglycosylated CTLD is sufficient for binding modified LDL.

  5. Crystallization and preliminary crystallographic analysis of the fourth FAS1 domain of human BigH3

    International Nuclear Information System (INIS)

    Yoo, Ji-Ho; Kim, EungKweon; Kim, Jongsun; Cho, Hyun-Soo

    2007-01-01

    The crystallization and X-ray diffraction analysis of the fourth FAS1 domain of human BigH3 are reported. The protein BigH3 is a cell-adhesion molecule induced by transforming growth factor-β (TGF-β). It consists of four homologous repeat domains known as FAS1 domains; mutations in these domains have been linked to corneal dystrophy. The fourth FAS1 domain was expressed in Escherichia coli B834 (DE3) (a methionine auxotroph) and purified by DEAE anion-exchange and gel-filtration chromatography. The FAS1 domain was crystallized using the vapour-diffusion method. A SAD diffraction data set was collected to a resolution of 2.5 Å at 100 K. The crystal belonged to space group P6 1 or P6 5 and had two molecules per asymmetric unit, with unit-cell parameters a = b = 62.93, c = 143.27 Å, α = β = 90.0, γ = 120.0°

  6. Crystal structure of the second fibronectin type III (FN3) domain from human collagen α1 type XX.

    Science.gov (United States)

    Zhao, Jingfeng; Ren, Jixia; Wang, Nan; Cheng, Zhong; Yang, Runmei; Lin, Gen; Guo, Yi; Cai, Dayong; Xie, Yong; Zhao, Xiaohong

    2017-12-01

    Collagen α1 type XX, which contains fibronectin type III (FN3) repeats involving six FN3 domains (referred to as the FN#1-FN#6 domains), is an unusual member of the fibril-associated collagens with interrupted triple helices (FACIT) subfamily of collagens. The results of standard protein BLAST suggest that the FN3 repeats might contribute to collagen α1 type XX acting as a cytokine receptor. To date, solution NMR structures of the FN#3, FN#4 and FN#6 domains have been determined. To obtain further structural evidence to understand the relationship between the structure and function of the FN3 repeats from collagen α1 type XX, the crystal structure of the FN#2 domain from human collagen α1 type XX (residues Pro386-Pro466; referred to as FN2-HCXX) was solved at 2.5 Å resolution. The crystal structure of FN2-HCXX shows an immunoglobulin-like fold containing a β-sandwich structure, which is formed by a three-stranded β-sheet (β1, β2 and β5) packed onto a four-stranded β-sheet (β3, β4, β6 and β7). Two consensus domains, tencon and fibcon, are structural analogues of FN2-HCXX. Fn8, an FN3 domain from human oncofoetal fibronectin, is the closest structural analogue of FN2-HCXX derived from a naturally occurring sequence. Based solely on the structural similarity of FN2-HCXX to other FN3 domains, the detailed functions of FN2-HCXX and the FN3 repeats in collagen α1 type XX cannot be identified.

  7. De novo identification of replication-timing domains in the human genome by deep learning.

    Science.gov (United States)

    Liu, Feng; Ren, Chao; Li, Hao; Zhou, Pingkun; Bo, Xiaochen; Shu, Wenjie

    2016-03-01

    The de novo identification of the initiation and termination zones-regions that replicate earlier or later than their upstream and downstream neighbours, respectively-remains a key challenge in DNA replication. Building on advances in deep learning, we developed a novel hybrid architecture combining a pre-trained, deep neural network and a hidden Markov model (DNN-HMM) for the de novo identification of replication domains using replication timing profiles. Our results demonstrate that DNN-HMM can significantly outperform strong, discriminatively trained Gaussian mixture model-HMM (GMM-HMM) systems and other six reported methods that can be applied to this challenge. We applied our trained DNN-HMM to identify distinct replication domain types, namely the early replication domain (ERD), the down transition zone (DTZ), the late replication domain (LRD) and the up transition zone (UTZ), using newly replicated DNA sequencing (Repli-Seq) data across 15 human cells. A subsequent integrative analysis revealed that these replication domains harbour unique genomic and epigenetic patterns, transcriptional activity and higher-order chromosomal structure. Our findings support the 'replication-domain' model, which states (1) that ERDs and LRDs, connected by UTZs and DTZs, are spatially compartmentalized structural and functional units of higher-order chromosomal structure, (2) that the adjacent DTZ-UTZ pairs form chromatin loops and (3) that intra-interactions within ERDs and LRDs tend to be short-range and long-range, respectively. Our model reveals an important chromatin organizational principle of the human genome and represents a critical step towards understanding the mechanisms regulating replication timing. Our DNN-HMM method and three additional algorithms can be freely accessed at https://github.com/wenjiegroup/DNN-HMM The replication domain regions identified in this study are available in GEO under the accession ID GSE53984. shuwj@bmi.ac.cn or boxc

  8. Crystal structure of a human single domain antibody dimer formed through V(H-V(H non-covalent interactions.

    Directory of Open Access Journals (Sweden)

    Toya Nath Baral

    Full Text Available Single-domain antibodies (sdAbs derived from human V(H are considered to be less soluble and prone to aggregate which makes it difficult to determine the crystal structures. In this study, we isolated and characterized two anti-human epidermal growth factor receptor-2 (HER2 sdAbs, Gr3 and Gr6, from a synthetic human V(H phage display library. Size exclusion chromatography and surface plasmon resonance analyses demonstrated that Gr3 is a monomer, but that Gr6 is a strict dimer. To understand this different molecular behavior, we solved the crystal structure of Gr6 to 1.6 Å resolution. The crystal structure revealed that the homodimer assembly of Gr6 closely mimics the V(H-V(L heterodimer of immunoglobulin variable domains and the dimerization interface is dominated by hydrophobic interactions.

  9. Functional interactions of the AF-2 activation domain core region of the human androgen receptor with the amino-terminal domain and with the transcriptional coactivator TIF2 (transcriptional intermediary factor2)

    NARCIS (Netherlands)

    C.A. Berrevoets (Cor); P. Doesburg (Paul); K. Steketee (Karine); J. Trapman (Jan); A.O. Brinkmann (Albert)

    1998-01-01

    textabstractPrevious studies in yeast and mammalian cells showed a functional interaction between the amino-terminal domain and the carboxy-terminal, ligand-binding domain (LBD) of the human androgen receptor (AR). In the present study, the AR subdomains involved in

  10. Targeted DNA demethylation of the Arabidopsis genome using the human TET1 catalytic domain

    Science.gov (United States)

    Gallego-Bartolomé, Javier; Gardiner, Jason; Liu, Wanlu; Papikian, Ashot; Ghoshal, Basudev; Kuo, Hsuan Yu; Zhao, Jenny Miao-Chi; Jacobsen, Steven E.

    2018-01-01

    DNA methylation is an important epigenetic modification involved in gene regulation and transposable element silencing. Changes in DNA methylation can be heritable and, thus, can lead to the formation of stable epialleles. A well-characterized example of a stable epiallele in plants is fwa, which consists of the loss of DNA cytosine methylation (5mC) in the promoter of the FLOWERING WAGENINGEN (FWA) gene, causing up-regulation of FWA and a heritable late-flowering phenotype. Here we demonstrate that a fusion between the catalytic domain of the human demethylase TEN-ELEVEN TRANSLOCATION1 (TET1cd) and an artificial zinc finger (ZF) designed to target the FWA promoter can cause highly efficient targeted demethylation, FWA up-regulation, and a heritable late-flowering phenotype. Additional ZF–TET1cd fusions designed to target methylated regions of the CACTA1 transposon also caused targeted demethylation and changes in expression. Finally, we have developed a CRISPR/dCas9-based targeted demethylation system using the TET1cd and a modified SunTag system. Similar to the ZF–TET1cd fusions, the SunTag–TET1cd system is able to target demethylation and activate gene expression when directed to the FWA or CACTA1 loci. Our study provides tools for targeted removal of 5mC at specific loci in the genome with high specificity and minimal off-target effects. These tools provide the opportunity to develop new epialleles for traits of interest, and to reactivate expression of previously silenced genes, transgenes, or transposons. PMID:29444862

  11. Optimized energy of spectral CT for infarct imaging: Experimental validation with human validation.

    Science.gov (United States)

    Sandfort, Veit; Palanisamy, Srikanth; Symons, Rolf; Pourmorteza, Amir; Ahlman, Mark A; Rice, Kelly; Thomas, Tom; Davies-Venn, Cynthia; Krauss, Bernhard; Kwan, Alan; Pandey, Ankur; Zimmerman, Stefan L; Bluemke, David A

    Late contrast enhancement visualizes myocardial infarction, but the contrast to noise ratio (CNR) is low using conventional CT. The aim of this study was to determine if spectral CT can improve imaging of myocardial infarction. A canine model of myocardial infarction was produced in 8 animals (90-min occlusion, reperfusion). Later, imaging was performed after contrast injection using CT at 90 kVp/150 kVpSn. The following reconstructions were evaluated: Single energy 90 kVp, mixed, iodine map, multiple monoenergetic conventional and monoenergetic noise optimized reconstructions. Regions of interest were measured in infarct and remote regions to calculate contrast to noise ratio (CNR) and Bhattacharya distance (a metric of the differentiation between regions). Blinded assessment of image quality was performed. The same reconstruction methods were applied to CT scans of four patients with known infarcts. For animal studies, the highest CNR for infarct vs. myocardium was achieved in the lowest keV (40 keV) VMo images (CNR 4.42, IQR 3.64-5.53), which was superior to 90 kVp, mixed and iodine map (p = 0.008, p = 0.002, p energy in conjunction with noise-optimized monoenergetic post-processing improves CNR of myocardial infarct delineation by approximately 20-25%. Published by Elsevier Inc.

  12. Volumetry of human molars with flat panel-based volume CT in vitro

    NARCIS (Netherlands)

    Hannig, C.; Krieger, E.; Dullin, C.; Merten, H.A.; Attin, T.; Grabbe, E.; Heidrich, G.

    2006-01-01

    The flat panel-based volume computed tomography (fpVCT) is a new CT device applicable for experimental, three-dimensional evaluation of teeth at a resolution of about 150 microm in the high contrast region. The aim of this study was to investigate whether fpVCT was suitable for quantification of the

  13. Detection of small traumatic hemorrhages using a computer-generated average human brain CT.

    Science.gov (United States)

    Afzali-Hashemi, Liza; Hazewinkel, Marieke; Tjepkema-Cloostermans, Marleen C; van Putten, Michel J A M; Slump, Cornelis H

    2018-04-01

    Computed tomography is a standard diagnostic imaging technique for patients with traumatic brain injury (TBI). A limitation is the poor-to-moderate sensitivity for small traumatic hemorrhages. A pilot study using an automatic method to detect hemorrhages [Formula: see text] in diameter in patients with TBI is presented. We have created an average image from 30 normal noncontrast CT scans that were automatically aligned using deformable image registration as implemented in Elastix software. Subsequently, the average image was aligned to the scans of TBI patients, and the hemorrhages were detected by a voxelwise subtraction of the average image from the CT scans of nine TBI patients. An experienced neuroradiologist and a radiologist in training assessed the presence of hemorrhages in the final images and determined the false positives and false negatives. The 9 CT scans contained 67 small haemorrhages, of which 97% was correctly detected by our system. The neuroradiologist detected three false positives, and the radiologist in training found two false positives. For one patient, our method showed a hemorrhagic contusion that was originally missed. Comparing individual CT scans with a computed average may assist the physicians in detecting small traumatic hemorrhages in patients with TBI.

  14. 18F FDG Uptake of Human Testis on PET/CT: Correlation with Age, Sex Hormones, and Vasectomy

    International Nuclear Information System (INIS)

    Moon, Seung Hwan; Eo, Jae Sun; Lee, Jong Jin; Chung, June Key; Lee, Dong Soo; Lee, Myung Chul

    2011-01-01

    The purpose of this study was to evaluate glucose metabolism of normal human testis on 18F FDG PET/CT and to assess possible correlation among age, the serum levels of sex hormones, and vasectomy. 18F FDG PET/CT was performed in 66 normal healthy men (50.8±13.6 years, range 22-81), and mean standard uptake values (SUV) of 18F FDG in testis and adductor muscle were measured. Testis muscle SUV ratios (T/M ratios) were calculated. Serum levels of total testosterone, free testosterone, estradiol, and of sex hormone binding globulin (SHBG) were measured. We searched for correlations between T/M ratios and age and the serum concentrations of sex hormones. 18F FDG PET/CT was also performed in 32 vasectomized men (55.7±7.8 years, range 38-71) and 52 nonvasectomized men (55.4±11.6 years, range 37-72). Mean SUVs of testis and adductor muscle were measured, and T/M ratios were calculated. A significant age related decline was found in T/M ratio (r=-0.509, p 18F FDG uptake may have attributed to testicular function and testicular histology. Our findings may have important implications for the interpretation of testicular 18F FDG uptake in the normal adult population.

  15. Morphometric anatomical and CT study of the human adult sacroiliac region.

    Science.gov (United States)

    Postacchini, Roberto; Trasimeni, Guido; Ripani, Francesca; Sessa, Pasquale; Perotti, Stefano; Postacchini, Franco

    2017-01-01

    To identify and describe the morphometry and CT features of the articular and extra-articular portions of the sacroiliac region. The resulting knowledge might help to avoid complications in sacroiliac joint (SIJ) fusion. We analyzed 102 dry hemi-sacra, 80 ilia, and 10 intact pelves and assessed the pelvic computerized tomography (CT) scans of 90 patients, who underwent the examination for conditions not involving the pelvis. We assessed both the posterior aspect of sacrum with regard to the depressions located externally to the lateral sacral crest at the level of the proximal three sacral vertebrae and the posteroinferior aspect of ilium. Coronal and axial CT scans of the SIJ of patients were obtained and the joint space was measured. On each side, the sacrum exhibits three bone depressions, not described in anatomic textbooks or studies, facing the medial aspect of the posteroinferior ilium, not yet described in detail. Both structures are extra-articular portions situated posteriorly to the SIJ. Coronal CT scans of patients showing the first three sacral foramens and the interval between sacrum and ilium as a continuous space display only the S1 and S3 portions of SIJ, the intermediate portion being extra-articular. The S2 portion is visible on the most anterior coronal scan. Axial scans show articular and extra-articular portions and features improperly described as anatomic variations. Extra-articular portions of the sacroiliac region, not yet described exhaustively, have often been confused with SIJ. Coronal CT scans through the middle part of sacrum, the most used to evaluate degenerative and inflammatory conditions of SIJ, show articular and extra-articular portions of the region.

  16. Solution structure of the first SH3 domain of human vinexin and its interaction with vinculin peptides

    International Nuclear Information System (INIS)

    Zhang, Jiahai; Li, Xiang; Yao, Bo; Shen, Weiqun; Sun, Hongbin; Xu, Chao; Wu, Jihui; Shi, Yunyu

    2007-01-01

    Solution structure of the first Src homology (SH) 3 domain of human vinexin (V S H3 1 ) was determined using nuclear magnetic resonance (NMR) method and revealed that it was a canonical SH3 domain, which has a typical β-β-β-β-α-β fold. Using chemical shift perturbation and surface plasmon resonance experiments, we studied the binding properties of the SH3 domain with two different peptides from vinculin hinge regions: P856 and P868. The observations illustrated slightly different affinities of the two peptides binding to V S H3 1 . The interaction between P868 and V S H3 1 belonged to intermediate exchange with a modest binding affinity, while the interaction between P856 and V S H3 1 had a low binding affinity. The structure and ligand-binding interface of V S H3 1 provide a structural basis for the further functional study of this important molecule

  17. Crystal structures of the human G3BP1 NTF2-like domain visualize FxFG Nup Repeat Specificity

    DEFF Research Database (Denmark)

    Vognsen, Tina Reinholdt; Möller, Ingvar Rúnar; Kristensen, Ole

    2013-01-01

    Ras GTPase Activating Protein SH3 Domain Binding Protein (G3BP) is a potential anti-cancer drug target implicated in several cellular functions. We have used protein crystallography to solve crystal structures of the human G3BP1 NTF2-like domain both alone and in complex with an FxFG Nup repeat...... peptide. Despite high structural similarity, the FxFG binding site is located between two alpha helices in the G3BP1 NTF2-like domain and not at the dimer interface as observed for nuclear transport factor 2. ITC studies showed specificity towards the FxFG motif but not FG and GLFG motifs. The unliganded...

  18. Connecting art and science: An interdisciplinary strategy and its impact on the affective domain of community college human anatomy students

    Science.gov (United States)

    Petti, Kevin

    Educational objectives are often described within the framework of a three-domain taxonomy: cognitive, affective and psychomotor. While most of the research on educational objectives has focused on the cognitive domain, the research that has been conducted on the affective domain, which speaks to emotions, attitudes, and values, has identified a number of positive outcomes. One approach to enhancing the affective domain is that of interdisciplinary education. Science education research in the realm of interdisciplinary education and affective outcomes is limited; especially research conducted on community college students of human anatomy. This project investigated the relationship between an interdisciplinary teaching strategy and the affective domain in science education by utilizing an interdisciplinary lecture in a human anatomy class. Subjects were anatomy students in a California community college who listened to a one-hour lecture describing the cultural, historical and scientific significance of selected pieces of art depicting human dissection in European medieval and Renaissance universities. The focus was on how these renderings represent the state of anatomy education during their respective eras. After listening to the lecture, subjects were administered a 35-question survey that was composed of 14 demographic questions and 21 Likert-style statements that asked respondents to rate the extent to which the intervention influenced their affective domain. Descriptive statistics were then used to determine which component of the affective domain was most influenced, and multiple regression analysis was used to examine the extent to which individual differences along the affective continuum were explained by select demographic measures such as gender, race/ethnicity, education level, and previous exposure to science courses. Results indicate that the interdisciplinary intervention had a positive impact on every component of the affective domain hierarchy

  19. A new technique to characterize CT scanner bow-tie filter attenuation and applications in human cadaver dosimetry simulations

    Science.gov (United States)

    Li, Xinhua; Shi, Jim Q.; Zhang, Da; Singh, Sarabjeet; Padole, Atul; Otrakji, Alexi; Kalra, Mannudeep K.; Xu, X. George; Liu, Bob

    2015-01-01

    Purpose: To present a noninvasive technique for directly measuring the CT bow-tie filter attenuation with a linear array x-ray detector. Methods: A scintillator based x-ray detector of 384 pixels, 307 mm active length, and fast data acquisition (model X-Scan 0.8c4-307, Detection Technology, FI-91100 Ii, Finland) was used to simultaneously detect radiation levels across a scan field-of-view. The sampling time was as short as 0.24 ms. To measure the body bow-tie attenuation on a GE Lightspeed Pro 16 CT scanner, the x-ray tube was parked at the 12 o’clock position, and the detector was centered in the scan field at the isocenter height. Two radiation exposures were made with and without the bow-tie in the beam path. Each readout signal was corrected for the detector background offset and signal-level related nonlinear gain, and the ratio of the two exposures gave the bow-tie attenuation. The results were used in the geant4 based simulations of the point doses measured using six thimble chambers placed in a human cadaver with abdomen/pelvis CT scans at 100 or 120 kV, helical pitch at 1.375, constant or variable tube current, and distinct x-ray tube starting angles. Results: Absolute attenuation was measured with the body bow-tie scanned at 80–140 kV. For 24 doses measured in six organs of the cadaver, the median or maximum difference between the simulation results and the measurements on the CT scanner was 8.9% or 25.9%, respectively. Conclusions: The described method allows fast and accurate bow-tie filter characterization. PMID:26520720

  20. Spatio-temporal re-organization of replication foci accompanies replication domain consolidation during human pluripotent stem cell lineage specification

    Science.gov (United States)

    Wilson, Korey A.; Elefanty, Andrew G.; Stanley, Edouard G.; Gilbert, David M.

    2016-01-01

    ABSTRACT Lineage specification of both mouse and human pluripotent stem cells (PSCs) is accompanied by spatial consolidation of chromosome domains and temporal consolidation of their replication timing. Replication timing and chromatin organization are both established during G1 phase at the timing decision point (TDP). Here, we have developed live cell imaging tools to track spatio-temporal replication domain consolidation during differentiation. First, we demonstrate that the fluorescence ubiquitination cell cycle indicator (Fucci) system is incapable of demarcating G1/S or G2/M cell cycle transitions. Instead, we employ a combination of fluorescent PCNA to monitor S phase progression, cytokinesis to demarcate mitosis, and fluorescent nucleotides to label early and late replication foci and track their 3D organization into sub-nuclear chromatin compartments throughout all cell cycle transitions. We find that, as human PSCs differentiate, the length of S phase devoted to replication of spatially clustered replication foci increases, coincident with global compartmentalization of domains into temporally clustered blocks of chromatin. Importantly, re-localization and anchorage of domains was completed prior to the onset of S phase, even in the context of an abbreviated PSC G1 phase. This approach can also be employed to investigate cell fate transitions in single PSCs, which could be seen to differentiate preferentially from G1 phase. Together, our results establish real-time, live-cell imaging methods for tracking cell cycle transitions during human PSC differentiation that can be applied to study chromosome domain consolidation and other aspects of lineage specification. PMID:27433885

  1. CTCF and CohesinSA-1 Mark Active Promoters and Boundaries of Repressive Chromatin Domains in Primary Human Erythroid Cells.

    Directory of Open Access Journals (Sweden)

    Laurie A Steiner

    Full Text Available CTCF and cohesinSA-1 are regulatory proteins involved in a number of critical cellular processes including transcription, maintenance of chromatin domain architecture, and insulator function. To assess changes in the CTCF and cohesinSA-1 interactomes during erythropoiesis, chromatin immunoprecipitation coupled with high throughput sequencing and mRNA transcriptome analyses via RNA-seq were performed in primary human hematopoietic stem and progenitor cells (HSPC and primary human erythroid cells from single donors.Sites of CTCF and cohesinSA-1 co-occupancy were enriched in gene promoters in HSPC and erythroid cells compared to single CTCF or cohesin sites. Cell type-specific CTCF sites in erythroid cells were linked to highly expressed genes, with the opposite pattern observed in HSPCs. Chromatin domains were identified by ChIP-seq with antibodies against trimethylated lysine 27 histone H3, a modification associated with repressive chromatin. Repressive chromatin domains increased in both number and size during hematopoiesis, with many more repressive domains in erythroid cells than HSPCs. CTCF and cohesinSA-1 marked the boundaries of these repressive chromatin domains in a cell-type specific manner.These genome wide data, changes in sites of protein occupancy, chromatin architecture, and related gene expression, support the hypothesis that CTCF and cohesinSA-1 have multiple roles in the regulation of gene expression during erythropoiesis including transcriptional regulation at gene promoters and maintenance of chromatin architecture. These data from primary human erythroid cells provide a resource for studies of normal and perturbed erythropoiesis.

  2. The 10 kDa domain of human erythrocyte protein 4.1 binds the Plasmodium falciparum EBA-181 protein

    Directory of Open Access Journals (Sweden)

    Coetzer Theresa L

    2006-11-01

    Full Text Available Abstract Background Erythrocyte invasion by Plasmodium falciparum parasites represents a key mechanism during malaria pathogenesis. Erythrocyte binding antigen-181 (EBA-181 is an important invasion protein, which mediates a unique host cell entry pathway. A novel interaction between EBA-181 and human erythrocyte membrane protein 4.1 (4.1R was recently demonstrated using phage display technology. In the current study, recombinant proteins were utilized to define and characterize the precise molecular interaction between the two proteins. Methods 4.1R structural domains (30, 16, 10 and 22 kDa domain and the 4.1R binding region in EBA-181 were synthesized in specific Escherichia coli strains as recombinant proteins and purified using magnetic bead technology. Recombinant proteins were subsequently used in blot-overlay and histidine pull-down assays to determine the binding domain in 4.1R. Results Blot overlay and histidine pull-down experiments revealed specific interaction between the 10 kDa domain of 4.1R and EBA-181. Binding was concentration dependent as well as saturable and was abolished by heat denaturation of 4.1R. Conclusion The interaction of EBA-181 with the highly conserved 10 kDa domain of 4.1R provides new insight into the molecular mechanisms utilized by P. falciparum during erythrocyte entry. The results highlight the potential multifunctional role of malaria invasion proteins, which may contribute to the success of the pathogenic stage of the parasite's life cycle.

  3. Crystallization and preliminary X-ray analysis of the vWA domain of human anthrax toxin receptor 1

    International Nuclear Information System (INIS)

    Cai, Chenguang; Zhao, Ying; Tong, Xiaohang; Fu, Sheng; Li, Yuanyuan; Wu, Yang; Li, Xumei; Lou, Zhiyong

    2010-01-01

    The vWA domain of human anthrax toxin receptor 1 was overexpressed in E. coli, purified and crystallized. Diffraction data were collected to 1.8 Å resolution. The Gram-positive spore-forming bacterium Bacillus anthracis causes anthrax by secreting anthrax toxin, which consists of protective antigen (PA), lethal factor and oedema factor. Binding of PA to receptors triggers the multi-step process of anthrax toxin entry into target cells. Two distinct cellular receptors, ANTXR1 (also known as tumour endothelial marker 8; TEM8) and ANTXR2 (also known as capillary morphogenesis protein 2; CMG2), for anthrax toxin have been identified. Although the crystal structure of the extracellular von Willebrand factor A (vWA) domain of CMG2 has been reported, the difference between the vWA domains of TEM8 and CMG2 remains unclear because there are no structural data for the TEM8 vWA domain. In this report, the TEM8 vWA domain was expressed, purified and crystallized. X-ray diffraction data were collected to 1.8 Å resolution from a single crystal, which belonged to space group P1 with unit-cell parameters a = 65.9, b = 66.1, c = 74.4 Å, α = 63.7, β = 88.2, γ = 59.9°

  4. Crystal structures of the UBX domain of human UBXD7 and its complex with p97 ATPase.

    Science.gov (United States)

    Li, Zhi-Hui; Wang, Yong; Xu, Min; Jiang, Tao

    2017-04-22

    In humans, UBXD7 (also called UBXN7), an adaptor of p97 ATPase, can participate in the degradation of misfolded or damaged proteins in the p97-mediated ubiquitin proteasome system (UPS). UBXD7 binds to ubiquitinated substrates via its UBA domain and interacts with p97 N-terminal domain through its UBX domain to recruit p97 or the p97 core complex (p97/NPL4/UFD1). Here, we report the crystal structures of the UBX domain of UBXD7 (UBXD7 UBX ) at 2.0 Å resolution and its complex with p97 N-terminal domain (p97 NTD -UBXD7 UBX complex) at 2.4 Å resolution. A structural analysis and isothermal titration calorimetry results provide detailed molecular basis of interaction between UBXD7 UBX and p97 NTD . Moreover, structural superpositions suggest that dimerization of UBXD7 UBX via an intermolecular disulfide bond could interfere with the formation of the p97 NTD -UBXD7 UBX complex. Interestingly, UBXD7 may have a cooperative effect on p97 interaction with UFD1. Together, these results provide structural and biochemical insights into the interaction between p97 NTD and UBXD7 UBX . Copyright © 2017 Elsevier Inc. All rights reserved.

  5. The crystal structure of the regulatory domain of the human sodium-driven chloride/bicarbonate exchanger.

    Science.gov (United States)

    Alvadia, Carolina M; Sommer, Theis; Bjerregaard-Andersen, Kaare; Damkier, Helle Hasager; Montrasio, Michele; Aalkjaer, Christian; Morth, J Preben

    2017-09-21

    The sodium-driven chloride/bicarbonate exchanger (NDCBE) is essential for maintaining homeostatic pH in neurons. The crystal structure at 2.8 Å resolution of the regulatory N-terminal domain of human NDCBE represents the first crystal structure of an electroneutral sodium-bicarbonate cotransporter. The crystal structure forms an equivalent dimeric interface as observed for the cytoplasmic domain of Band 3, and thus establishes that the consensus motif VTVLP is the key minimal dimerization motif. The VTVLP motif is highly conserved and likely to be the physiologically relevant interface for all other members of the SLC4 family. A novel conserved Zn 2+ -binding motif present in the N-terminal domain of NDCBE is identified and characterized in vitro. Cellular studies confirm the Zn 2+ dependent transport of two electroneutral bicarbonate transporters, NCBE and NBCn1. The Zn 2+ site is mapped to a cluster of histidines close to the conserved ETARWLKFEE motif and likely plays a role in the regulation of this important motif. The combined structural and bioinformatics analysis provides a model that predicts with additional confidence the physiologically relevant interface between the cytoplasmic domain and the transmembrane domain.

  6. Purification, crystallization and preliminary X-ray analysis of the IgV domain of human nectin-4

    International Nuclear Information System (INIS)

    Xu, Xiang; Zhang, Xiaoai; Lu, Guangwen; Cai, Yongping

    2012-01-01

    Nectin-4 belongs to a family of immunoglobulin-like cell adhesion molecules and is highly expressed in cancer cells. Recently, nectin-4 was found to be a receptor of measles virus and the IgV domain sustains strong binding to measles virus H protein. In this study, the successful expression and purification of human nectin-4 V domain (nectin-4v) is reported Nectin-4 belongs to a family of immunoglobulin-like cell adhesion molecules and is highly expressed in cancer cells. Recently, nectin-4 was found to be a receptor of measles virus and the IgV domain sustains strong binding to measles virus H protein. In this study, the successful expression and purification of human nectin-4 V domain (nectin-4v) is reported. The purified protein was crystallized using the sitting-drop vapour-diffusion method. The crystals diffracted to 1.8 Å resolution and belonged to space group P2 1 , with unit-cell parameters a = 33.1, b = 51.7, c = 56.9 Å, β = 94.7°. Preliminary analysis of the diffraction data was also performed

  7. Expression, purification, crystallization and preliminary X-ray crystallographic analysis of the SH3 domain of human AHI1

    International Nuclear Information System (INIS)

    Shi, Zhuliang; Liang, Ning; Xu, Wei; Li, Kuai; Sheng, Guoqing; Liu, Jinsong; Xu, Aimin; Li, Xiao-Jiang; Wu, Donghai

    2009-01-01

    The SH3 domain of human AHI1 has been cloned and expressed in Escherichia coli. The protein was purified by affinity and size-exclusion chromatography and was crystallized using the sitting-drop vapour-diffusion method at 293 K. The SH3 domain of human AHI1 was cloned and expressed in Escherichia coli. The protein was purified by affinity and size-exclusion chromatography and was crystallized using the sitting-drop vapour-diffusion method at 293 K. A complete data set was collected to 2.5 Å resolution at 110 K. The crystal belonged to space group P4 1 2 1 2, with unit-cell parameters a = 67.377, b = 67.377, c = 98.549 Å

  8. Assessment of regional ventilation and deformation using 4D-CT imaging for healthy human lungs during tidal breathing.

    Science.gov (United States)

    Jahani, Nariman; Choi, Sanghun; Choi, Jiwoong; Iyer, Krishna; Hoffman, Eric A; Lin, Ching-Long

    2015-11-15

    This study aims to assess regional ventilation, nonlinearity, and hysteresis of human lungs during dynamic breathing via image registration of four-dimensional computed tomography (4D-CT) scans. Six healthy adult humans were studied by spiral multidetector-row CT during controlled tidal breathing as well as during total lung capacity and functional residual capacity breath holds. Static images were utilized to contrast static vs. dynamic (deep vs. tidal) breathing. A rolling-seal piston system was employed to maintain consistent tidal breathing during 4D-CT spiral image acquisition, providing required between-breath consistency for physiologically meaningful reconstructed respiratory motion. Registration-derived variables including local air volume and anisotropic deformation index (ADI, an indicator of preferential deformation in response to local force) were employed to assess regional ventilation and lung deformation. Lobar distributions of air volume change during tidal breathing were correlated with those of deep breathing (R(2) ≈ 0.84). Small discrepancies between tidal and deep breathing were shown to be likely due to different distributions of air volume change in the left and the right lungs. We also demonstrated an asymmetric characteristic of flow rate between inhalation and exhalation. With ADI, we were able to quantify nonlinearity and hysteresis of lung deformation that can only be captured in dynamic images. Nonlinearity quantified by ADI is greater during inhalation, and it is stronger in the lower lobes (P < 0.05). Lung hysteresis estimated by the difference of ADI between inhalation and exhalation is more significant in the right lungs than that in the left lungs. Copyright © 2015 the American Physiological Society.

  9. Point Organ Radiation Dose in Abdominal CT: Effect of Patient Off-Centering in an Experimental Human Cadaver Study.

    Science.gov (United States)

    Ali Khawaja, Ranish Deedar; Singh, Sarabjeet; Padole, Atul; Otrakji, Alexi; Lira, Diego; Zhang, Da; Liu, Bob; Primak, Andrew; Xu, George; Kalra, Mannudeep K

    2017-08-01

    To determine the effect of patient off-centering on point organ radiation dose measurements in a human cadaver scanned with routine abdominal CT protocol. A human cadaver (88 years, body-mass-index 20 kg/m2) was scanned with routine abdominal CT protocol on 128-slice dual source MDCT (Definition Flash, Siemens). A total of 18 scans were performed using two scan protocols (a) 120 kV-200 mAs fixed-mA (CTDIvol 14 mGy) (b) 120 kV-125 ref mAs (7 mGy) with automatic exposure control (AEC, CareDose 4D) at three different positions (a) gantry isocenter, (b) upward off-centering and (c) downward off-centering. Scanning was repeated three times at each position. Six thimble (in liver, stomach, kidney, pancreas, colon and urinary bladder) and four MOSFET dosimeters (on cornea, thyroid, testicle and breast) were placed for calculation of measured point organ doses. Organ dose estimations were retrieved from dose-tracking software (eXposure, Radimetrics). Statistical analysis was performed using analysis of variance. There was a significant difference between the trends of point organ doses with AEC and fixed-mA at all three positions (p 92% for both protocols; p < 0.0001). For both protocols, the highest mean difference in point doses was found for stomach and lowest for colon. Measured absorbed point doses in abdominal CT vary with patient-centering in the gantry isocenter. Due to lack of consideration of patient positioning in the dose estimation on automatic software-over estimation of the doses up to 92% was reported. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  10. Reconstruction of Cochlea Based on Micro-CT and Histological Images of the Human Inner Ear

    Directory of Open Access Journals (Sweden)

    Christos Bellos

    2014-01-01

    Full Text Available The study of the normal function and pathology of the inner ear has unique difficulties as it is inaccessible during life and, so, conventional techniques of pathologic studies such as biopsy and surgical excision are not feasible, without further impairing function. Mathematical modelling is therefore particularly attractive as a tool in researching the cochlea and its pathology. The first step towards efficient mathematical modelling is the reconstruction of an accurate three dimensional (3D model of the cochlea that will be presented in this paper. The high quality of the histological images is being exploited in order to extract several sections of the cochlea that are not visible on the micro-CT (mCT images (i.e., scala media, spiral ligament, and organ of Corti as well as other important sections (i.e., basilar membrane, Reissner membrane, scala vestibule, and scala tympani. The reconstructed model is being projected in the centerline of the coiled cochlea, extracted from mCT images, and represented in the 3D space. The reconstruction activities are part of the SIFEM project, which will result in the delivery of an infrastructure, semantically interlinking various tools and libraries (i.e., segmentation, reconstruction, and visualization tools with the clinical knowledge, which is represented by existing data, towards the delivery of a robust multiscale model of the inner ear.

  11. Determination of the optical properties of melanin-pigmented human skin equivalents using terahertz time-domain spectroscopy

    Science.gov (United States)

    Lipscomb, Dawn; Echchgadda, Ibtissam; Peralta, Xomalin G.; Wilmink, Gerald J.

    2013-02-01

    Terahertz time-domain spectroscopy (THz-TDS) methods have been utilized in previous studies in order to characterize the optical properties of skin and its primary constituents (i.e., water, collagen, and keratin). However, similar experiments have not yet been performed to investigate whether melanocytes and the melanin pigment that they synthesize contribute to skin's optical properties. In this study, we used THz-TDS methods operating in transmission geometry to measure the optical properties of in vitro human skin equivalents with or without normal human melanocytes. Skin equivalents were cultured for three weeks to promote gradual melanogenesis, and THz time domain data were collected at various time intervals. Frequency-domain analysis techniques were performed to determine the index of refraction (n) and absorption coefficient (μa) for each skin sample over the frequency range of 0.1-2.0 THz. We found that for all samples as frequency increased, n decreased exponentially and the μa increased linearly. Additionally, we observed that skin samples with higher levels of melanin exhibited greater n and μa values than the non-pigmented samples. Our results indicate that melanocytes and the degree of melanin pigmentation contribute in an appreciable manner to the skin's optical properties. Future studies will be performed to examine whether these contributions are observed in human skin in vivo.

  12. Signatures of pleiotropy, economy and convergent evolution in a domain-resolved map of human-virus protein-protein interaction networks.

    Directory of Open Access Journals (Sweden)

    Sara Garamszegi

    Full Text Available A central challenge in host-pathogen systems biology is the elucidation of general, systems-level principles that distinguish host-pathogen interactions from within-host interactions. Current analyses of host-pathogen and within-host protein-protein interaction networks are largely limited by their resolution, treating proteins as nodes and interactions as edges. Here, we construct a domain-resolved map of human-virus and within-human protein-protein interaction networks by annotating protein interactions with high-coverage, high-accuracy, domain-centric interaction mechanisms: (1 domain-domain interactions, in which a domain in one protein binds to a domain in a second protein, and (2 domain-motif interactions, in which a domain in one protein binds to a short, linear peptide motif in a second protein. Analysis of these domain-resolved networks reveals, for the first time, significant mechanistic differences between virus-human and within-human interactions at the resolution of single domains. While human proteins tend to compete with each other for domain binding sites by means of sequence similarity, viral proteins tend to compete with human proteins for domain binding sites in the absence of sequence similarity. Independent of their previously established preference for targeting human protein hubs, viral proteins also preferentially target human proteins containing linear motif-binding domains. Compared to human proteins, viral proteins participate in more domain-motif interactions, target more unique linear motif-binding domains per residue, and contain more unique linear motifs per residue. Together, these results suggest that viruses surmount genome size constraints by convergently evolving multiple short linear motifs in order to effectively mimic, hijack, and manipulate complex host processes for their survival. Our domain-resolved analyses reveal unique signatures of pleiotropy, economy, and convergent evolution in viral

  13. Signatures of pleiotropy, economy and convergent evolution in a domain-resolved map of human-virus protein-protein interaction networks.

    Science.gov (United States)

    Garamszegi, Sara; Franzosa, Eric A; Xia, Yu

    2013-01-01

    A central challenge in host-pathogen systems biology is the elucidation of general, systems-level principles that distinguish host-pathogen interactions from within-host interactions. Current analyses of host-pathogen and within-host protein-protein interaction networks are largely limited by their resolution, treating proteins as nodes and interactions as edges. Here, we construct a domain-resolved map of human-virus and within-human protein-protein interaction networks by annotating protein interactions with high-coverage, high-accuracy, domain-centric interaction mechanisms: (1) domain-domain interactions, in which a domain in one protein binds to a domain in a second protein, and (2) domain-motif interactions, in which a domain in one protein binds to a short, linear peptide motif in a second protein. Analysis of these domain-resolved networks reveals, for the first time, significant mechanistic differences between virus-human and within-human interactions at the resolution of single domains. While human proteins tend to compete with each other for domain binding sites by means of sequence similarity, viral proteins tend to compete with human proteins for domain binding sites in the absence of sequence similarity. Independent of their previously established preference for targeting human protein hubs, viral proteins also preferentially target human proteins containing linear motif-binding domains. Compared to human proteins, viral proteins participate in more domain-motif interactions, target more unique linear motif-binding domains per residue, and contain more unique linear motifs per residue. Together, these results suggest that viruses surmount genome size constraints by convergently evolving multiple short linear motifs in order to effectively mimic, hijack, and manipulate complex host processes for their survival. Our domain-resolved analyses reveal unique signatures of pleiotropy, economy, and convergent evolution in viral-host interactions that are

  14. Clinical evaluation of the radiolanthanide terbium-152: first-in-human PET/CT with 152Tb-DOTATOC.

    Science.gov (United States)

    Baum, Richard P; Singh, Aviral; Benešová, Martina; Vermeulen, Christiaan; Gnesin, Silvano; Köster, Ulli; Johnston, Karl; Müller, Dirk; Senftleben, Stefan; Kulkarni, Harshad R; Türler, Andreas; Schibli, Roger; Prior, John O; van der Meulen, Nicholas P; Müller, Cristina

    2017-10-31

    The existence of theragnostic pairs of radionuclides allows the preparation of radiopharmaceuticals for diagnostic and therapeutic purposes. Radiolanthanides, such as 177 Lu, are successfully used for therapeutic purposes; however, a perfect diagnostic match is currently not available for clinical use. A unique, multi-disciplinary study was performed using 152 Tb (T 1/2 = 17.5 h, Eβ + average = 1140 keV, Iβ + = 20.3%), which resulted in the first-in-human PET/CT images with this promising radionuclide. For this purpose, 152 Tb was produced via a spallation process followed by mass separation at ISOLDE, CERN. The chemical separation and quality control, performed at PSI, resulted in a pure product in sufficient yields. Clinical PET phantom studies revealed an increased image noise level, because of the smaller β + branching ratio of 152 Tb as compared to standard PET nuclides at matched activity concentrations; however, the expected recovery would be comparable at matched signal-to-noise ratios in clinical PET. 152 Tb was used for labeling DOTATOC, at Zentralklinik Bad Berka, and administered to a patient for a first-in-human clinical study. PET scans were performed over a period of 24 h, allowing the visualization of even small metastases with increased tumor-to-background contrast over time. Based on the results obtained in this work, it can be deduced that PET/CT imaging with 152 Tb-labeled targeting agents has promise for clinical application and may be particularly interesting for pre-therapeutic dosimetry.

  15. Domain-restricted mutation analysis to identify novel driver events in human cancer

    Directory of Open Access Journals (Sweden)

    Sanket Desai

    2017-10-01

    Full Text Available Analysis of mutational spectra across various cancer types has given valuable insights into tumorigenesis. Different approaches have been used to identify novel drivers from the set of somatic mutations, including the methods which use sequence conservation, geometric localization and pathway information. Recent computational methods suggest use of protein domain information for analysis and understanding of the functional consequence of non-synonymous mutations. Similarly, evidence suggests recurrence at specific position in proteins is robust indicators of its functional impact. Building on this, we performed a systematic analysis of TCGA exome derived somatic mutations across 6089 PFAM domains and significantly mutated domains were identified using randomization approach. Multiple alignment of individual domain allowed us to prioritize for conserved residues mutated at analogous positions across different proteins in a statistically disciplined manner. In addition to the known frequently mutated genes, this analysis independently identifies low frequency Meprin and TRAF-Homology (MATH domain in Speckle Type BTB/POZ (SPOP protein, in prostate adenocarcinoma. Results from this analysis will help generate hypotheses about the downstream molecular mechanism resulting in cancer phenotypes.

  16. Human ribosomal protein L37 has motifs predicting serine/threonine phosphorylation and a zinc-finger domain.

    Science.gov (United States)

    Barnard, G F; Staniunas, R J; Puder, M; Steele, G D; Chen, L B

    1994-08-02

    Ribosomal protein L37 mRNA is overexpressed in colon cancer. The nucleotide sequences of human L37 from several tumor and normal, colon and liver cDNA sources were determined to be identical. L37 mRNA was approximately 375 nucleotides long encoding 97 amino acids with M(r) = 11,070, pI = 12.6, multiple potential serine/threonine phosphorylation sites and a zinc-finger domain. The human sequence is compared to other species.

  17. Purification, crystallization and preliminary crystallographic characterization of the caspase-recruitment domain of human Nod1

    International Nuclear Information System (INIS)

    Srimathi, Thiagarajan; Robbins, Sheila L.; Dubas, Rachel L.; Seo, Jang-Hoon; Park, Young Chul

    2006-01-01

    The caspase-recruitment domain of the cytosolic pathogen receptor Nod1 was crystallized. X-ray diffraction data were collected to 1.9 Å resolution. The caspase-recruitment domain (CARD) is known to play an important role in apoptosis and inflammation as an essential protein–protein interaction domain. The CARD of the cytosolic pathogen receptor Nod1 was overexpressed in Escherichia coli and purified by affinity chromatography and gel filtration. The purified CARD was crystallized at 277 K using the microseeding method. X-ray diffraction data were collected to 1.9 Å resolution. The crystals belong to space group P3 1 or P3 2 , with unit-cell parameters a = b = 79.1, c = 80.9 Å. Preliminary analysis indicates that there is one dimeric CARD molecule in the asymmetric unit

  18. The metal chaperone Atox1 regulates the activity of the human copper transporter ATP7B by modulating domain dynamics.

    Science.gov (United States)

    Yu, Corey H; Yang, Nan; Bothe, Jameson; Tonelli, Marco; Nokhrin, Sergiy; Dolgova, Natalia V; Braiterman, Lelita; Lutsenko, Svetlana; Dmitriev, Oleg Y

    2017-11-03

    The human transporter ATP7B delivers copper to the biosynthetic pathways and maintains copper homeostasis in the liver. Mutations in ATP7B cause the potentially fatal hepatoneurological disorder Wilson disease. The activity and intracellular localization of ATP7B are regulated by copper, but the molecular mechanism of this regulation is largely unknown. We show that the copper chaperone Atox1, which delivers copper to ATP7B, and the group of the first three metal-binding domains (MBD1-3) are central to the activity regulation of ATP7B. Atox1-Cu binding to ATP7B changes domain dynamics and interactions within the MBD1-3 group and activates ATP hydrolysis. To understand the mechanism linking Atox1-MBD interactions and enzyme activity, we have determined the MBD1-3 conformational space using small angle X-ray scattering and identified changes in MBD dynamics caused by apo -Atox1 and Atox1-Cu by solution NMR. The results show that copper transfer from Atox1 decreases domain interactions within the MBD1-3 group and increases the mobility of the individual domains. The N-terminal segment of MBD1-3 was found to interact with the nucleotide-binding domain of ATP7B, thus physically coupling the domains involved in copper binding and those involved in ATP hydrolysis. Taken together, the data suggest a regulatory mechanism in which Atox1-mediated copper transfer activates ATP7B by releasing inhibitory constraints through increased freedom of MBD1-3 motions. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  19. Predicting detection performance with model observers: Fourier domain or spatial domain?

    Science.gov (United States)

    Chen, Baiyu; Yu, Lifeng; Leng, Shuai; Kofler, James; Favazza, Christopher; Vrieze, Thomas; McCollough, Cynthia

    2016-02-27

    The use of Fourier domain model observer is challenged by iterative reconstruction (IR), because IR algorithms are nonlinear and IR images have noise texture different from that of FBP. A modified Fourier domain model observer, which incorporates nonlinear noise and resolution properties, has been proposed for IR and needs to be validated with human detection performance. On the other hand, the spatial domain model observer is theoretically applicable to IR, but more computationally intensive than the Fourier domain method. The purpose of this study is to compare the modified Fourier domain model observer to the spatial domain model observer with both FBP and IR images, using human detection performance as the gold standard. A phantom with inserts of various low contrast levels and sizes was repeatedly scanned 100 times on a third-generation, dual-source CT scanner at 5 dose levels and reconstructed using FBP and IR algorithms. The human detection performance of the inserts was measured via a 2-alternative-forced-choice (2AFC) test. In addition, two model observer performances were calculated, including a Fourier domain non-prewhitening model observer and a spatial domain channelized Hotelling observer. The performance of these two mode observers was compared in terms of how well they correlated with human observer performance. Our results demonstrated that the spatial domain model observer correlated well with human observers across various dose levels, object contrast levels, and object sizes. The Fourier domain observer correlated well with human observers using FBP images, but overestimated the detection performance using IR images.

  20. NMR assignments of SPOC domain of the human transcriptional corepressor SHARP in complex with a C-terminal SMRT peptide.

    Science.gov (United States)

    Mikami, Suzuka; Kanaba, Teppei; Ito, Yutaka; Mishima, Masaki

    2013-10-01

    The transcriptional corepressor SMRT/HDAC1-associated repressor protein (SHARP) recruits histone deacetylases. Human SHARP protein is thought to function in processes involving steroid hormone responses and the Notch signaling pathway. SHARP consists of RNA recognition motifs (RRMs) in the N-terminal region and the spen paralog and ortholog C-terminal (SPOC) domain in the C-terminal region. It is known that the SPOC domain binds the LSD motif in the C-terminal tail of corepressors silencing mediator for retinoid and thyroid receptor (SMRT)/nuclear receptor corepressor (NcoR). We are interested in delineating the mechanism by which the SPOC domain recognizes the LSD motif of the C-terminal tail of SMRT/NcoR. To this end, we are investigating the tertiary structure of the SPOC/SMRT peptide using NMR. Herein, we report on the (1)H, (13)C and (15)N resonance assignments of the SPOC domain in complex with a SMRT peptide, which contributes towards a structural understanding of the SPOC/SMRT peptide and its molecular recognition.

  1. The study on mutations of the gene of extracellular domain of human thyrotropin receptor in the patients with thyroid diseases

    International Nuclear Information System (INIS)

    Zhang Zuncheng; Fang Peihua; Tan Jian; Lu Mei

    2002-01-01

    Objective: To define the sequence of the gene of extracellular domain of normal human thyrotropin receptor (hTSHR) and to investigate the mutations of the gene in the patients with thyroid diseases. Methods: Total RNAs were extracted from the thyroid tissue of four normal controls, twelve Graves' disease, four Hashimoto's thyroiditis and eleven nodular goiter patients. The extracellular domain of hTSHR genes were amplified by reverse transcription-polymerase chain reaction (RT-PCR) and sequenced with CEQ 2000 Genetic Analyzer. Results: The normal controls and the patients with thyroid disease had the same gene sequences of the extracellular domain of hTSHR. No mutation was found, except a silent base exchange in exon 7 (Asn187) at 661 base, in which 20 samples were 'T', 11 samples were 'C', without changes of amino acid of the TSHR. Conclusions: This study has not revealed mutations in the gene of extracellular domain of hTSHR. Other molecular pathogenetic mechanisms may be involved and more research is demanded

  2. Identification of an evolutionary conserved SURF-6 domain in a family of nucleolar proteins extending from human to yeast

    International Nuclear Information System (INIS)

    Polzikov, Mikhail; Zatsepina, Olga; Magoulas, Charalambos

    2005-01-01

    The mammalian SURF-6 protein is localized in the nucleolus, yet its function remains elusive in the recently characterized nucleolar proteome. We discovered by searching the Protein families database that a unique evolutionary conserved SURF-6 domain is present in the carboxy-terminal of a novel family of eukaryotic proteins extending from human to yeast. By using the enhanced green fluorescent protein as a fusion protein marker in mammalian cells, we show that proteins from distantly related taxonomic groups containing the SURF-6 domain are localized in the nucleolus. Deletion sequence analysis shows that multiple regions of the SURF-6 protein are capable of nucleolar targeting independently of the evolutionary conserved domain. We identified that the Saccharomyces cerevisiae member of the SURF-6 family, named rrp14 or ykl082c, has been categorized in yeast databases to interact with proteins involved in ribosomal biogenesis and cell polarity. These results classify SURF-6 as a new family of nucleolar proteins in the eukaryotic kingdom and point out that SURF-6 has a distinct domain within the known nucleolar proteome that may mediate complex protein-protein interactions for analogous processes between yeast and mammalian cells

  3. {sup 18F} FDG Uptake of Human Testis on PET/CT: Correlation with Age, Sex Hormones, and Vasectomy

    Energy Technology Data Exchange (ETDEWEB)

    Moon, Seung Hwan; Eo, Jae Sun; Lee, Jong Jin; Chung, June Key; Lee, Dong Soo; Lee, Myung Chul [Seoul National Univ. Hospital, Seoul (Korea, Republic of)

    2011-12-15

    The purpose of this study was to evaluate glucose metabolism of normal human testis on {sup 18F} FDG PET/CT and to assess possible correlation among age, the serum levels of sex hormones, and vasectomy. {sup 18F} FDG PET/CT was performed in 66 normal healthy men (50.8{+-}13.6 years, range 22-81), and mean standard uptake values (SUV) of {sup 18F} FDG in testis and adductor muscle were measured. Testis muscle SUV ratios (T/M ratios) were calculated. Serum levels of total testosterone, free testosterone, estradiol, and of sex hormone binding globulin (SHBG) were measured. We searched for correlations between T/M ratios and age and the serum concentrations of sex hormones. {sup 18F} FDG PET/CT was also performed in 32 vasectomized men (55.7{+-}7.8 years, range 38-71) and 52 nonvasectomized men (55.4{+-}11.6 years, range 37-72). Mean SUVs of testis and adductor muscle were measured, and T/M ratios were calculated. A significant age related decline was found in T/M ratio (r=-0.509, p<0.0001). Serum levels of total testosterone and free testosterone were also found to be positively correlated with T/M ratio (r=-0.427, p=0.0003; r=0.435, p=0.0003, respectively). The mean SUV and T/M ratio of vasectomized men were significantly lower than those of nonvasectomized men (p<0.0378 and p=0.0001, respectively). Glucose metabolism in the testis in an adult population was found to be correlated with age, serum sex hormone level, and vasectomy history. These results indicate that testicular {sup 18F} FDG uptake may have attributed to testicular function and testicular histology. Our findings may have important implications for the interpretation of testicular {sup 18F} FDG uptake in the normal adult population.

  4. Plasmodium falciparum CS protein - prime malaria vaccine candidate: definition of the human CTL domain and analysis of its variation

    Directory of Open Access Journals (Sweden)

    Denise L. Doolan

    1992-01-01

    Full Text Available Studies in mice have shown that immunity to malaria sporozoites is mediated primarily by citotoxic T lymphocytes (CTL specific for epitopes within the circumsporozoite (CS protein. Humans, had never been shown to generate CTL against any malaria or other parasite protein. The design of a sub-unit vaccine for humans ralies on the epitopes recognized by CTL being identified and polymorphisms therein being defined. We have developed a novel technique using an entire series of overlapping synthetic peptides to define the epitopes of the Plasmodium falciparum CS protein recognized by human CTL and have analyzed the sequence variation of the protein with respect to the identified CTL epitopic domain. We have demonstrated that some humans can indeed generate CTL. against the P. falciparum CS protein. Furthermore, the extent of variation observed for the CTL recognition domain is finite and the combination of peptides necessary for inclusion in a polyvalent vaccine may be small. If ways can be found to increase immune responsiveness, then a vaccine designed to stimulate CS protein-specific CTL activity may prevent malaria.

  5. Identification of Amino Acids in the Human Tetherin Transmembrane Domain Responsible for HIV-1 Vpu Interaction and Susceptibility▿ †

    Science.gov (United States)

    Kobayashi, Tomoko; Ode, Hirotaka; Yoshida, Takeshi; Sato, Kei; Gee, Peter; Yamamoto, Seiji P.; Ebina, Hirotaka; Strebel, Klaus; Sato, Hironori; Koyanagi, Yoshio

    2011-01-01

    Tetherin, also known as BST-2/CD317/HM1.24, is an antiviral cellular protein that inhibits the release of HIV-1 particles from infected cells. HIV-1 viral protein U (Vpu) is a specific antagonist of human tetherin that might contribute to the high virulence of HIV-1. In this study, we show that three amino acid residues (I34, L37, and L41) in the transmembrane (TM) domain of human tetherin are critical for the interaction with Vpu by using a live cell-based assay. We also found that the conservation of an additional amino acid at position 45 and two residues downstream of position 22, which are absent from monkey tetherins, are required for the antagonism by Vpu. Moreover, computer-assisted structural modeling and mutagenesis studies suggest that an alignment of these four amino acid residues (I34, L37, L41, and T45) on the same helical face in the TM domain is crucial for the Vpu-mediated antagonism of human tetherin. These results contribute to the molecular understanding of human tetherin-specific antagonism by HIV-1 Vpu. PMID:21068238

  6. Identification of amino acids in the human tetherin transmembrane domain responsible for HIV-1 Vpu interaction and susceptibility.

    Science.gov (United States)

    Kobayashi, Tomoko; Ode, Hirotaka; Yoshida, Takeshi; Sato, Kei; Gee, Peter; Yamamoto, Seiji P; Ebina, Hirotaka; Strebel, Klaus; Sato, Hironori; Koyanagi, Yoshio

    2011-01-01

    Tetherin, also known as BST-2/CD317/HM1.24, is an antiviral cellular protein that inhibits the release of HIV-1 particles from infected cells. HIV-1 viral protein U (Vpu) is a specific antagonist of human tetherin that might contribute to the high virulence of HIV-1. In this study, we show that three amino acid residues (I34, L37, and L41) in the transmembrane (TM) domain of human tetherin are critical for the interaction with Vpu by using a live cell-based assay. We also found that the conservation of an additional amino acid at position 45 and two residues downstream of position 22, which are absent from monkey tetherins, are required for the antagonism by Vpu. Moreover, computer-assisted structural modeling and mutagenesis studies suggest that an alignment of these four amino acid residues (I34, L37, L41, and T45) on the same helical face in the TM domain is crucial for the Vpu-mediated antagonism of human tetherin. These results contribute to the molecular understanding of human tetherin-specific antagonism by HIV-1 Vpu.

  7. An ankyrin-like protein with transmembrane domains is specifically lost after oncogenic transformation of human fibroblasts.

    Science.gov (United States)

    Jaquemar, D; Schenker, T; Trueb, B

    1999-03-12

    We have identified a novel transformation-sensitive mRNA, which is present in cultured fibroblasts but is lacking in SV40 transformed cells as well as in many mesenchymal tumor cell lines. The corresponding gene is located on human chromosome 8 in band 8q13. The open reading frame of the mRNA encodes a protein of 1119 amino acids forming two distinct domains. The N-terminal domain consists of 18 repeats that are related to the cytoskeletal protein ankyrin. The C-terminal domain contains six putative transmembrane segments that resemble many ion channels. This overall structure is reminiscent of TRP-like proteins that function as store-operated calcium channels. The novel protein with an Mr of 130 kDa is expressed at a very low level in human fibroblasts and at a moderate level in liposarcoma cells. Overexpression in eukaryotic cells appears to interfere with normal growth, suggesting that it might play a direct or indirect role in signal transduction and growth control.

  8. Localization of the fourth membrane spanning domain as a ligand binding site in the human platelet α2-adrenergic receptor

    International Nuclear Information System (INIS)

    Matsui, Hiroaki; Lefkowitz, R.J.; Caron, M.G.; Regan, J.W.

    1989-01-01

    The human platelet α 2 -adrenergic receptor is an integral membrane protein which binds epinephrine. The gene for this receptor has been cloned, and the primary structure is thus known. A model of its secondary structure predicts that the receptor has seven transmembrane spanning domains. By covalent labeling and peptide mapping, the authors have identified a region of the receptor that is directly involved with ligand binding. Partially purified preparations of the receptor were covalently radiolabeled with either of two specific photoaffinity ligands: [ 3 H]SKF 102229 (an antagonist) or p-azido[ 3 H]clonidine (an agonist). The radiolabeled receptors were then digested with specific endopeptidases, and peptides containing the covalently bound radioligands were identified. Lysylendopeptidase treatment of [ 3 H]SKF 102229 labeled receptor yielded one peptide of M r 2400 as the product of a complete digest. Endopeptidase Arg-C gave a labeled peptide of M r 4000, which was further digested to the M r 2400 peptide by additional treatment with lysylendopeptidase. Using p-azido[ 3 H]clonidine-labeled receptor, a similar M r 2400 peptide was obtained by lysylendopeptidase cleavage. This M r 2400 peptide corresponds to the fourth transmembrane spanning domain of the receptor. These data suggest that this region forms part of the ligand binding domain of the human platelet α 2 -adrenergic receptor

  9. Sexual difference of human hyoid bones. Quantitative analysis of CT three-dimensional image

    International Nuclear Information System (INIS)

    Terashima, Yoshiharu; Izumi, Masahiro; Hanamura, Hajime; Takada, Yasushi

    2007-01-01

    We investigated sexual differences in hyoid bones of 50 dissected Japanese cadavers: 26 males (aged 52 to 101, averaged 81.9 years) and 24 females (aged 61 to 94, averaged 83.6 years). All extracted hyoid bones were scanned by multi-slice CT. Length of body, distance between bilateral greater horns, length of greater horns, distance between bilateral lesser horns, and length of lesser horns were measured on CT three-dimensional image, and were analyzed by univariate and multivariate statistics. t-tests showed significant sexual differences in all the dimensions; being about 20% longer in males than in females. In principal component analysis using five hyoid dimensions, factor 1, expressing the overall size of the bone, fairly separated each sex, but factors 2 and 3, expressing the shape, did not. Discriminant analysis by a stepwise model, using all the eight dimensions, classified sex rightly (88.6% of the bone) by a function of two dimensions: length of body and distance between bilateral tips of lesser horns. In conclusion, a sexual difference of the hyoid bone was evident in size rather than in shape. (author)

  10. CT scans in studies of the dynamics of liposynthesis and lipolysis of the human liver

    International Nuclear Information System (INIS)

    Huebener, K.H.; Schmitt, W.G.H.

    1981-01-01

    Following an analysis of the most important metabolic disturbances and other diseases associated ith fatty degeneration of the liver, it is shown, basing on a previous study, by means of examples, that subtle diagnosis of changes of the fatty content of the liver can be achieved via CT density measurement. Lipolysis in alcoholic fatty liver is quantified during abstention from alcohol, and a lipolysis factor of 1% fatty decomposition per day is determined from the measurements. Observations in pancreatitis with liver lipid storage during the course of the disease, point to an enzymatic weakness in lipid metabolism. Case histories show fatty degeneration associated with an enhanced cortisone serum level (in adrenal adenoma and cortisone therapy) and in poorly stabilized diabetes mellitus. It is concluded from the studies that determination of the relative fatty content by CT densitometry should be conducted as a routine procedure and that pathological lipid content should stimulate subtle diagnosis of the pancreas and the adrenals. In addition, examinations repeated during the course of the disease with assessment of changes in fatty content, will be useful for assessing the value of the therapy employed. (orig.) [de

  11. Segmentation of human skull in MRI using statistical shape information from CT data.

    Science.gov (United States)

    Wang, Defeng; Shi, Lin; Chu, Winnie C W; Cheng, Jack C Y; Heng, Pheng Ann

    2009-09-01

    To automatically segment the skull from the MRI data using a model-based three-dimensional segmentation scheme. This study exploited the statistical anatomy extracted from the CT data of a group of subjects by means of constructing an active shape model of the skull surfaces. To construct a reliable shape model, a novel approach was proposed to optimize the automatic landmarking on the coupled surfaces (i.e., the skull vault) by minimizing the description length that incorporated local thickness information. This model was then used to locate the skull shape in MRI of a different group of patients. Compared with performing landmarking separately on the coupled surfaces, the proposed landmarking method constructed models that had better generalization ability and specificity. The segmentation accuracies were measured by the Dice coefficient and the set difference, and compared with the method based on mathematical morphology operations. The proposed approach using the active shape model based on the statistical skull anatomy presented in the head CT data contributes to more reliable segmentation of the skull from MRI data.

  12. CT scans in studies of the dynamics of liposynthesis and lipolysis of the human liver

    Energy Technology Data Exchange (ETDEWEB)

    Huebener, K H; Schmitt, W G.H.

    1981-06-01

    Following an analysis of the most important metabolic disturbances and other diseases associated with fatty degeneration of the liver, it is shown, basing on a previous study, by means of examples, that subtle diagnosis of changes of the fatty content of the liver can be achieved via CT density measurement. Lipolysis in alcoholic fatty liver is quantified during abstention from alcohol, and a lipolysis factor of 1% fatty decomposition per day is determined from the measurements. Observations in pancreatitis with liver lipid storage during the course of the disease, point to an enzymatic weakness in lipid metabolism. Case histories show fatty degeneration associated with an enhanced cortisone serum level (in adrenal adenoma and cortisone therapy) and in poorly stabilized diabetes mellitus. It is concluded from the studies that determination of the relative fatty content by CT densitometry should be conducted as a routine procedure and that pathological lipid content should stimulate subtle diagnosis of the pancreas and the adrenals. In addition, examinations repeated during the course of the disease with assessment of changes in fatty content, will be useful for assessing the value of the therapy employed.

  13. Purification, crystallization and preliminary X-ray analysis of the IgV domain of human nectin-4.

    Science.gov (United States)

    Xu, Xiang; Zhang, Xiaoai; Lu, Guangwen; Cai, Yongping

    2012-08-01

    Nectin-4 belongs to a family of immunoglobulin-like cell adhesion molecules and is highly expressed in cancer cells. Recently, nectin-4 was found to be a receptor of measles virus and the IgV domain sustains strong binding to measles virus H protein. In this study, the successful expression and purification of human nectin-4 V domain (nectin-4v) is reported. The purified protein was crystallized using the sitting-drop vapour-diffusion method. The crystals diffracted to 1.8 Å resolution and belonged to space group P2(1), with unit-cell parameters a = 33.1, b = 51.7, c = 56.9 Å, β = 94.7°. Preliminary analysis of the diffraction data was also performed.

  14. The central domain of yeast transcription factor Rpn4 facilitates degradation of reporter protein in human cells.

    Science.gov (United States)

    Morozov, A V; Spasskaya, D S; Karpov, D S; Karpov, V L

    2014-10-16

    Despite high interest in the cellular degradation machinery and protein degradation signals (degrons), few degrons with universal activity along species have been identified. It has been shown that fusion of a target protein with a degradation signal from mammalian ornithine decarboxylase (ODC) induces fast proteasomal degradation of the chimera in both mammalian and yeast cells. However, no degrons from yeast-encoded proteins capable to function in mammalian cells were identified so far. Here, we demonstrate that the yeast transcription factor Rpn4 undergoes fast proteasomal degradation and its central domain can destabilize green fluorescent protein and Alpha-fetoprotein in human HEK 293T cells. Furthermore, we confirm the activity of this degron in yeast. Thus, the Rpn4 central domain is an effective interspecies degradation signal. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  15. Structure-Based Sequence Alignment of the Transmembrane Domains of All Human GPCRs: Phylogenetic, Structural and Functional Implications

    Science.gov (United States)

    Cvicek, Vaclav; Goddard, William A.; Abrol, Ravinder

    2016-01-01

    The understanding of G-protein coupled receptors (GPCRs) is undergoing a revolution due to increased information about their signaling and the experimental determination of structures for more than 25 receptors. The availability of at least one receptor structure for each of the GPCR classes, well separated in sequence space, enables an integrated superfamily-wide analysis to identify signatures involving the role of conserved residues, conserved contacts, and downstream signaling in the context of receptor structures. In this study, we align the transmembrane (TM) domains of all experimental GPCR structures to maximize the conserved inter-helical contacts. The resulting superfamily-wide GpcR Sequence-Structure (GRoSS) alignment of the TM domains for all human GPCR sequences is sufficient to generate a phylogenetic tree that correctly distinguishes all different GPCR classes, suggesting that the class-level differences in the GPCR superfamily are encoded at least partly in the TM domains. The inter-helical contacts conserved across all GPCR classes describe the evolutionarily conserved GPCR structural fold. The corresponding structural alignment of the inactive and active conformations, available for a few GPCRs, identifies activation hot-spot residues in the TM domains that get rewired upon activation. Many GPCR mutations, known to alter receptor signaling and cause disease, are located at these conserved contact and activation hot-spot residue positions. The GRoSS alignment places the chemosensory receptor subfamilies for bitter taste (TAS2R) and pheromones (Vomeronasal, VN1R) in the rhodopsin family, known to contain the chemosensory olfactory receptor subfamily. The GRoSS alignment also enables the quantification of the structural variability in the TM regions of experimental structures, useful for homology modeling and structure prediction of receptors. Furthermore, this alignment identifies structurally and functionally important residues in all human GPCRs

  16. Comparison of Measured and Estimated CT Organ Doses for Modulated and Fixed Tube Current:: A Human Cadaver Study.

    Science.gov (United States)

    Padole, Atul; Deedar Ali Khawaja, Ranish; Otrakji, Alexi; Zhang, Da; Liu, Bob; Xu, X George; Kalra, Mannudeep K

    2016-05-01

    The aim of this study was to compare the directly measured and the estimated computed tomography (CT) organ doses obtained from commercial radiation dose-tracking (RDT) software for CT performed with modulated tube current or automatic exposure control (AEC) technique and fixed tube current (mAs). With the institutional review board (IRB) approval, the ionization chambers were surgically implanted in a human cadaver (88 years old, male, 68 kg) in six locations such as liver, stomach, colon, left kidney, small intestine, and urinary bladder. The cadaver was scanned with routine abdomen pelvis protocol on a 128-slice, dual-source multidetector computed tomography (MDCT) scanner using both AEC and fixed mAs. The effective and quality reference mAs of 100, 200, and 300 were used for AEC and fixed mAs, respectively. Scanning was repeated three times for each setting, and measured and estimated organ doses (from RDT software) were recorded (N = 3*3*2 = 18). Mean CTDIvol for AEC and fixed mAs were 4, 8, 13 mGy and 7, 14, 21 mGy, respectively. The most estimated organ doses were significantly greater (P < 0.01) than the measured organ doses for both AEC and fixed mAs. At AEC, the mean estimated organ doses (for six organs) were 14.7 mGy compared to mean measured organ doses of 12.3 mGy. Similarly, at fixed mAs, the mean estimated organ doses (for six organs) were 24 mGy compared to measured organ doses of 22.3 mGy. The differences among the measured and estimated organ doses were higher for AEC technique compared to the fixed mAs for most organs (P < 0.01). The most CT organ doses estimated from RDT software are greater compared to directly measured organ doses, particularly when AEC technique is used for CT scanning. Copyright © 2016 The Association of University Radiologists. Published by Elsevier Inc. All rights reserved.

  17. Structure of the C-type lectin carbohydrate recognition domain of human Tetranectin

    DEFF Research Database (Denmark)

    Kastrup, Jette Sandholm Jensen; Nielsen, Bettina Bryde; Rasmussen, Hanne B.

    1998-01-01

    Tetranectin (TN) is a C-type lectin involved in fibrinolysis, being the only endogenous ligand known to bind specifically to the kringle 4 domain of plasminogen. TN was originally isolated from plasma, but shows a wide tissue distribution. Furthermore, TN has been found in the extracellular matri...... molecules. One sulfate ion has been located at the surface of TN3, forming contacts to Glu120, Lys148, Asn106 of a symmetry-related molecule, and to an ethanol molecule....

  18. submitter Clinical evaluation of the radiolanthanide terbium-152: first-in-human PET/CT with $^152$Tb-DOTATOC

    CERN Document Server

    Baum, Richard P; Benešová, Martina; Vermeulen, Christiaan; Gnesin, Silvano; Köster, Ulli; Johnston, Karl; Müller, Dirk; Senftleben, Stefan; Kulkarni, Harshad R; Türler, Andreas; Schibli, Roger; Prior, John O; van der Meulen, Nicholas P; Müller, Cristina

    2017-01-01

    The existence of theragnostic pairs of radionuclides allows the preparation of radiopharmaceuticals for diagnostic and therapeutic purposes. Radiolanthanides, such as $^{177}$Lu, are successfully used for therapeutic purposes; however, a perfect diagnostic match is currently not available for clinical use. A unique, multi-disciplinary study was performed using $^{152}$Tb ($T_{1/2}$ = 17.5 h, E$\\beta ^+_{average}$ = 1140 keV, I$β^+$ = 20.3%), which resulted in the first-in-human PET/CT images with this promising radionuclide. For this purpose, $^{152}$Tb was produced via a spallation process followed by mass separation at ISOLDE, CERN. The chemical separation and quality control, performed at PSI, resulted in a pure product in sufficient yields. Clinical PET phantom studies revealed an increased image noise level, because of the smaller $\\beta^+$ branching ratio of $^{152}$Tb as compared to standard PET nuclides at matched activity concentrations; however, the expected recovery would be comparable at matched ...

  19. Site-specific chemical conjugation of human Fas ligand extracellular domain using trans-cyclooctene - methyltetrazine reactions.

    Science.gov (United States)

    Muraki, Michiro; Hirota, Kiyonori

    2017-07-03

    Fas ligand plays a key role in the human immune system as a major cell death inducing protein. The extracellular domain of human Fas ligand (hFasLECD) triggers apoptosis of malignant cells, and therefore is expected to have substantial potentials in medical biotechnology. However, the current application of this protein to clinical medicine is hampered by a shortage of the benefits relative to the drawbacks including the side-effects in systemic administration. Effective procedures for the engineering of the protein by attaching useful additional functions are required to overcome the problem. A procedure for the site-specific chemical conjugation of hFasLECD with a fluorochrome and functional proteins was devised using an inverse-electron-demand Diels-Alder reaction between trans-cyclooctene group and methyltetrazine group. The conjugations in the present study were attained by using much less molar excess amounts of the compounds to be attached as compared with the conventional chemical modification reactions using maleimide derivatives in the previous study. The isolated conjugates of hFasLECD with sulfo-Cy3, avidin and rabbit IgG Fab' domain presented the functional and the structural integrities of the attached molecules without impairing the specific binding activity toward human Fas receptor extracellular domain. The present study provided a new fundamental strategy for the production of the engineered hFasLECDs with additional beneficial functions, which will lead to the developments of the improved diagnostic systems and the effective treatment methods of serious diseases by using this protein as a component of novel molecular tools.

  20. A survey on using domain and contextual knowledge for human activity recognition in video streams

    NARCIS (Netherlands)

    Onofri, L.; Soda, P.; Pechenizkiy, M.; Iannello, G.

    2016-01-01

    Human activity recognition has gained an increasing relevance in computer vision and it can be tackled with either non-hierarchical or hierarchical approaches. The former, also known as single-layered approaches, are those that represent and recognize human activities directly from the extracted

  1. Immunohistochemical Localization of Fibrinogen C Domain Containing 1 on Epithelial and Mucosal Surfaces in Human Tissues

    DEFF Research Database (Denmark)

    von Huth, Sebastian; Moeller, Jesper B; Schlosser, Anders

    2018-01-01

    Fibrinogen C domain containing 1 (FIBCD1) is a transmembrane receptor that binds chitin and other acetylated compounds with high affinity. FIBCD1 has previously been shown to be present in the epithelium of the gastrointestinal tract. In the present study, we performed a detailed analysis...... high expression of FIBCD1 and also mesodermal-derived cells in the genitourinary system and ectodermal-derived epidermis and sebaceous glands cells expressed FIBCD1. In some columnar epithelial cells, for example, in the salivary gland and gall bladder, the FIBCD1 expression was clearly polarized...

  2. Determinants in the Ig Variable Domain of Human HAVCR1 (TIM-1) Are Required To Enhance Hepatitis C Virus Entry.

    Science.gov (United States)

    Kachko, Alla; Costafreda, Maria Isabel; Zubkova, Iryna; Jacques, Jerome; Takeda, Kazuyo; Wells, Frances; Kaplan, Gerardo; Major, Marian E

    2018-03-15

    Hepatitis C virus (HCV) is the leading cause of chronic hepatitis in humans. Several host molecules participate in HCV cell entry, but this process remains unclear. The complete unraveling of the HCV entry process is important to further understand viral pathogenesis and develop therapeutics. Human hepatitis A virus (HAV) cellular receptor 1 (HAVCR1), CD365, also known as TIM-1, functions as a phospholipid receptor involved in cell entry of several enveloped viruses. Here, we studied the role of HAVCR1 in HCV infection. HAVCR1 antibody inhibited entry in a dose-dependent manner. HAVCR1 soluble constructs neutralized HCV, which did not require the HAVCR1 mucinlike region and was abrogated by a mutation of N to A at position 94 (N94A) in the Ig variable (IgV) domain phospholipid-binding pocket, indicating a direct interaction of the HAVCR1 IgV domain with HCV virions. However, knockout of HAVCR1 in Huh7 cells reduced but did not prevent HCV growth. Interestingly, the mouse HAVCR1 ortholog, also a phospholipid receptor, did not enhance infection and a soluble form failed to neutralize HCV, although replacement of the mouse IgV domain with the human HAVCR1 IgV domain restored the enhancement of HCV infection. Mutations in the cytoplasmic tail revealed that direct HAVCR1 signaling is not required to enhance HCV infection. Our data show that the phospholipid-binding function and other determinant(s) in the IgV domain of human HAVCR1 enhance HCV infection. Although the exact mechanism is not known, it is possible that HAVCR1 facilitates entry by stabilizing or enhancing attachment, leading to direct interactions with specific receptors, such as CD81. IMPORTANCE Hepatitis C virus (HCV) enters cells through a multifaceted process. We identified the human hepatitis A virus cellular receptor 1 (HAVCR1), CD365, also known as TIM-1, as a facilitator of HCV entry. Antibody blocking and silencing or knockout of HAVCR1 in hepatoma cells reduced HCV entry. Our findings that the

  3. The 1.7 Å X-ray crystal structure of the porcine factor VIII C2 domain and binding analysis to anti-human C2 domain antibodies and phospholipid surfaces.

    Directory of Open Access Journals (Sweden)

    Caileen M Brison

    Full Text Available The factor VIII C2 domain is essential for binding to activated platelet surfaces as well as the cofactor activity of factor VIII in blood coagulation. Inhibitory antibodies against the C2 domain commonly develop following factor VIII replacement therapy for hemophilia A patients, or they may spontaneously arise in cases of acquired hemophilia. Porcine factor VIII is an effective therapeutic for hemophilia patients with inhibitor due to its low cross-reactivity; however, the molecular basis for this behavior is poorly understood. In this study, the X-ray crystal structure of the porcine factor VIII C2 domain was determined, and superposition of the human and porcine C2 domains demonstrates that most surface-exposed differences cluster on the face harboring the "non-classical" antibody epitopes. Furthermore, antibody-binding results illustrate that the "classical" 3E6 antibody can bind both the human and porcine C2 domains, although the inhibitory titer to human factor VIII is 41 Bethesda Units (BU/mg IgG versus 0.8 BU/mg IgG to porcine factor VIII, while the non-classical G99 antibody does not bind to the porcine C2 domain nor inhibit porcine factor VIII activity. Further structural analysis of differences between the electrostatic surface potentials suggest that the C2 domain binds to the negatively charged phospholipid surfaces of activated platelets primarily through the 3E6 epitope region. In contrast, the G99 face, which contains residue 2227, should be distal to the membrane surface. Phospholipid binding assays indicate that both porcine and human factor VIII C2 domains bind with comparable affinities, and the human K2227A and K2227E mutants bind to phospholipid surfaces with similar affinities as well. Lastly, the G99 IgG bound to PS-immobilized factor VIII C2 domain with an apparent dissociation constant of 15.5 nM, whereas 3E6 antibody binding to PS-bound C2 domain was not observed.

  4. [Expression, crystallization and crystallographic study of the 1st IgV domain of human CD96].

    Science.gov (United States)

    Jiang, Wenjing; Zhang, Shuijun; Yan, Jinghua; Guo, Ning

    2013-05-01

    CD96 (Tactile) is an adhesion receptor expressed mainly on activated T cells, NK cells. As a family member of the immunoglobulin-like cell receptor, CD96 consists of three immunoglobulin-like domains (V1, V2/C and C) in the extracellular region. Recent studies have shown that the 1st IgV domain of CD96 (CD96V1) plays an essential role in cell adhesion and NK cell-mediated killing. In this study, the 1st IgV domain of human CD96 (hCD96V1) was cloned and expressed in Escherichia coli (BL21). The soluble protein was obtained by refolding of the hCD96V1 inclusion bodies. From analytical ultracentrifugation, we could predict that CD96 V1 maily exists as dimer with approximate molecular weight of 26.9 kDa. The protein was then successfully crystallized using the sitting-drop vapour-diffusion method. The crystals diffracted to 1.9 angstrom resolution and belonged to space group P21, with unit-cell parameters a = 35.1, b = 69.5, c = 49.6A, alpha=gamma=90 degrees, beta=105.4 degrees.

  5. The human 64-kDa polyadenylylation factor contains a ribonucleoprotein-type RNA binding domain and unusual auxiliary motifs

    International Nuclear Information System (INIS)

    Takagaki, Yoshio; Manley, J.L.; MacDonald, C.C.; Shenk, T.

    1992-01-01

    Cleavage stimulation factor is one of the multiple factors required for 3'-end cleavage of mammalian pre-mRNAs. The authors have shown previously that this factor is composed of three subunits with estimated molecular masses of 77, 64, and 50 kDa and that the 64-kDa subunit can be UV-cross linked to RNA in a polyadenylylation signal (AAUAAA)-dependent manner. They have now isolated cDNAs encoding the 64-kDa subunit of human cleavage stimulation factor. The 64-kDa subunit contains a ribonucleoprotein-type RNA binding domain in the N-terminal region and a repeat structure in the C-terminal region in which a pentapeptide sequence (consensus MEARA/G) is repeated 12 times and the formation of a long α-helix stabilized by salt bridges is predicted. An ∼270-amino acid segment surrounding this repeat structure is highly enriched in proline and glycine residues (∼20% for each). When cloned 64-kDa subunit was expressed in Escherichia coli, an N-terminal fragment containing the RNA binding domain bound to RNAs in a polyadenylylation-signal-independent manner, suggesting that the RNA binding domain is directly involved in the binding of the 64-kDa subunit to pre-mRNAs

  6. The B-subdomain of the Xenopus laevis XFIN KRAB-AB domain is responsible for its weaker transcriptional repressor activity compared to human ZNF10/Kox1.

    Science.gov (United States)

    Born, Nadine; Thiesen, Hans-Jürgen; Lorenz, Peter

    2014-01-01

    The Krüppel-associated box (KRAB) domain interacts with the nuclear hub protein TRIM28 to initiate or mediate chromatin-dependent processes like transcriptional repression, imprinting or suppression of endogenous retroviruses. The prototype KRAB domain initially identified in ZNF10/KOX1 encompasses two subdomains A and B that are found in hundreds of zinc finger transcription factors studied in human and murine genomes. Here we demonstrate for the first time transcriptional repressor activity of an amphibian KRAB domain. After sequence correction, the updated KRAB-AB domain of zinc finger protein XFIN from the frog Xenopus laevis was found to confer transcriptional repression in reporter assays in Xenopus laevis A6 kidney cells as well as in human HeLa, but not in the minnow Pimephales promelas fish cell line EPC. Binding of the XFIN KRAB-AB domain to human TRIM28 was demonstrated in a classical co-immunoprecipitation approach and visualized in a single-cell compartmentalization assay. XFIN-AB displayed reduced potency in repression as well as lower strength of interaction with TRIM28 compared to ZNF10 KRAB-AB. KRAB-B subdomain swapping between the two KRAB domains indicated that it was mainly the KRAB-B subdomain of XFIN that was responsible for its lower capacity in repression and binding to human TRIM28. In EPC fish cells, ZNF10 and XFIN KRAB repressor activity could be partially restored to low levels by adding exogenous human TRIM28. In contrast to XFIN, we did not find any transcriptional repression activity for the KRAB-like domain of human PRDM9 in HeLa cells. PRDM9 is thought to harbor an evolutionary older domain related to KRAB whose homologs even occur in invertebrates. Our results support the notion that functional bona fide KRAB domains which confer transcriptional repression and interact with TRIM28 most likely co-evolved together with TRIM28 at the beginning of tetrapode evolution.

  7. Delectability of low contrast in CT. Comparison between human observes and observer model

    International Nuclear Information System (INIS)

    Hernandez-Giron, I.; Geleijins, J.; Calzado, A.; Joemai, R. M. S.; Veldkamp, W. J. H.

    2013-01-01

    The objective of this work is to study the real images of TC and other simulated LCD with white noise through a model of observer. The results are compared with those obtained in a similar experiment by human observers. (Author)

  8. Purification, crystallization and preliminary X-ray diffraction study of human ribosomal protein L10 core domain

    International Nuclear Information System (INIS)

    Nishimura, Mitsuhiro; Kaminishi, Tatsuya; Kawazoe, Masahito; Shirouzu, Mikako; Takemoto, Chie; Yokoyama, Shigeyuki; Tanaka, Akiko; Sugano, Sumio; Yoshida, Takuya; Ohkubo, Tadayasu; Kobayashi, Yuji

    2007-01-01

    A truncated variant of human ribosomal protien L10 was prepared and crystallized. Diffraction data were collected to 2.5 Å resolution. Eukaryotic ribosomal protein L10 is an essential component of the large ribosomal subunit, which organizes the architecture of the aminoacyl-tRNA binding site. The human L10 protein is also called the QM protein and consists of 214 amino-acid residues. For crystallization, the L10 core domain (L10CD, Phe34–Glu182) was recombinantly expressed in Escherichia coli and purified to homogeneity. A hexagonal crystal of L10CD was obtained by the sitting-drop vapour-diffusion method. The L10CD crystal diffracted to 2.5 Å resolution and belongs to space group P3 1 21 or P3 2 21

  9. Methods to uncover an antibody epitope in the KPI domain of Alzheimer's amyloid precursor protein for immunohistochemistry in human brain.

    Science.gov (United States)

    Campbell, E; Pearson, R C; Parkinson, D

    1999-11-15

    A novel polyclonal antibody (Ab993), specific for a KPI domain epitope of APP, was characterised for use in immunoprecipitation, Western blotting and immunohistochemistry. Conditioned medium from NTera2/D1 cells was used for immunoprecipitation and Western blots. Paraffin-embedded human brain sections were used for immunohistochemistry. The antibody recognised KPI-containing APP on Western blots after standard solubilisation but immunoprecipitation of soluble APP required reduction with 2-mercaptoethanol followed by alkylation of reduced sulphydryl bonds with sodium iodoacetate. Immunohistochemical staining of human brain sections was significantly enhanced by this pre-treatment. Microwaving of sections also increased immunolabelling, by a mechanism that was additive to reduction and alkylation. Incubation in 80% formic acid did not confer any enhancement of immunoreactivity. Ab993, applied with the methods reported here, is expected to be valuable in investigations of the pathogenesis of Alzheimer's disease to determine the source of the beta-amyloid peptide.

  10. Automated segmentation of muscle and adipose tissue on CT images for human body composition analysis

    Science.gov (United States)

    Chung, Howard; Cobzas, Dana; Birdsell, Laura; Lieffers, Jessica; Baracos, Vickie

    2009-02-01

    The ability to compute body composition in cancer patients lends itself to determining the specific clinical outcomes associated with fat and lean tissue stores. For example, a wasting syndrome of advanced disease associates with shortened survival. Moreover, certain tissue compartments represent sites for drug distribution and are likely determinants of chemotherapy efficacy and toxicity. CT images are abundant, but these cannot be fully exploited unless there exist practical and fast approaches for tissue quantification. Here we propose a fully automated method for segmenting muscle, visceral and subcutaneous adipose tissues, taking the approach of shape modeling for the analysis of skeletal muscle. Muscle shape is represented using PCA encoded Free Form Deformations with respect to a mean shape. The shape model is learned from manually segmented images and used in conjunction with a tissue appearance prior. VAT and SAT are segmented based on the final deformed muscle shape. In comparing the automatic and manual methods, coefficients of variation (COV) (1 - 2%), were similar to or smaller than inter- and intra-observer COVs reported for manual segmentation.

  11. Expression, purification, crystallization and preliminary X-ray diffraction analysis of the aspartate transcarbamoylase domain of human CAD

    International Nuclear Information System (INIS)

    Ruiz-Ramos, Alba; Lallous, Nada; Grande-García, Araceli; Ramón-Maiques, Santiago

    2013-01-01

    The recombinant aspartate transcarbamoylase domain of human CAD was expressed in E. coli, purified and crystallized in the presence and absence of the inhibitor PALA. X-ray diffraction data sets were collected for both crystal forms at 2.1 Å resolution. Aspartate transcarbamoylase (ATCase) catalyzes the synthesis of N-carbamoyl-l-aspartate from carbamoyl phosphate and aspartate in the second step of the de novo biosynthesis of pyrimidines. In prokaryotes, the first three activities of the pathway, namely carbamoyl phosphate synthetase (CPSase), ATCase and dihydroorotase (DHOase), are encoded as distinct proteins that function independently or in noncovalent association. In animals, CPSase, ATCase and DHOase are part of a 243 kDa multifunctional polypeptide named CAD. Up-regulation of CAD is essential for normal and tumour cell proliferation. Although the structures of numerous prokaryotic ATCases have been determined, there is no structural information about any eukaryotic ATCase. In fact, the only detailed structural information about CAD is that it self-assembles into hexamers and trimers through interactions of the ATCase domains. Here, the expression, purification and crystallization of the ATCase domain of human CAD is reported. The recombinant protein, which was expressed in bacteria and purified with good yield, formed homotrimers in solution. Crystallization experiments both in the absence and in the presence of the inhibitor PALA yielded small crystals that diffracted X-rays to 2.1 Å resolution using synchrotron radiation. The crystals appeared to belong to the hexagonal space group P6 3 22, and Matthews coefficient calculation indicated the presence of one ATCase subunit per asymmetric unit, with a solvent content of 48%. However, analysis of the intensity statistics suggests a special case of the P2 1 lattice with pseudo-symmetry and possibly twinning

  12. Unexpected role of the L-domain of calpastatin during the autoproteolytic activation of human erythrocyte calpain.

    Science.gov (United States)

    De Tullio, Roberta; Franchi, Alice; Martines, Antonino; Averna, Monica; Pedrazzi, Marco; Melloni, Edon; Sparatore, Bianca

    2018-04-26

    Autoproteolysis of human erythrocyte calpain-1 proceeds in vitro at high [Ca 2+ ], through the conversion of the 80-kDa catalytic subunit into a 75-kDa activated enzyme that requires lower [Ca 2+ ] for catalysis. Importantly, here we detect a similar 75 kDa calpain-1 form also in vivo , in human meningiomas. Although calpastatin is so far considered the specific inhibitor of calpains, we have previously identified in rat brain a calpastatin transcript truncated at the end of the L-domain (cast110, L-DOM), coding for a protein lacking the inhibitory units. Aim of the present study was to characterize the possible biochemical role of the L-DOM during calpain-1 autoproteolysis in vitro , at high (100 µM) and low (5 µM) [Ca 2+ ]. Here we demonstrate that the L-DOM binds the 80 kDa proenzyme in the absence of Ca 2+ Consequently, we have explored the ability of the 75 kDa activated protease to catalyze at 5 µM Ca 2+ the intermolecular activation of native calpain-1 associated with the L-DOM. Notably, this [Ca 2+ ] is too low to promote the autoproteolytic activation of calpain-1 but enough to support the catalysis of the 75 kDa calpain. We show for the first time that the L-DOM preserves native calpain-1 from the degradation mediated by the 75 kDa form. Taken together, our data suggest that the free L-domain of calpastatin is a novel member of the calpain/calpastatin system endowed with a function alternative to calpain inhibition. For this reason, it will be crucial to define the intracellular relevance of the L-domain in controlling calpain activation/activity in physiopathological conditions having altered Ca 2+ homeostasis. © 2018 The Author(s).

  13. The crystal structure of the Dachshund domain of human SnoN reveals flexibility in the putative protein interaction surface.

    Directory of Open Access Journals (Sweden)

    Tomas Nyman

    2010-09-01

    Full Text Available The human SnoN is an oncoprotein that interacts with several transcription-regulatory proteins such as the histone-deacetylase, N-CoR containing co-repressor complex and Smad proteins. This study presents the crystal structure of the Dachshund homology domain of human SnoN. The structure reveals a groove composed of conserved residues with characteristic properties of a protein-interaction surface. A comparison of the 12 monomers in the asymmetric unit reveals the presence of two major conformations: an open conformation with a well accessible groove and a tight conformation with a less accessible groove. The variability in the backbone between the open and the tight conformations matches the differences seen in previously determined structures of individual Dachshund homology domains, suggesting a general plasticity within this fold family. The flexibility observed in the putative protein binding groove may enable SnoN to recognize multiple interaction partners.This article can also be viewed as an enhanced version in which the text of the article is integrated with interactive 3D representations and animated transitions. Please note that a web plugin is required to access this enhanced functionality. Instructions for the installation and use of the web plugin are available in Text S1.

  14. The N-terminal domain of human DNA helicase Rtel1 contains a redox active iron-sulfur cluster.

    Science.gov (United States)

    Landry, Aaron P; Ding, Huangen

    2014-01-01

    Human telomere length regulator Rtel1 is a superfamily II DNA helicase and is essential for maintaining proper length of telomeres in chromosomes. Here we report that the N-terminal domain of human Rtel1 (RtelN) expressed in Escherichia coli cells produces a protein that contains a redox active iron-sulfur cluster with the redox midpoint potential of -248 ± 10 mV (pH 8.0). The iron-sulfur cluster in RtelN is sensitive to hydrogen peroxide and nitric oxide, indicating that reactive oxygen/nitrogen species may modulate the DNA helicase activity of Rtel1 via modification of its iron-sulfur cluster. Purified RtelN retains a weak binding affinity for the single-stranded (ss) and double-stranded (ds) DNA in vitro. However, modification of the iron-sulfur cluster by hydrogen peroxide or nitric oxide does not significantly affect the DNA binding activity of RtelN, suggesting that the iron-sulfur cluster is not directly involved in the DNA interaction in the N-terminal domain of Rtel1.

  15. The N-Terminal Domain of Human DNA Helicase Rtel1 Contains a Redox Active Iron-Sulfur Cluster

    Directory of Open Access Journals (Sweden)

    Aaron P. Landry

    2014-01-01

    Full Text Available Human telomere length regulator Rtel1 is a superfamily II DNA helicase and is essential for maintaining proper length of telomeres in chromosomes. Here we report that the N-terminal domain of human Rtel1 (RtelN expressed in Escherichia coli cells produces a protein that contains a redox active iron-sulfur cluster with the redox midpoint potential of −248 ± 10 mV (pH 8.0. The iron-sulfur cluster in RtelN is sensitive to hydrogen peroxide and nitric oxide, indicating that reactive oxygen/nitrogen species may modulate the DNA helicase activity of Rtel1 via modification of its iron-sulfur cluster. Purified RtelN retains a weak binding affinity for the single-stranded (ss and double-stranded (ds DNA in vitro. However, modification of the iron-sulfur cluster by hydrogen peroxide or nitric oxide does not significantly affect the DNA binding activity of RtelN, suggesting that the iron-sulfur cluster is not directly involved in the DNA interaction in the N-terminal domain of Rtel1.

  16. Structural insights into RISC assembly facilitated by dsRNA-binding domains of human RNA helicase A (DHX9).

    Science.gov (United States)

    Fu, Qinqin; Yuan, Y Adam

    2013-03-01

    Intensive research interest has focused on small RNA-processing machinery and the RNA-induced silencing complex (RISC), key cellular machines in RNAi pathways. However, the structural mechanism regarding RISC assembly, the primary step linking small RNA processing and RNA-mediated gene silencing, is largely unknown. Human RNA helicase A (DHX9) was reported to function as an RISC-loading factor, and such function is mediated mainly by its dsRNA-binding domains (dsRBDs). Here, we report the crystal structures of human RNA helicase A (RHA) dsRBD1 and dsRBD2 domains in complex with dsRNAs, respectively. Structural analysis not only reveals higher siRNA duplex-binding affinity displayed by dsRBD1, but also identifies a crystallographic dsRBD1 pair of physiological significance in cooperatively recognizing dsRNAs. Structural observations are further validated by isothermal titration calorimetric (ITC) assay. Moreover, co-immunoprecipitation (co-IP) assay coupled with mutagenesis demonstrated that both dsRBDs are required for RISC association, and such association is mediated by dsRNA. Hence, our structural and functional efforts have revealed a potential working model for siRNA recognition by RHA tandem dsRBDs, and together they provide direct structural insights into RISC assembly facilitated by RHA.

  17. 18F-Alfatide II PET/CT in healthy human volunteers and patients with brain metastases

    International Nuclear Information System (INIS)

    Yu, Chunjing; Mi, Baoming; Wan, Weixing; Pan, Donghui; Xu, Yuping; Yang, Min; Lang, Lixin; Niu, Gang; Chen, Xiaoyuan

    2015-01-01

    We report the biodistribution and radiation dosimetry of an integrin α v β 3 specific PET tracer 18 F-AlF-NOTA-E[PEG 4 -c(RGDfk)] 2 (denoted as 18 F-Alfatide II). We also assessed the value of 18 F-Alfatide II in patients with brain metastases. A series of torso (from the skull to the thigh) static images were acquired in five healthy volunteers (3 M, 2 F) at 5, 10, 15, 30, 45, and 60 min after injection of 18 F-Alfatide II (257 ± 48 MBq). Regions of interest (ROIs) were drawn manually, and the time-activity curves (TACs) were obtained for major organs. Nine patients with brain metastases were examined by static PET imaging with 18 F-FDG (5.55 MBq/kg) and 18 F-Alfatide II. Injection of 18 F-Alfatide II was well tolerated in all healthy volunteers, with no serious tracer-related adverse events found. 18 F-Alfatide II showed rapid clearance from the blood pool and kidneys. The total effective dose equivalent (EDE) and effective dose (ED) were 0.0277 ± 0.003 mSv/MBq and 0.0198 ± 0.002 mSv/MBq, respectively. The organs with the highest absorbed dose were the kidneys and the spleen. Nine patients with 20 brain metastatic lesions identified by MRI and/or CT were enrolled in this study. All 20 brain lesions were visualized by 18 F-Alfatide II PET, while only ten lesions were visualized by 18 F-FDG, and 13 by CT. F-Alfatide II is a safe PET tracer with a favorable dosimetry profile. The observed ED suggests that 18 F-Alfatide II is feasible for human studies. 18 F-Alfatide II has potential value in finding brain metastases of different cancers as a biomarker of angiogenesis. (orig.)

  18. Experimental demonstration of passive acoustic imaging in the human skull cavity using CT-based aberration corrections.

    Science.gov (United States)

    Jones, Ryan M; O'Reilly, Meaghan A; Hynynen, Kullervo

    2015-07-01

    Experimentally verify a previously described technique for performing passive acoustic imaging through an intact human skull using noninvasive, computed tomography (CT)-based aberration corrections Jones et al. [Phys. Med. Biol. 58, 4981-5005 (2013)]. A sparse hemispherical receiver array (30 cm diameter) consisting of 128 piezoceramic discs (2.5 mm diameter, 612 kHz center frequency) was used to passively listen through ex vivo human skullcaps (n = 4) to acoustic emissions from a narrow-band fixed source (1 mm diameter, 516 kHz center frequency) and from ultrasound-stimulated (5 cycle bursts, 1 Hz pulse repetition frequency, estimated in situ peak negative pressure 0.11-0.33 MPa, 306 kHz driving frequency) Definity™ microbubbles flowing through a thin-walled tube phantom. Initial in vivo feasibility testing of the method was performed. The performance of the method was assessed through comparisons to images generated without skull corrections, with invasive source-based corrections, and with water-path control images. For source locations at least 25 mm from the inner skull surface, the modified reconstruction algorithm successfully restored a single focus within the skull cavity at a location within 1.25 mm from the true position of the narrow-band source. The results obtained from imaging single bubbles are in good agreement with numerical simulations of point source emitters and the authors' previous experimental measurements using source-based skull corrections O'Reilly et al. [IEEE Trans. Biomed. Eng. 61, 1285-1294 (2014)]. In a rat model, microbubble activity was mapped through an intact human skull at pressure levels below and above the threshold for focused ultrasound-induced blood-brain barrier opening. During bursts that led to coherent bubble activity, the location of maximum intensity in images generated with CT-based skull corrections was found to deviate by less than 1 mm, on average, from the position obtained using source-based corrections. Taken

  19. The study on three-dimensional measurement of the Human Eustachian tube examined by Multislice CT

    International Nuclear Information System (INIS)

    Yoshioka, Satoshi; Naito, Kensei; Fujii, Naoko; Katada, Kazuhiro; Takeuchi, Kenji

    2007-01-01

    Morphological aberration of the Eustachian tube is a significant factor of various middle ear diseases. Traditionally, cadaveric specimens have been used for studies on the morphology of Eustachian tubes. However, this approach was not too efficient, as samples were limited in number as they were difficult to obtain, and biological conditions were not reflected due to rigor mortis and atrophy during specimen preparation. We thus decided to use Multi-Slice CT (MSCT) to perform 3-dimensional (3-D) anatomic measurements of the Eustachian tube. MSCT has benefits of isotropy and high resolution, and it is useful in preparing images of any plane. Forty-eight adults were studied. For the purpose of measurement, various anatomic indices were carefully and precisely defined to identify each area on the image. Calculations based on each coordinate value enabled the measurement of length, diameter and angle of the Eustachian tube of normal adults. Therefore, measurements of the Eustachian tube, which were traditionally difficult as it is located in the deep part of the cranium, were simplified in many specimens. Mean value of total length was 39.2±3.2 mm, cartilage part length of the tube 30.0±2.7 mm and bony part 9.2±1.6 mm. Mean values of diameter of tympanic orifice were 5.2 x 3.2 mm, and pharyngeal orifice 9.7 x 4.4 mm. Mean value of angle between bony part and cartilage part was 160.9±13.6 degrees. This approach to anatomic measurement is expected to contribute greatly to investigation on various middle ear diseases. (author)

  20. Construction of a novel lentiviral vector carrying human B-domain ...

    African Journals Online (AJOL)

    ... integration were detected in all cell lines after transfection. A novel lentiviral vector carrying human FVIII³BD was constructed, which was able to transfect different mammalian cell types accompanied by high-level activity. This lentiviral vector may provide a theoretical basis for the gene therapy of patients with hemophilia ...

  1. Engineering planetary exploration systems : Integrating novel technologies and the human element using work domain analysis

    NARCIS (Netherlands)

    Baker, C.; Naikar, N.; Neerincx, M.

    2008-01-01

    The realisation of sustainable space exploration and utilisation requires not only the development of novel concepts and technologies, but also their successful integration. Hardware, software, and the human element must be integrated effectively to make the dream for which these technologies were

  2. Domain Decomposition for Computing Extremely Low Frequency Induced Current in the Human Body

    OpenAIRE

    Perrussel , Ronan; Voyer , Damien; Nicolas , Laurent; Scorretti , Riccardo; Burais , Noël

    2011-01-01

    International audience; Computation of electromagnetic fields in high resolution computational phantoms requires solving large linear systems. We present an application of Schwarz preconditioners with Krylov subspace methods for computing extremely low frequency induced fields in a phantom issued from the Visible Human.

  3. Human SIRT6 promotes DNA end resection through CtIP deacetylation

    DEFF Research Database (Denmark)

    Kaidi, Abderrahmane; Weinert, Brian T; Choudhary, Chunaram

    2010-01-01

    SIRT6 belongs to the sirtuin family of protein lysine deacetylases, which regulate aging and genome stability. We found that human SIRT6 has a role in promoting DNA end resection, a crucial step in DNA double-strand break (DSB) repair by homologous recombination. SIRT6 depletion impaired the accu...

  4. Human mitochondrial Hsp70 (mortalin): shedding light on ATPase activity, interaction with adenosine nucleotides, solution structure and domain organization.

    Science.gov (United States)

    Dores-Silva, Paulo R; Barbosa, Leandro R S; Ramos, Carlos H I; Borges, Júlio C

    2015-01-01

    The human mitochondrial Hsp70, also called mortalin, is of considerable importance for mitochondria biogenesis and the correct functioning of the cell machinery. In the mitochondrial matrix, mortalin acts in the importing and folding process of nucleus-encoded proteins. The in vivo deregulation of mortalin expression and/or function has been correlated with age-related diseases and certain cancers due to its interaction with the p53 protein. In spite of its critical biological roles, structural and functional studies on mortalin are limited by its insoluble recombinant production. This study provides the first report of the production of folded and soluble recombinant mortalin when co-expressed with the human Hsp70-escort protein 1, but it is still likely prone to self-association. The monomeric fraction of mortalin presented a slightly elongated shape and basal ATPase activity that is higher than that of its cytoplasmic counterpart Hsp70-1A, suggesting that it was obtained in the functional state. Through small angle X-ray scattering, we assessed the low-resolution structural model of monomeric mortalin that is characterized by an elongated shape. This model adequately accommodated high resolution structures of Hsp70 domains indicating its quality. We also observed that mortalin interacts with adenosine nucleotides with high affinity. Thermally induced unfolding experiments indicated that mortalin is formed by at least two domains and that the transition is sensitive to the presence of adenosine nucleotides and that this process is dependent on the presence of Mg2+ ions. Interestingly, the thermal-induced unfolding assays of mortalin suggested the presence of an aggregation/association event, which was not observed for human Hsp70-1A, and this finding may explain its natural tendency for in vivo aggregation. Our study may contribute to the structural understanding of mortalin as well as to contribute for its recombinant production for antitumor compound screenings.

  5. Viral RNA annealing activities of human immunodeficiency virus type 1 nucleocapsid protein require only peptide domains outside the zinc fingers.

    Science.gov (United States)

    De Rocquigny, H; Gabus, C; Vincent, A; Fournié-Zaluski, M C; Roques, B; Darlix, J L

    1992-07-15

    The nucleocapsid (NC) of human immunodeficiency virus type 1 consists of a large number of NC protein molecules, probably wrapping the dimeric RNA genome within the virion inner core. NC protein is a gag-encoded product that contains two zinc fingers flanked by basic residues. In human immunodeficiency virus type 1 virions, NCp15 is ultimately processed into NCp7 and p6 proteins. During virion assembly the retroviral NC protein is necessary for core formation and genomic RNA encapsidation, which are essential for virus infectivity. In vitro NCp15 activates viral RNA dimerization, a process most probably linked in vivo to genomic RNA packaging, and replication primer tRNA(Lys,3) annealing to the initiation site of reverse transcription. To characterize the domains of human immunodeficiency virus type 1 NC protein necessary for its various functions, the 72-amino acid NCp7 and several derived peptides were synthesized in a pure form. We show here that synthetic NCp7 with or without the two zinc fingers has the RNA annealing activities of NCp15. Further deletions of the N-terminal 12 and C-terminal 8 amino acids, leading to a 27-residue peptide lacking the finger domains, have little or no effect on NC protein activity in vitro. However deletion of short sequences containing basic residues flanking the first finger leads to a complete loss of NC protein activity. It is proposed that the basic residues and the zinc fingers cooperate to select and package the genomic RNA in vivo. Inhibition of the viral RNA binding and annealing activities associated with the basic residues flanking the first zinc finger of NC protein could therefore be used as a model for the design of antiviral agents.

  6. Domains and domain loss

    DEFF Research Database (Denmark)

    Haberland, Hartmut

    2005-01-01

    politicians and in the media, especially in the discussion whether some languages undergo ‘domain loss’ vis-à-vis powerful international languages like English. An objection that has been raised here is that domains, as originally conceived, are parameters of language choice and not properties of languages...

  7. Unique biological properties of catalytic domain directed human anti-CAIX antibodies discovered through phage-display technology.

    Directory of Open Access Journals (Sweden)

    Chen Xu

    2010-03-01

    Full Text Available Carbonic anhydrase IX (CAIX, gene G250/MN-encoded transmembrane protein is highly expressed in various human epithelial tumors such as renal clear cell carcinoma (RCC, but absent from the corresponding normal tissues. Besides the CA signal transduction activity, CAIX may serve as a biomarker in early stages of oncogenesis and also as a reliable marker of hypoxia, which is associated with tumor resistance to chemotherapy and radiotherapy. Although results from preclinical and clinical studies have shown CAIX as a promising target for detection and therapy for RCC, only a limited number of murine monoclonal antibodies (mAbs and one humanized mAb are available for clinical testing and development. In this study, paramagnetic proteoliposomes of CAIX (CAIX-PMPLs were constructed and used for anti-CAIX antibody selection from our 27 billion human single-chain antibody (scFv phage display libraries. A panel of thirteen human scFvs that specifically recognize CAIX expressed on cell surface was identified, epitope mapped primarily to the CA domain, and affinity-binding constants (KD determined. These human anti-CAIX mAbs are diverse in their functions including induction of surface CAIX internalization into endosomes and inhibition of the carbonic anhydrase activity, the latter being a unique feature that has not been previously reported for anti-CAIX antibodies. These human anti-CAIX antibodies are important reagents for development of new immunotherapies and diagnostic tools for RCC treatment as well as extending our knowledge on the basic structure-function relationships of the CAIX molecule.

  8. Low statistical power in biomedical science: a review of three human research domains

    Science.gov (United States)

    Dumas-Mallet, Estelle; Button, Katherine S.; Boraud, Thomas; Gonon, Francois

    2017-01-01

    Studies with low statistical power increase the likelihood that a statistically significant finding represents a false positive result. We conducted a review of meta-analyses of studies investigating the association of biological, environmental or cognitive parameters with neurological, psychiatric and somatic diseases, excluding treatment studies, in order to estimate the average statistical power across these domains. Taking the effect size indicated by a meta-analysis as the best estimate of the likely true effect size, and assuming a threshold for declaring statistical significance of 5%, we found that approximately 50% of studies have statistical power in the 0–10% or 11–20% range, well below the minimum of 80% that is often considered conventional. Studies with low statistical power appear to be common in the biomedical sciences, at least in the specific subject areas captured by our search strategy. However, we also observe evidence that this depends in part on research methodology, with candidate gene studies showing very low average power and studies using cognitive/behavioural measures showing high average power. This warrants further investigation. PMID:28386409

  9. Structure of the SPRY domain of the human RNA helicase DDX1, a putative interaction platform within a DEAD-box protein

    Energy Technology Data Exchange (ETDEWEB)

    Kellner, Julian N.; Meinhart, Anton, E-mail: anton.meinhart@mpimf-heidelberg.mpg.de [Max Planck Institute for Medical Research, Jahnstrasse 29, 69120 Heidelberg (Germany)

    2015-08-25

    The structure of the SPRY domain of the human RNA helicase DDX1 was determined at 2.0 Å resolution. The SPRY domain provides a putative protein–protein interaction platform within DDX1 that differs from other SPRY domains in its structure and conserved regions. The human RNA helicase DDX1 in the DEAD-box family plays an important role in RNA processing and has been associated with HIV-1 replication and tumour progression. Whereas previously described DEAD-box proteins have a structurally conserved core, DDX1 shows a unique structural feature: a large SPRY-domain insertion in its RecA-like consensus fold. SPRY domains are known to function as protein–protein interaction platforms. Here, the crystal structure of the SPRY domain of human DDX1 (hDSPRY) is reported at 2.0 Å resolution. The structure reveals two layers of concave, antiparallel β-sheets that stack onto each other and a third β-sheet beneath the β-sandwich. A comparison with SPRY-domain structures from other eukaryotic proteins showed that the general β-sandwich fold is conserved; however, differences were detected in the loop regions, which were identified in other SPRY domains to be essential for interaction with cognate partners. In contrast, in hDSPRY these loop regions are not strictly conserved across species. Interestingly, though, a conserved patch of positive surface charge is found that may replace the connecting loops as a protein–protein interaction surface. The data presented here comprise the first structural information on DDX1 and provide insights into the unique domain architecture of this DEAD-box protein. By providing the structure of a putative interaction domain of DDX1, this work will serve as a basis for further studies of the interaction network within the hetero-oligomeric complexes of DDX1 and of its recruitment to the HIV-1 Rev protein as a viral replication factor.

  10. An open library of human kinase domain constructs for automated bacterial expression

    OpenAIRE

    Rodríguez-Laureano, Lucelenie; Işık, Mehtap; Chodera, John; Seeliger, Markus; Jeans, Chris; Gradia, Scott; Hanson, Sonya; Parton, Daniel; Albanese, Steven; Levinson, Nicholas; Behr, Julie

    2017-01-01

    Kinases play a critical role in many cellular signaling pathways and are dysregulated in a number of diseases, such as cancer, diabetes, and neurodegeneration. Since the FDA approval of imatinib in 2001, therapeutics targeting kinases now account for roughly 50% of current cancer drug discovery efforts. The ability to explore human kinase biochemistry, biophysics, and structural biology in the laboratory is essential to making rapid progress in understanding kinase regulation, designing selec...

  11. Interactive domains in the molecular chaperone human alphaB crystallin modulate microtubule assembly and disassembly.

    Directory of Open Access Journals (Sweden)

    Joy G Ghosh

    2007-06-01

    Full Text Available Small heat shock proteins regulate microtubule assembly during cell proliferation and in response to stress through interactions that are poorly understood.Novel functions for five interactive sequences in the small heat shock protein and molecular chaperone, human alphaB crystallin, were investigated in the assembly/disassembly of microtubules and aggregation of tubulin using synthetic peptides and mutants of human alphaB crystallin.The interactive sequence (113FISREFHR(120 exposed on the surface of alphaB crystallin decreased microtubule assembly by approximately 45%. In contrast, the interactive sequences, (131LTITSSLSSDGV(142 and (156ERTIPITRE(164, corresponding to the beta8 strand and the C-terminal extension respectively, which are involved in complex formation, increased microtubule assembly by approximately 34-45%. The alphaB crystallin peptides, (113FISREFHR(120 and (156ERTIPITRE(164, inhibited microtubule disassembly by approximately 26-36%, and the peptides (113FISREFHR(120 and (131LTITSSLSSDGV(142 decreased the thermal aggregation of tubulin by approximately 42-44%. The (131LTITSSLSSDGV(142 and (156ERTIPITRE(164 peptides were more effective than the widely used anti-cancer drug, Paclitaxel, in modulating tubulinmicrotubule dynamics. Mutagenesis of these interactive sequences in wt human alphaB crystallin confirmed the effects of the alphaB crystallin peptides on microtubule assembly/disassembly and tubulin aggregation. The regulation of microtubule assembly by alphaB crystallin varied over a narrow range of concentrations. The assembly of microtubules was maximal at alphaB crystallin to tubulin molar ratios between 1:4 and 2:1, while molar ratios >2:1 inhibited microtubule assembly.Interactive sequences on the surface of human alphaB crystallin collectively modulate microtubule assembly through a dynamic subunit exchange mechanism that depends on the concentration and ratio of alphaB crystallin to tubulin. These are the first

  12. Direct visualization of lipid domains in human skin stratum corneum's lipid membranes

    DEFF Research Database (Denmark)

    Plasencia, I; Norlen, Lars; Bagatolli, Luis

    2007-01-01

    scanning calorimetry, fluorescence spectroscopy, and two-photon excitation and laser scanning confocal fluorescence microscopy. Here we show that hydrated bilayers of human skin stratum corneum lipids express a giant sponge-like morphology with dimensions corresponding to the global three......-dimensional morphology of the stratum corneum extracellular space. These structures can be directly visualized using the aforementioned fluorescence microscopy techniques. At skin physiological temperatures (28 degrees C-32 degrees C), the phase state of these hydrated bilayers correspond microscopically (radial...

  13. Identification of a putative nuclear export signal motif in human NANOG homeobox domain

    International Nuclear Information System (INIS)

    Park, Sung-Won; Do, Hyun-Jin; Huh, Sun-Hyung; Sung, Boreum; Uhm, Sang-Jun; Song, Hyuk; Kim, Nam-Hyung; Kim, Jae-Hwan

    2012-01-01

    Highlights: ► We found the putative nuclear export signal motif within human NANOG homeodomain. ► Leucine-rich residues are important for human NANOG homeodomain nuclear export. ► CRM1-specific inhibitor LMB blocked the potent human NANOG NES-mediated nuclear export. -- Abstract: NANOG is a homeobox-containing transcription factor that plays an important role in pluripotent stem cells and tumorigenic cells. To understand how nuclear localization of human NANOG is regulated, the NANOG sequence was examined and a leucine-rich nuclear export signal (NES) motif ( 125 MQELSNILNL 134 ) was found in the homeodomain (HD). To functionally validate the putative NES motif, deletion and site-directed mutants were fused to an EGFP expression vector and transfected into COS-7 cells, and the localization of the proteins was examined. While hNANOG HD exclusively localized to the nucleus, a mutant with both NLSs deleted and only the putative NES motif contained (hNANOG HD-ΔNLSs) was predominantly cytoplasmic, as observed by nucleo/cytoplasmic fractionation and Western blot analysis as well as confocal microscopy. Furthermore, site-directed mutagenesis of the putative NES motif in a partial hNANOG HD only containing either one of the two NLS motifs led to localization in the nucleus, suggesting that the NES motif may play a functional role in nuclear export. Furthermore, CRM1-specific nuclear export inhibitor LMB blocked the hNANOG potent NES-mediated export, suggesting that the leucine-rich motif may function in CRM1-mediated nuclear export of hNANOG. Collectively, a NES motif is present in the hNANOG HD and may be functionally involved in CRM1-mediated nuclear export pathway.

  14. SU-G-IeP4-10: Microimaging for Different Degrees of Human Cavernous Hemangioma of Liver by Using In-Line Phase-Contrast Imaging CT

    International Nuclear Information System (INIS)

    Duan, J

    2016-01-01

    Purpose: Cavernous hemangioma of the liver (CHL) is the most common benign solid tumor of the liver. In this study, we quantitative assessment the different degrees of CHL from microscopic viewpoint by using in-line phase-contrast imaging CT (ILPCI-CT). Methods: The experiments were performed at x-ray imaging and biomedical application beamline (BL13W1) of Shanghai Synchrotron Radiation Facility (SSRF) in China. Three typical specimens at different stages, i.e., mild, moderate and severe human CHL were imaged using ILPCI-CT at 16keV without contrast agents. The 3D visualization of different degrees of CHL samples were presented using ILPCI-CT. Additionally, quantitative evaluation of the CHL features, such as the range of hepatic sinusoid equivalent diameters in different degrees of CHL samples, the ratio of the hepatic sinusoid to the CHL tissue, were measured. Results: The planar image clearly displayed the dilated hepatic sinusoids in microns. There was no normal hepatic vascular found in the all CHL samples. Different stages of CHL samples were presented with vivid shapes and stereoscopic effects by using 3D visualization. The equivalent diameters of hepatic sinusoids in three degrees CHL were different. The equivalent diameters of the hepatic sinusoids in mild CHL, range from 60 to 120 µm. The equivalent diameters of the hepatic sinusoids in moderate CHL, range from 65 to 190 µm. The equivalent diameters of the hepatic sinusoids in severe CHL, range from 95 to 215 µm. The ratio of the hepatic sinusoid to the mild, moderate and severe CHL tissue were 3%, 16% and 21%, respectively. Conclusion: The results show that the high degree of sensitivity of the ILPCI-CT technique and demonstrate the feasibility of accurate visualization of different stage human CHL. ILPCI-CT may offers a potential use in non-invasive study and analysis of CHL.

  15. NMR investigation of the isolated second voltage-sensing domain of human Nav1.4 channel.

    Science.gov (United States)

    Paramonov, A S; Lyukmanova, E N; Myshkin, M Yu; Shulepko, M A; Kulbatskii, D S; Petrosian, N S; Chugunov, A O; Dolgikh, D A; Kirpichnikov, M P; Arseniev, A S; Shenkarev, Z O

    2017-03-01

    Voltage-gated Na + channels are essential for the functioning of cardiovascular, muscular, and nervous systems. The α-subunit of eukaryotic Na + channel consists of ~2000 amino acid residues and encloses 24 transmembrane (TM) helices, which form five membrane domains: four voltage-sensing (VSD) and one pore domain. The structural complexity significantly impedes recombinant production and structural studies of full-sized Na + channels. Modular organization of voltage-gated channels gives an idea for studying of the isolated second VSD of human skeletal muscle Nav1.4 channel (VSD-II). Several variants of VSD-II (~150a.a., four TM helices) with different N- and C-termini were produced by cell-free expression. Screening of membrane mimetics revealed low stability of VSD-II samples in media containing phospholipids (bicelles, nanodiscs) associated with the aggregation of electrically neutral domain molecules. The almost complete resonance assignment of 13 C, 15 N-labeled VSD-II was obtained in LPPG micelles. The secondary structure of VSD-II showed similarity with the structures of bacterial Na + channels. The fragment of S4 TM helix between the first and second conserved Arg residues probably adopts 3 10 -helical conformation. Water accessibility of S3 helix, observed by the Mn 2+ titration, pointed to the formation of water-filled crevices in the micelle embedded VSD-II. 15 N relaxation data revealed characteristic pattern of μs-ms time scale motions in the VSD-II regions sharing expected interhelical contacts. VSD-II demonstrated enhanced mobility at ps-ns time scale as compared to isolated VSDs of K + channels. These results validate structural studies of isolated VSDs of Na + channels and show possible pitfalls in application of this 'divide and conquer' approach. Copyright © 2017 Elsevier B.V. All rights reserved.

  16. Constraints imposed by transmembrane domains affect enzymatic activity of membrane-associated human CD39/NTPDase1 mutants.

    Science.gov (United States)

    Musi, Elgilda; Islam, Naziba; Drosopoulos, Joan H F

    2007-05-01

    Human CD39/NTPDase1 is an endothelial cell membrane-associated nucleotidase. Its large extracellular domain rapidly metabolizes nucleotides, especially ADP released from activated platelets, inhibiting further platelet activation/recruitment. Previous studies using our recombinant soluble CD39 demonstrated the importance of residues S57, D54, and D213 for enzymatic/biological activity. We now report effects of S57A, D54A, and D213A mutations on full-length (FL)CD39 function. Enzymatic activity of alanine modified FLCD39s was less than wild-type, contrasting the enhanced activity of their soluble counterparts. Furthermore, conservative substitutions D54E and D213E led to enzymes with activities greater than the alanine modified FLCD39s, but less than wild-type. Reductions in mutant activities were primarily associated with reduced catalytic rates. Differences in enzymatic activity were not attributable to gross changes in the nucleotide binding pocket or the enzyme's ability to multimerize. Thus, composition of the active site of wild-type CD39 appears optimized for ADPase function in the context of the transmembrane domains.

  17. Entamoeba lysyl-tRNA synthetase contains a cytokine-like domain with chemokine activity towards human endothelial cells.

    Directory of Open Access Journals (Sweden)

    Manuel Castro de Moura

    2011-11-01

    Full Text Available Immunological pressure encountered by protozoan parasites drives the selection of strategies to modulate or avoid the immune responses of their hosts. Here we show that the parasite Entamoeba histolytica has evolved a chemokine that mimics the sequence, structure, and function of the human cytokine HsEMAPII (Homo sapiens endothelial monocyte activating polypeptide II. This Entamoeba EMAPII-like polypeptide (EELP is translated as a domain attached to two different aminoacyl-tRNA synthetases (aaRS that are overexpressed when parasites are exposed to inflammatory signals. EELP is dispensable for the tRNA aminoacylation activity of the enzymes that harbor it, and it is cleaved from them by Entamoeba proteases to generate a standalone cytokine. Isolated EELP acts as a chemoattractant for human cells, but its cell specificity is different from that of HsEMAPII. We show that cell specificity differences between HsEMAPII and EELP can be swapped by site directed mutagenesis of only two residues in the cytokines' signal sequence. Thus, Entamoeba has evolved a functional mimic of an aaRS-associated human cytokine with modified cell specificity.

  18. Concerted action of the PHD, chromo and motor domains regulates the human chromatin remodelling ATPase CHD4

    OpenAIRE

    Morra, Rosa; Lee, Benjamin M; Shaw, Heather; Tuma, Roman; Mancini, Erika J

    2012-01-01

    CHD4, the core subunit of the Nucleosome Remodelling and Deacetylase (NuRD) complex, is a chromatin remodelling ATPase that, in addition to a helicase domain, harbors tandem plant homeo finger and chromo domains. By using a panel of domain constructs we dissect their roles and demonstrate that DNA binding, histone binding and ATPase activities are allosterically regulated. Molecular shape reconstruction from small-angle X-ray scattering reveals extensive domain-domain interactions, which prov...

  19. PDZ domain-binding motif of Tax sustains T-cell proliferation in HTLV-1-infected humanized mice.

    Science.gov (United States)

    Pérès, Eléonore; Blin, Juliana; Ricci, Emiliano P; Artesi, Maria; Hahaut, Vincent; Van den Broeke, Anne; Corbin, Antoine; Gazzolo, Louis; Ratner, Lee; Jalinot, Pierre; Duc Dodon, Madeleine

    2018-03-01

    Human T-cell leukemia virus type 1 (HTLV-1) is the etiological agent of adult T-cell leukemia/lymphoma (ATLL), an aggressive malignant proliferation of activated CD4+ T lymphocytes. The viral Tax oncoprotein is critically involved in both HTLV-1-replication and T-cell proliferation, a prerequisite to the development of ATLL. In this study, we investigated the in vivo contribution of the Tax PDZ domain-binding motif (PBM) to the lymphoproliferative process. To that aim, we examined T-cell proliferation in humanized mice (hu-mice) carrying a human hemato-lymphoid system infected with either a wild type (WT) or a Tax PBM-deleted (ΔPBM) provirus. We observed that the frequency of CD4+ activated T-cells in the peripheral blood and in the spleen was significantly higher in WT than in ΔPBM hu-mice. Likewise, human T-cells collected from WT hu-mice and cultivated in vitro in presence of interleukin-2 were proliferating at a higher level than those from ΔPBM animals. We next examined the association of Tax with the Scribble PDZ protein, a prominent regulator of T-cell polarity, in human T-cells analyzed either after ex vivo isolation or after in vitro culture. We confirmed the interaction of Tax with Scribble only in T-cells from the WT hu-mice. This association correlated with the presence of both proteins in aggregates at the leading edge of the cells and with the formation of long actin filopods. Finally, data from a comparative genome-wide transcriptomic analysis suggested that the PBM-PDZ association is implicated in the expression of genes regulating proliferation, apoptosis and cytoskeletal organization. Collectively, our findings suggest that the Tax PBM is an auxiliary motif that contributes to the sustained growth of HTLV-1 infected T-cells in vivo and in vitro and is essential to T-cell immortalization.

  20. Common-path Fourier domain optical coherence tomography of irradiated human skin and ventilated isolated rabbit lungs

    Science.gov (United States)

    Popp, A.; Wendel, M.; Knels, L.; Knuschke, P.; Mehner, M.; Koch, T.; Boller, D.; Koch, P.; Koch, E.

    2005-08-01

    A compact common path Fourier domain optical coherence tomography (FD-OCT) system based on a broadband superluminescence diode is used for biomedical imaging. The epidermal thickening of human skin after exposure to ultraviolet radiation is measured to proof the feasibility of FD-OCT for future substitution of invasive biopsies in a long term study on natural UV skin protection. The FD-OCT system is also used for imaging lung parenchyma. FD-OCT images of a formalin fixated lung show the same alveolar structure as scanning electron microscopy images. In the ventilated and blood-free perfused isolated rabbit lung FD-OCT is used for real-time cross-sectional image capture of alveolar mechanics throughout tidal ventilation. The alveolar mechanics changing from alternating recruitment-derecruitment at zero positive end-expiratory pressure (PEEP) to persistent recruitment after applying a PEEP of 5 cm H2O is observed in the OCT images.

  1. Suramin blocks interaction between human FGF1 and FGFR2 D2 domain and reduces downstream signaling activity

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Zong-Sian, E-mail: gary810426@hotmail.com [Department of Chemistry, National Tsing Hua University, No. 101, Section 2, Kuang-Fu Road, Hsinchu 30013, Taiwan (China); Liu, Che Fu, E-mail: s9823002@m98.nthu.edu.tw [Department of Chemistry, National Tsing Hua University, No. 101, Section 2, Kuang-Fu Road, Hsinchu 30013, Taiwan (China); Fu, Brian, E-mail: brianfu9@gmail.com [Northwood High School, Irvine, CA (United States); Chou, Ruey-Hwang, E-mail: rhchou@mail.cmu.edu.tw [Graduate Institute of Cancer Biology and Center for Molecular Medicine, China Medical University, No.91, Hsueh-Shih Road, Taichung 40402, Taiwan (China); Department of Biotechnology, Asia University, Taiwan (China); Yu, Chin, E-mail: cyu.nthu@gmail.com [Department of Chemistry, National Tsing Hua University, No. 101, Section 2, Kuang-Fu Road, Hsinchu 30013, Taiwan (China)

    2016-09-02

    The extracellular portion of the human fibroblast growth factor receptor2 D2 domain (FGFR2 D2) interacts with human fibroblast growth factor 1 (hFGF1) to activate a downstream signaling cascade that ultimately affects mitosis and differentiation. Suramin is an antiparasiticdrug and a potent inhibitor of FGF-induced angiogenesis. Suramin has been shown to bind to hFGF1, and might block the interaction between hFGF1 and FGFR2 D2. Here, we titrated hFGF1 with FGFR2 D2 and suramin to elucidate their interactions using the detection of NMR. The docking results of both hFGF1-FGFR2 D2 domain and hFGF1-suramin complex were superimposed. The results indicate that suramin blocks the interaction between hFGF1 and FGFR2 D2. We used the PyMOL software to show the hydrophobic interaction of hFGF1-suramin. In addition, we used a Water-soluble Tetrazolium salts assay (WST1) to assess hFGF1 bioactivity. The results will be useful for the development of new antimitogenic activity drugs. - Highlights: • The interfacial residues on hFGF1-FGFR2 D2 and hFGF1-Suramin contact surface were mapped by {sup 1}H-{sup 15}N HSQC experiments. • hFGF1-FGFR2 D2 and hFGF1-Suramin complex models were generated from NMR restraints by using HADDOCK program. • We analyzed hFGF1-Suramin complex models and found the interaction between hFGF1-Suramin is hydrophobic. • The bioactivity of the hFGF1-FGFR2 D2 and hFGF1-Suramin complex was studied by using WST1 assay.

  2. 1H NMR structural characterization of a recombinant kringle 2 domain from human tissue-type plasminogen activator

    International Nuclear Information System (INIS)

    Byeon, I.J.L.; Llinas, M.; Kelley, R.F.

    1989-01-01

    The kringle 2 domain of human tissue-type plasminogen activator (t-PA) has been characterized via 1 H NMR spectroscopy at 300 and 620 MHz. The experiments were performed on the isolated domain obtained by expression of the 174-263 portion of t-PA in Escherichia coli. The spectrum of t-Pa kringle 2 is characteristic of a globular structure and shows overall similarity to that of the plasminogen (PGN) kringle 4. Spectral comparison with human and bovine PGN kringle 4 identified side-chain resonances from Leu 46 , which afford a fingerprint of kringle folding, and from most of the aromatic ring spin systems. Ligand-binding studies confirm that t-PA kringle 2 binds L-lysine with an association constant K a ∼ 11.9 mM -1 . The data indicate that homologous or conserved residues relative to those that compose the lysine-binding sites of PGN kringles 1 and 4 are involved in the binding of L-lysine to t-PA kringle 2. These include Tyr 36 and, within the kringle inner loop, Trp 62 , His 64 , Trp 72 , and Tyr 74 . Several labile NH protons of t-PA kringle 2 exhibit retarded H-exchange kinetics, requiring more than a week in 2 H 2 O for full deuteration in the presence of L-lysine at 37 degree C. This reveals that kringle 2 is endowed with a compact, dynamically stable conformation. Proton Overhauser experiments in 1 H 2 O, centered on well-resolved NH resonances between 9.8 and 12 ppm, identify signals arising from the His 48a imidazole NH3 proton and the three Trp indole NH1 protons. Overall, the data indicate a highly structured conformation for the recombinant t-PA kringle 2 that is closely related to that of the previously investigated PGN kringles 1, 4, and 5

  3. Hydroxychloroquine binding to cytoplasmic domain of Band 3 in human erythrocytes: Novel mechanistic insights into drug structure, efficacy and toxicity.

    Science.gov (United States)

    Nakagawa, Mizuki; Sugawara, Kotomi; Goto, Tatsufumi; Wakui, Hideki; Nunomura, Wataru

    2016-05-13

    Hydroxychloroquine (HCQ) is a widely used drug in the treatment of autoimmune diseases, such as arthritis and systemic lupus erythematosus. It has also been prescribed for the treatment of malaria owing to its lower toxicity compared to its closely related compound chloroquine (CQ). However, the mechanisms of action of HCQ in erythrocytes (which bind preferentially this drug) have not been documented and the reasons underlying the lower side effects of HCQ compared to CQ remain unclear. Here we show that, although the activity of erythrocyte lactate dehydrogenase (LDH), but not GAPDH, was inhibited by both HCQ and CQ in vitro, LDH activity in erythrocytes incubated with 20 mM HCQ was not significantly reduced within 5 h in contrast to CQ did. Using HCQ coupled Sepharose chromatography (HCQ-Sepharose), we identified Band 3, spectrin, ankyrin, protein 4.1R and protein 4.2 as HCQ binding proteins in human erythrocyte plasma membrane. Recombinant cytoplasmic N-terminal 43 kDa domain of Band 3 bound to HCQ-Sepharose and was eluted with 40 mM (but not 20 mM) HCQ. Band 3 transport activity was reduced by only 23% in the presence of 20 mM HCQ. Taken together, these data demonstrate that HCQ binds to the cytoplasmic N-terminal domain of Band 3 in human erythrocytes but does not inhibit dramatically its transport activity. We hypothesize that the trapping of HCQ on Band 3 contributes to the lower side effects of the drug on energy production in erythrocytes. Copyright © 2016 Elsevier Inc. All rights reserved.

  4. Suramin blocks interaction between human FGF1 and FGFR2 D2 domain and reduces downstream signaling activity

    International Nuclear Information System (INIS)

    Wu, Zong-Sian; Liu, Che Fu; Fu, Brian; Chou, Ruey-Hwang; Yu, Chin

    2016-01-01

    The extracellular portion of the human fibroblast growth factor receptor2 D2 domain (FGFR2 D2) interacts with human fibroblast growth factor 1 (hFGF1) to activate a downstream signaling cascade that ultimately affects mitosis and differentiation. Suramin is an antiparasiticdrug and a potent inhibitor of FGF-induced angiogenesis. Suramin has been shown to bind to hFGF1, and might block the interaction between hFGF1 and FGFR2 D2. Here, we titrated hFGF1 with FGFR2 D2 and suramin to elucidate their interactions using the detection of NMR. The docking results of both hFGF1-FGFR2 D2 domain and hFGF1-suramin complex were superimposed. The results indicate that suramin blocks the interaction between hFGF1 and FGFR2 D2. We used the PyMOL software to show the hydrophobic interaction of hFGF1-suramin. In addition, we used a Water-soluble Tetrazolium salts assay (WST1) to assess hFGF1 bioactivity. The results will be useful for the development of new antimitogenic activity drugs. - Highlights: • The interfacial residues on hFGF1-FGFR2 D2 and hFGF1-Suramin contact surface were mapped by "1H-"1"5N HSQC experiments. • hFGF1-FGFR2 D2 and hFGF1-Suramin complex models were generated from NMR restraints by using HADDOCK program. • We analyzed hFGF1-Suramin complex models and found the interaction between hFGF1-Suramin is hydrophobic. • The bioactivity of the hFGF1-FGFR2 D2 and hFGF1-Suramin complex was studied by using WST1 assay.

  5. Intrasurgical Human Retinal Imaging With Manual Instrument Tracking Using a Microscope-Integrated Spectral-Domain Optical Coherence Tomography Device.

    Science.gov (United States)

    Hahn, Paul; Carrasco-Zevallos, Oscar; Cunefare, David; Migacz, Justin; Farsiu, Sina; Izatt, Joseph A; Toth, Cynthia A

    2015-07-01

    To characterize the first in-human intraoperative imaging using a custom prototype spectral-domain microscope-integrated optical coherence tomography (MIOCT) device during vitreoretinal surgery with instruments in the eye. Under institutional review board approval for a prospective intraoperative study, MIOCT images were obtained at surgical pauses with instruments held static in the vitreous cavity and then concurrently with surgical maneuvers. Postoperatively, MIOCT images obtained at surgical pauses were compared with images obtained with a high-resolution handheld spectral-domain OCT (HHOCT) system with objective endpoints, including acquisition of images acceptable for analysis and identification of predefined macular morphologic or pathologic features. Human MIOCT images were successfully obtained before incision and during pauses in surgical maneuvers. MIOCT imaging confirmed preoperative diagnoses, such as epiretinal membrane, full-thickness macular hole, and vitreomacular traction and demonstrated successful achievement of surgical goals. MIOCT and HHOCT images obtained at surgical pauses in two cohorts of five patients were comparable with greater than or equal to 80% correlation in 80% of patients. Real-time video-imaging concurrent with surgical manipulations enabled, for the first time using this device, visualization of dynamic instrument-retina interaction with targeted OCT tracking. MIOCT is successful for imaging at surgical pauses and for real-time image guidance with implementation of targeted OCT tracking. Even faster acquisition speeds are currently being developed with incorporation of a swept-source MIOCT engine. Further refinements and investigations will be directed toward continued integration for real-time volumetric imaging of surgical maneuvers. Ongoing development of seamless MIOCT systems will likely transform surgical visualization, approaches, and decision-making.

  6. Mutational definition of functional domains within the Rev homolog encoded by human endogenous retrovirus K.

    Science.gov (United States)

    Bogerd, H P; Wiegand, H L; Yang, J; Cullen, B R

    2000-10-01

    Nuclear export of the incompletely spliced mRNAs encoded by several complex retroviruses, including human immunodeficiency virus type 1 (HIV-1), is dependent on a virally encoded adapter protein, termed Rev in HIV-1, that directly binds both to a cis-acting viral RNA target site and to the cellular Crm1 export factor. Human endogenous retrovirus K, a family of ancient endogenous retroviruses that is not related to the exogenous retrovirus HIV-1, was recently shown to also encode a Crm1-dependent nuclear RNA export factor, termed K-Rev. Although HIV-1 Rev and K-Rev display little sequence identity, they share the ability not only to bind to Crm1 and to RNA but also to form homomultimers and shuttle between nucleus and cytoplasm. We have used mutational analysis to identify sequences in the 105-amino-acid K-Rev protein required for each of these distinct biological activities. While mutations in K-Rev that inactivate any one of these properties also blocked K-Rev-dependent nuclear RNA export, several K-Rev mutants were comparable to wild type when assayed for any of these individual activities yet nevertheless defective for RNA export. Although several nonfunctional K-Rev mutants acted as dominant negative inhibitors of K-Rev-, but not HIV-1 Rev-, dependent RNA export, these were not defined by their inability to bind to Crm1, as is seen with HIV-1 Rev. In total, this analysis suggests a functional architecture for K-Rev that is similar to, but distinct from, that described for HIV-1 Rev and raises the possibility that viral RNA export mediated by the approximately 25 million-year-old K-Rev protein may require an additional cellular cofactor that is not required for HIV-1 Rev function.

  7. EyeFrame: Real-time memory aid improves human multitasking via domain-general eye tracking procedures

    Directory of Open Access Journals (Sweden)

    P. eTaylor

    2015-09-01

    Full Text Available OBJECTIVE: We developed an extensively general closed-loop system to improve human interaction in various multitasking scenarios, with semi-autonomous agents, processes, and robots. BACKGROUND: Much technology is converging toward semi-independent processes with intermittent human supervision distributed over multiple computerized agents. Human operators multitask notoriously poorly, in part due to cognitive load and limited working memory. To multitask optimally, users must remember task order, e.g., the most neglected task, since longer times not monitoring an element indicates greater probability of need for user input. The secondary task of monitoring attention history over multiple spatial tasks requires similar cognitive resources as primary tasks themselves. Humans can not reliably make more than ~2 decisions/s. METHODS: Participants managed a range of 4-10 semi-autonomous agents performing rescue tasks. To optimize monitoring and controlling multiple agents, we created an automated short term memory aid, providing visual cues from users' gaze history. Cues indicated when and where to look next, and were derived from an inverse of eye fixation recency. RESULTS: Contingent eye tracking algorithms drastically improved operator performance, increasing multitasking capacity. The gaze aid reduced biases, and reduced cognitive load, measured by smaller pupil dilation. CONCLUSIONS: Our eye aid likely helped by delegating short-term memory to the computer, and by reducing decision making load. Past studies used eye position for gaze-aware control and interactive updating of displays in application-specific scenarios, but ours is the first to successfully implement domain-general algorithms. Procedures should generalize well to: process control, factory operations, robot control, surveillance, aviation, air traffic control, driving, military, mobile search and rescue, and many tasks where probability of utility is predicted by duration since last

  8. 3D osteocyte lacunar morphometric properties and distributions in human femoral cortical bone using synchrotron radiation micro-CT images.

    Science.gov (United States)

    Dong, Pei; Haupert, Sylvain; Hesse, Bernhard; Langer, Max; Gouttenoire, Pierre-Jean; Bousson, Valérie; Peyrin, Françoise

    2014-03-01

    Osteocytes, the most numerous bone cells, are thought to be actively involved in the bone modeling and remodeling processes. The morphology of osteocyte is hypothesized to adapt according to the physiological mechanical loading. Three-dimensional micro-CT has recently been used to study osteocyte lacunae. In this work, we proposed a computationally efficient and validated automated image analysis method to quantify the 3D shape descriptors of osteocyte lacunae and their distribution in human femurs. Thirteen samples were imaged using Synchrotron Radiation (SR) micro-CT at ID19 of the ESRF with 1.4μm isotropic voxel resolution. With a field of view of about 2.9×2.9×1.4mm(3), the 3D images include several tens of thousands of osteocyte lacunae. We designed an automated quantification method to segment and extract 3D cell descriptors from osteocyte lacunae. An image moment-based approach was used to calculate the volume, length, width, height and anisotropy of each osteocyte lacuna. We employed a fast algorithm to further efficiently calculate the surface area, the Euler number and the structure model index (SMI) of each lacuna. We also introduced the 3D lacunar density map to directly visualize the lacunar density variation over a large field of view. We reported the lacunar morphometric properties and distributions as well as cortical bone histomorphometric indices on the 13 bone samples. The mean volume and surface were found to be 409.5±149.7μm(3) and 336.2±94.5μm(2). The average dimensions were of 18.9±4.9μm in length, 9.2±2.1μm in width and 4.8±1.1μm in depth. We found lacunar number density and six osteocyte lacunar descriptors, three axis lengths, two anisotropy ratios and SMI, that are significantly correlated to bone porosity at a same local region. The proposed method allowed an automatic and efficient direct 3D analysis of a large population of bone cells and is expected to provide reliable biological information for better understanding the

  9. {sup 18}F-Alfatide II PET/CT in healthy human volunteers and patients with brain metastases

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Chunjing; Mi, Baoming; Wan, Weixing [Affiliated Hospital of Jiangnan University (Wuxi No. 4 People' s Hospital), Department of Nuclear Medicine, Wuxi (China); Pan, Donghui; Xu, Yuping; Yang, Min [Jiangsu Institute of Nuclear Medicine, Key Laboratory of Nuclear Medicine, Ministry of Health, Jiangsu Key Laboratory of Molecular Nuclear Medicine, Wuxi (China); Lang, Lixin; Niu, Gang; Chen, Xiaoyuan [National Institutes of Health, Laboratory of Molecular Imaging and Nanomedicine, National Institute of Biomedical Imaging and Bioengineering, Bethesda, MD (United States)

    2015-12-15

    We report the biodistribution and radiation dosimetry of an integrin α{sub v}β{sub 3} specific PET tracer {sup 18}F-AlF-NOTA-E[PEG{sub 4}-c(RGDfk)]{sub 2} (denoted as {sup 18}F-Alfatide II). We also assessed the value of {sup 18}F-Alfatide II in patients with brain metastases. A series of torso (from the skull to the thigh) static images were acquired in five healthy volunteers (3 M, 2 F) at 5, 10, 15, 30, 45, and 60 min after injection of {sup 18}F-Alfatide II (257 ± 48 MBq). Regions of interest (ROIs) were drawn manually, and the time-activity curves (TACs) were obtained for major organs. Nine patients with brain metastases were examined by static PET imaging with {sup 18}F-FDG (5.55 MBq/kg) and {sup 18}F-Alfatide II. Injection of {sup 18}F-Alfatide II was well tolerated in all healthy volunteers, with no serious tracer-related adverse events found. {sup 18}F-Alfatide II showed rapid clearance from the blood pool and kidneys. The total effective dose equivalent (EDE) and effective dose (ED) were 0.0277 ± 0.003 mSv/MBq and 0.0198 ± 0.002 mSv/MBq, respectively. The organs with the highest absorbed dose were the kidneys and the spleen. Nine patients with 20 brain metastatic lesions identified by MRI and/or CT were enrolled in this study. All 20 brain lesions were visualized by {sup 18}F-Alfatide II PET, while only ten lesions were visualized by {sup 18}F-FDG, and 13 by CT. F-Alfatide II is a safe PET tracer with a favorable dosimetry profile. The observed ED suggests that {sup 18}F-Alfatide II is feasible for human studies. {sup 18}F-Alfatide II has potential value in finding brain metastases of different cancers as a biomarker of angiogenesis. (orig.)

  10. Synthetic alleles at position 121 define a functional domain of human interleukin-1 beta.

    Science.gov (United States)

    Ambrosetti, D C; Palla, E; Mirtella, A; Galeotti, C; Solito, E; Navarra, P; Parente, L; Melli, M

    1996-06-01

    The non-conservative substitution of the tyrosine residue at position 121 of human interleukin-1 beta (IL-1 beta) generates protein mutants showing strong reduction of the capacity to induce (a) prostaglandin E2 (PGE2) release from fibroblasts and smooth muscle cells, (b) murine T-cells proliferation and (c) activation of interleukin-6 (IL-6) gene expression. It is generally accepted that these functions are mediated by the type-I interleukin-1 receptor (IL-1RI). However, the mutant proteins maintain the binding affinity to the types-I and II IL-1 receptors, which is the same as the control IL-1 beta, suggesting that this amino acid substitution does not alter the structure of the molecule, except locally. Thus we have identified a new functional site of IL-1 beta different from the known receptor binding region, responsible for fundamental IL-1 beta functions. Moreover, we show that the same mutants maintain at least two hypothalamic functions, that is, the in vitro short-term PGE2 release from rat hypothalamus and the induction of fever in rabbits. This result suggests that there is yet another site of the molecule responsible for the hypothalamic functions, implying that multiple active sites on the IL-1 beta molecule, possibly binding to more than one receptor chain, trigger different signals.

  11. N- versus C-domain selectivity of catalytic inactivation of human angiotensin converting enzyme by lisinopril-coupled transition metal chelates.

    Science.gov (United States)

    Hocharoen, Lalintip; Joyner, Jeff C; Cowan, J A

    2013-12-27

    The N- and C-terminal domains of human somatic angiotensin I converting enzyme (sACE-1) demonstrate distinct physiological functions, with resulting interest in the development of domain-selective inhibitors for specific therapeutic applications. Herein, the activity of lisinopril-coupled transition metal chelates was tested for both reversible binding and irreversible catalytic inactivation of each domain of sACE-1. C/N domain binding selectivity ratios ranged from 1 to 350, while rates of irreversible catalytic inactivation of the N- and C-domains were found to be significantly greater for the N-domain, suggesting a more optimal orientation of M-chelate-lisinopril complexes within the active site of the N-domain of sACE-1. Finally, the combined effect of binding selectivity and inactivation selectivity was assessed for each catalyst (double-filter selectivity factors), and several catalysts were found to cause domain-selective catalytic inactivation. The results of this study demonstrate the ability to optimize the target selectivity of catalytic metallopeptides through both binding and catalytic factors (double-filter effect).

  12. Full-length model of the human galectin-4 and insights into dynamics of inter-domain communication

    Science.gov (United States)

    Rustiguel, Joane K.; Soares, Ricardo O. S.; Meisburger, Steve P.; Davis, Katherine M.; Malzbender, Kristina L.; Ando, Nozomi; Dias-Baruffi, Marcelo; Nonato, Maria Cristina

    2016-09-01

    Galectins are proteins involved in diverse cellular contexts due to their capacity to decipher and respond to the information encoded by β-galactoside sugars. In particular, human galectin-4, normally expressed in the healthy gastrointestinal tract, displays differential expression in cancerous tissues and is considered a potential drug target for liver and lung cancer. Galectin-4 is a tandem-repeat galectin characterized by two carbohydrate recognition domains connected by a linker-peptide. Despite their relevance to cell function and pathogenesis, structural characterization of full-length tandem-repeat galectins has remained elusive. Here, we investigate galectin-4 using X-ray crystallography, small- and wide-angle X-ray scattering, molecular modelling, molecular dynamics simulations, and differential scanning fluorimetry assays and describe for the first time a structural model for human galectin-4. Our results provide insight into the structural role of the linker-peptide and shed light on the dynamic characteristics of the mechanism of carbohydrate recognition among tandem-repeat galectins.

  13. The effect of dimerizing domains and basic residues on in vitro and in vivo assembly of Mason-Pfizer monkey virus and Human immunodeficiency virus

    Czech Academy of Sciences Publication Activity Database

    Böhmová, Karolína; Hadravová, Romana; Štokrová, Jitka; Tůma, R.; Ruml, T.; Pichová, Iva; Rumlová, Michaela

    2010-01-01

    Roč. 84, č. 4 (2010), s. 1977-1988 ISSN 0022-538X R&D Projects: GA MŠk 1M0508; GA ČR GA204/09/1388 Grant - others:EUROCORES(XE) ERAS-CT-2003-980409 Institutional research plan: CEZ:AV0Z40550506 Keywords : Mason-Pfizer Monkey Virus * Human Immunodeficiency Virus * assembly * NC Subject RIV: CC - Organic Chemistry Impact factor: 5.189, year: 2010

  14. Correlation between model observers in uniform background and human observers in patient liver background for a low-contrast detection task in CT

    Science.gov (United States)

    Gong, Hao; Yu, Lifeng; Leng, Shuai; Dilger, Samantha; Zhou, Wei; Ren, Liqiang; McCollough, Cynthia H.

    2018-03-01

    Channelized Hotelling observer (CHO) has demonstrated strong correlation with human observer (HO) in both single-slice viewing mode and multi-slice viewing mode in low-contrast detection tasks with uniform background. However, it remains unknown if the simplest single-slice CHO in uniform background can be used to predict human observer performance in more realistic tasks that involve patient anatomical background and multi-slice viewing mode. In this study, we aim to investigate the correlation between CHO in a uniform water background and human observer performance at a multi-slice viewing mode on patient liver background for a low-contrast lesion detection task. The human observer study was performed on CT images from 7 abdominal CT exams. A noise insertion tool was employed to synthesize CT scans at two additional dose levels. A validated lesion insertion tool was used to numerically insert metastatic liver lesions of various sizes and contrasts into both phantom and patient images. We selected 12 conditions out of 72 possible experimental conditions to evaluate the correlation at various radiation doses, lesion sizes, lesion contrasts and reconstruction algorithms. CHO with both single and multi-slice viewing modes were strongly correlated with HO. The corresponding Pearson's correlation coefficient was 0.982 (with 95% confidence interval (CI) [0.936, 0.995]) and 0.989 (with 95% CI of [0.960, 0.997]) in multi-slice and single-slice viewing modes, respectively. Therefore, this study demonstrated the potential to use the simplest single-slice CHO to assess image quality for more realistic clinically relevant CT detection tasks.

  15. Human ClC-6 is a late endosomal glycoprotein that associates with detergent-resistant lipid domains.

    Directory of Open Access Journals (Sweden)

    Sofie Ignoul

    and ClC-7 when cotransfected in COS-1 cells. CONCLUSIONS: We conclude that human ClC-6 is an endosomal glycoprotein that partitions in detergent resistant lipid domains. The differential sorting of endogenous (late endosomal versus overexpressed (early and recycling endosomal ClC-6 is reminiscent of that of other late endosomal/lysosomal membrane proteins (e.g. LIMP II, and is consistent with a rate-limiting sorting step for ClC-6 between early endosomes and its final destination in late endosomes.

  16. Spectral-domain optical coherence tomography study on dynamic changes of human tears after instillation of artificial tears.

    Science.gov (United States)

    Napoli, Pietro Emanuele; Satta, Giovanni Maria; Coronella, Franco; Fossarello, Maurizio

    2014-07-01

    To analyze in vivo the dynamic changes induced by different artificial tears (ATs) in the precorneal tear film (PCTF) and lower tear meniscus (LTM) by using spectral-domain (SD) anterior segment optical coherence tomography (OCT). We prospectively examined 42 normal human eyes by using SD-OCT imaging. On the day before OCT imaging, all enrolled subjects were evaluated for abnormalities of ocular surface. All tear film images were obtained before and after instillation of three different types of ATs (mucomimetic, lipid-based, and saline) in five serial scans: immediately (within 30 seconds), at the first, fifth, 10th, and 20th minute. Subjects received a drop of 35 μL in one randomly selected eye. All examinations were conducted in the same conditions of temperature, brightness, humidity, and time of day. Changes in the morphological pattern of both LTM and PCTF were associated with the type of artificial tear instilled on the ocular surface. Similarly, the radius of curvature (rc), the height (h), and the depth (d) showed dynamic variations depending on treatment. Although by the 20th minute, both h and d returned to baseline values in all groups, a significant difference in rc (compared with baseline) was detected for mucomimetic ATs (P = 0.04) and lipid ATs (P = 0.02). Spectral-domain OCT imaging has preliminarily proved to be a noninvasive tool to evaluate, in real time, the different changes induced by ATs instillation. An important stride in understanding the clinical response to various tear substitutes can be achieved by this objective and quantitative approach. Copyright 2014 The Association for Research in Vision and Ophthalmology, Inc.

  17. Selective targeting of tumour neovasculature by a radiohalogenated human antibody fragment specific for the ED-B domain of fibronectin

    International Nuclear Information System (INIS)

    Demartis, S.; Tarli, L.; Neri, D.; Borsi, L.; Zardi, L.

    2001-01-01

    Angiogenesis is a characteristic feature of many aggressive tumours and other disorders. Antibodies capable of binding to new blood vessels, but not to mature vessels, could be used as selective targeting agents for immunoscintigraphic and radioimmunotherapeutic applications. Here we show that scFv(L19), a recombinant human antibody fragment with sub-nanomolar affinity for the ED-B domain of fibronectin, a marker of angiogenesis, can be stably labelled with iodine-125 and astatine-211 with full retention of immunoreactivity, using a trimethyl-stannyl benzoate bifunctional derivative. Biodistribution studies in mice bearing two different types of tumour grafted subcutaneously, followed by ex vivo micro-autoradiographic analysis, revealed that scFv(L19) rapidly localises around tumour blood vessels, but not around normal vessels. Four hours after intravenous injection of the stably radioiodinated scFv(L19), tumour to blood ratios were 6:1 in mice bearing the F9 murine teratocarcinoma and 9:1 in mice bearing an FE8 rat sarcoma. As expected, all other organs (including kidney) contained significantly less radioactivity than the tumour. Since the ED-B domain of fibronectin has an identical sequence in mouse and man, scFv(L19) is a pan-species antibody and the results presented here suggest clinical utility of radiolabelled scFv(L19) for the scintigraphic detection of angiogenesis in vivo. Furthermore, it should now be possible to investigate scFv(L19) for the selective delivery of 211 At to the tumour neovasculature, causing the selective death of tumour endothelial cells and tumour collapse. (orig.)

  18. The disordered C-terminal domain of human DNA glycosylase NEIL1 contributes to its stability via intramolecular interactions.

    Science.gov (United States)

    Hegde, Muralidhar L; Tsutakawa, Susan E; Hegde, Pavana M; Holthauzen, Luis Marcelo F; Li, Jing; Oezguen, Numan; Hilser, Vincent J; Tainer, John A; Mitra, Sankar

    2013-07-10

    NEIL1 [Nei (endonuclease VIII)-like protein 1], one of the five mammalian DNA glycosylases that excise oxidized DNA base lesions in the human genome to initiate base excision repair, contains an intrinsically disordered C-terminal domain (CTD; ~100 residues), not conserved in its Escherichia coli prototype Nei. Although dispensable for NEIL1's lesion excision and AP lyase activities, this segment is required for efficient in vivo enzymatic activity and may provide an interaction interface for many of NEIL1's interactions with other base excision repair proteins. Here, we show that the CTD interacts with the folded domain in native NEIL1 containing 389 residues. The CTD is poised for local folding in an ordered structure that is induced in the purified fragment by osmolytes. Furthermore, deletion of the disordered tail lacking both Tyr and Trp residues causes a red shift in NEIL1's intrinsic Trp-specific fluorescence, indicating a more solvent-exposed environment for the Trp residues in the truncated protein, which also exhibits reduced stability compared to the native enzyme. These observations are consistent with stabilization of the native NEIL1 structure via intramolecular, mostly electrostatic, interactions that were disrupted by mutating a positively charged (Lys-rich) cluster of residues (amino acids 355-360) near the C-terminus. Small-angle X-ray scattering (SAXS) analysis confirms the flexibility and dynamic nature of NEIL1's CTD, a feature that may be critical to providing specificity for NEIL1's multiple, functional interactions. Copyright © 2013 Elsevier Ltd. All rights reserved.

  19. Activation of nucleotide oligomerization domain 2 (NOD2 by human cytomegalovirus initiates innate immune responses and restricts virus replication.

    Directory of Open Access Journals (Sweden)

    Arun Kapoor

    Full Text Available Nucleotide-binding oligomerization domain 2 (NOD2 is an important innate immune sensor of bacterial pathogens. Its induction results in activation of the classic NF-κB pathway and alternative pathways including type I IFN and autophagy. Although the importance of NOD2 in recognizing RNA viruses has recently been identified, its role in sensing DNA viruses has not been studied. We report that infection with human cytomegalovirus (HCMV results in significant induction of NOD2 expression, beginning as early as 2 hours post infection and increasing steadily 24 hours post infection and afterwards. Infection with human herpesvirus 1 and 2 does not induce NOD2 expression. While the HCMV-encoded glycoprotein B is not required for NOD2 induction, a replication competent virion is necessary. Lentivirus-based NOD2 knockdown in human foreskin fibroblasts (HFFs and U373 glioma cells leads to enhanced HCMV replication along with decreased levels of interferon beta (IFN-β and the pro-inflammatory cytokine, IL8. NOD2 induction in HCMV-infected cells activates downstream NF-κB and interferon pathways supported by reduced nuclear localization of NF-κB and pIRF3 in NOD2 knockdown HFFs. Stable overexpression of NOD2 in HFFs restricts HCMV replication in association with increased levels of IFN-β and IL8. Similarly, transient overexpression of NOD2 in U373 cells or its downstream kinase, RIPK2, results in decreased HCMV replication and enhanced cytokine responses. However, overexpression of a mutant NOD2, 3020insC, associated with severe Crohn's disease, results in enhanced HCMV replication and decreased levels of IFN-β in U373 cells. These results show for the first time that NOD2 plays a significant role in HCMV replication and may provide a model for studies of HCMV recognition by the host cell and HCMV colitis in Crohn's disease.

  20. Identification and characterization of human polyserase-3, a novel protein with tandem serine-protease domains in the same polypeptide chain

    Directory of Open Access Journals (Sweden)

    Garabaya Cecilia

    2006-03-01

    Full Text Available Abstract Background We have previously described the identification and characterization of polyserase-1 and polyserase-2, two human serine proteases containing three different catalytic domains within the same polypeptide chain. Polyserase-1 shows a complex organization and it is synthesized as a membrane-bound protein which can generate three independent serine protease domains as a consequence of post-translational processing events. The two first domains are enzymatically active. By contrast, polyserase-2 is an extracellular glycosylated protein whose three protease domains remain embedded in the same chain, and only the first domain possesses catalytic activity. Results Following our interest in the study of the human degradome, we have cloned a human liver cDNA encoding polyserase-3, a new protease with tandem serine protease domains in the same polypeptide chain. Comparative analysis of polyserase-3 with the two human polyserases described to date, revealed that this novel polyprotein is more closely related to polyserase-2 than to polyserase-1. Thus, polyserase-3 is a secreted protein such as polyserase-2, but lacks additional domains like the type II transmembrane motif and the low-density lipoprotein receptor module present in the membrane-anchored polyserase-1. Moreover, analysis of post-translational mechanisms operating in polyserase-3 maturation showed that its two protease domains remain as integral parts of the same polypeptide chain. This situation is similar to that observed in polyserase-2, but distinct from polyserase-1 whose protease domains are proteolytically released from the original chain to generate independent units. Immunolocalization studies indicated that polyserase-3 is secreted as a non-glycosylated protein, thus being also distinct from polyserase-2, which is a heavily glycosylated protein. Enzymatic assays indicated that recombinant polyserase-3 degrades the α-chain of fibrinogen as well as pro

  1. 3D printing from microfocus computed tomography (micro-CT) in human specimens: education and future implications.

    Science.gov (United States)

    Shelmerdine, Susan C; Simcock, Ian C; Hutchinson, John Ciaran; Aughwane, Rosalind; Melbourne, Andrew; Nikitichev, Daniil I; Ong, Ju-Ling; Borghi, Alessandro; Cole, Garrard; Kingham, Emilia; Calder, Alistair D; Capelli, Claudio; Akhtar, Aadam; Cook, Andrew C; Schievano, Silvia; David, Anna; Ourselin, Sebastian; Sebire, Neil J; Arthurs, Owen J

    2018-06-14

    Microfocus CT (micro-CT) is an imaging method that provides three-dimensional digital data sets with comparable resolution to light microscopy. Although it has traditionally been used for non-destructive testing in engineering, aerospace industries and in preclinical animal studies, new applications are rapidly becoming available in the clinical setting including post-mortem fetal imaging and pathological specimen analysis. Printing three-dimensional models from imaging data sets for educational purposes is well established in the medical literature, but typically using low resolution (0.7 mm voxel size) data acquired from CT or MR examinations. With higher resolution imaging (voxel sizes below 1 micron, printing of micro-CT imaged specimens can provide insight into craniofacial surgical applications, developmental cardiac anatomy, placental imaging, archaeological remains and high-resolution bone imaging. We conclude with other potential future usages of this emerging technique.

  2. PDZ domain-binding motif of Tax sustains T-cell proliferation in HTLV-1-infected humanized mice

    Science.gov (United States)

    Artesi, Maria; Jalinot, Pierre

    2018-01-01

    Human T-cell leukemia virus type 1 (HTLV-1) is the etiological agent of adult T-cell leukemia/lymphoma (ATLL), an aggressive malignant proliferation of activated CD4+ T lymphocytes. The viral Tax oncoprotein is critically involved in both HTLV-1-replication and T-cell proliferation, a prerequisite to the development of ATLL. In this study, we investigated the in vivo contribution of the Tax PDZ domain-binding motif (PBM) to the lymphoproliferative process. To that aim, we examined T-cell proliferation in humanized mice (hu-mice) carrying a human hemato-lymphoid system infected with either a wild type (WT) or a Tax PBM-deleted (ΔPBM) provirus. We observed that the frequency of CD4+ activated T-cells in the peripheral blood and in the spleen was significantly higher in WT than in ΔPBM hu-mice. Likewise, human T-cells collected from WT hu-mice and cultivated in vitro in presence of interleukin-2 were proliferating at a higher level than those from ΔPBM animals. We next examined the association of Tax with the Scribble PDZ protein, a prominent regulator of T-cell polarity, in human T-cells analyzed either after ex vivo isolation or after in vitro culture. We confirmed the interaction of Tax with Scribble only in T-cells from the WT hu-mice. This association correlated with the presence of both proteins in aggregates at the leading edge of the cells and with the formation of long actin filopods. Finally, data from a comparative genome-wide transcriptomic analysis suggested that the PBM-PDZ association is implicated in the expression of genes regulating proliferation, apoptosis and cytoskeletal organization. Collectively, our findings suggest that the Tax PBM is an auxiliary motif that contributes to the sustained growth of HTLV-1 infected T-cells in vivo and in vitro and is essential to T-cell immortalization. PMID:29566098

  3. Preclinical evaluation and intraoperative human retinal imaging with a high-resolution microscope-integrated spectral domain optical coherence tomography device.

    Science.gov (United States)

    Hahn, Paul; Migacz, Justin; O'Donnell, Rachelle; Day, Shelley; Lee, Annie; Lin, Phoebe; Vann, Robin; Kuo, Anthony; Fekrat, Sharon; Mruthyunjaya, Prithvi; Postel, Eric A; Izatt, Joseph A; Toth, Cynthia A

    2013-01-01

    The authors have recently developed a high-resolution microscope-integrated spectral domain optical coherence tomography (MIOCT) device designed to enable OCT acquisition simultaneous with surgical maneuvers. The purpose of this report is to describe translation of this device from preclinical testing into human intraoperative imaging. Before human imaging, surgical conditions were fully simulated for extensive preclinical MIOCT evaluation in a custom model eye system. Microscope-integrated spectral domain OCT images were then acquired in normal human volunteers and during vitreoretinal surgery in patients who consented to participate in a prospective institutional review board-approved study. Microscope-integrated spectral domain OCT images were obtained before and at pauses in surgical maneuvers and were compared based on predetermined diagnostic criteria to images obtained with a high-resolution spectral domain research handheld OCT system (HHOCT; Bioptigen, Inc) at the same time point. Cohorts of five consecutive patients were imaged. Successful end points were predefined, including ≥80% correlation in identification of pathology between MIOCT and HHOCT in ≥80% of the patients. Microscope-integrated spectral domain OCT was favorably evaluated by study surgeons and scrub nurses, all of whom responded that they would consider participating in human intraoperative imaging trials. The preclinical evaluation identified significant improvements that were made before MIOCT use during human surgery. The MIOCT transition into clinical human research was smooth. Microscope-integrated spectral domain OCT imaging in normal human volunteers demonstrated high resolution comparable to tabletop scanners. In the operating room, after an initial learning curve, surgeons successfully acquired human macular MIOCT images before and after surgical maneuvers. Microscope-integrated spectral domain OCT imaging confirmed preoperative diagnoses, such as full-thickness macular hole

  4. The role of glycosylation and domain interactions in the thermal stability of human angiotensin-converting enzyme.

    Science.gov (United States)

    O'Neill, Hester G; Redelinghuys, Pierre; Schwager, Sylva L U; Sturrock, Edward D

    2008-09-01

    The N and C domains of somatic angiotensin-converting enzyme (sACE) differ in terms of their substrate specificity, inhibitor profiling, chloride dependency and thermal stability. The C domain is thermally less stable than sACE or the N domain. Since both domains are heavily glycosylated, the effect of glycosylation on their thermal stability was investigated by assessing their catalytic and physicochemical properties. Testis ACE (tACE) expressed in mammalian cells, mammalian cells in the presence of a glucosidase inhibitor and insect cells yielded proteins with altered catalytic and physicochemical properties, indicating that the more complex glycans confer greater thermal stabilization. Furthermore, a decrease in tACE and N-domain N-glycans using site-directed mutagenesis decreased their thermal stability, suggesting that certain N-glycans have an important effect on the protein's thermodynamic properties. Evaluation of the thermal stability of sACE domain swopover and domain duplication mutants, together with sACE expressed in insect cells, showed that the C domain contained in sACE is less dependent on glycosylation for thermal stabilization than a single C domain, indicating that stabilizing interactions between the two domains contribute to the thermal stability of sACE and are decreased in a C-domain-duplicating mutant.

  5. Concerted action of the PHD, chromo and motor domains regulates the human chromatin remodelling ATPase CHD4.

    Science.gov (United States)

    Morra, Rosa; Lee, Benjamin M; Shaw, Heather; Tuma, Roman; Mancini, Erika J

    2012-07-30

    CHD4, the core subunit of the Nucleosome Remodelling and Deacetylase (NuRD) complex, is a chromatin remodelling ATPase that, in addition to a helicase domain, harbors tandem plant homeo finger and chromo domains. By using a panel of domain constructs we dissect their roles and demonstrate that DNA binding, histone binding and ATPase activities are allosterically regulated. Molecular shape reconstruction from small-angle X-ray scattering reveals extensive domain-domain interactions, which provide a structural explanation for the regulation of CHD4 activities by intramolecular domain communication. Our results demonstrate functional interdependency between domains within a chromatin remodeller. Copyright © 2012 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  6. Spiral CT and optimization of the modalities of the iodinated intravenous contrast material: Experimental studies in human pathology

    International Nuclear Information System (INIS)

    Bonaldi, V.

    1998-01-01

    Spiral (or helical) CT represents the most recent improvement in the field of computed assisted tomography (CAT scan). The capabilities of this new imaging modality are much superior to these of conventional CT scanning; then result from the rapid acquisition and from the volumetric nature of the derived data set. The short time of data acquisition had made mandatory the revision of protocols for intravenous administration of iodinated contrast material. By the means of several studies, carried out on pathologic and healthy patients, we have attempted to improve knowledge in factors influencing CT attenuation values after injection of contrast material, in the aim of improving contrast administration performed during spiral CT scanning. Anatomical landmarks that we have studied till now have been liver, the pancreas, the kidney and the cervical spine. In addition, a paired based methodology has been used. The volumetric set of data derived from spiral CT scanning leads to optimal post-processing tasks, the most interesting being related to cine-display and multiplanar reformatting; both modalities have been evaluated, about the pancreas and the musculo-skeletal system respectively. Conversely, this new modality, as for other imaging modalities, is responsible for additional costs derived from restless increase in the number of images to be dealt with and from the occurrence of new tasks (in post-processing particularly). The place of spiral CT in diagnostic strategies among other modern imaging modalities should be assessed, especially with respect to Magnetic Resonance Imaging (MRI). (author)

  7. Fingerprinting of near-homogeneous DNA ligase I and II from human cells. Similarity of their AMP-binding domains.

    Science.gov (United States)

    Yang, S W; Becker, F F; Chan, J Y

    1990-10-25

    DNA ligases play obligatory roles during replication, repair, and recombination. Multiple forms of DNA ligase have been reported in mammalian cells including DNA ligase I, the high molecular mass species which functions during replication, and DNA ligase II, the low molecular mass species which is associated with repair. In addition, alterations in DNA ligase activities have been reported in acute lymphocytic leukemia cells, Bloom's syndrome cells, and cells undergoing differentiation and development. To better distinguish the biochemical and molecular properties of the various DNA ligases from human cells, we have developed a method of purifying multiple species of DNA ligase from HeLa cells by chromatography through DEAE-Bio-Gel, CM-Bio-Gel, hydroxylapatite, Sephacryl S-300, Mono P, and DNA-cellulose. DNA-cellulose chromatography of the partially purified enzymes resolved multiple species of DNA ligase after labeling the enzyme with [alpha-32P]ATP to form the ligase-[32P]AMP adduct. The early eluting enzyme activity (0.25 M NaCl) contained a major 67-kDa-labeled protein, while the late eluting activity (0.48 M NaCl) contained two major labeled proteins of 90 and 78 kDa. Neutralization experiments with antiligase I antibodies indicated that the early and late eluting activity peaks were DNA ligase II and I, respectively. The three major ligase-[32P]AMP polypeptides (90, 78, and 67 kDa) were subsequently purified to near homogeneity by elution from preparative sodium dodecyl sulfate-polyacrylamide gels. All three polypeptides retained DNA ligase activities after gel elution and renaturation. To further reveal the relationship between these enzymes, partial digestion by V8-protease was performed. All three purified polypeptides gave rise to a common 22-kDa-labeled fragment for their AMP-binding domains, indicating that the catalytic sites of ligase I and II are quite similar, if not identical. Similar findings were obtained from the two-dimensional gel

  8. N- vs. C-Domain Selectivity of Catalytic Inactivation of Human Angiotensin Converting Enzyme by Lisinopril-Coupled Transition Metal Chelates

    Science.gov (United States)

    Hocharoen, Lalintip; Joyner, Jeff C.; Cowan, J. A.

    2014-01-01

    The N- and C-terminal domains of human somatic Angiotensin I Converting Enzyme (sACE-1) demonstrate distinct physiological functions, with resulting interest in the development of domain-selective inhibitors for specific therapeutic applications. Herein, the activity of lisinopril-coupled transition metal chelates were tested for both reversible binding and irreversible catalytic inactivation of sACE-1. C/N domain binding selectivity ratios ranged from 1 to 350, while rates of irreversible catalytic inactivation of the N- and C-domains were found to be significantly greater for the N-domain, suggesting a more optimal orientation of the M-chelate-lisinopril complexes within the active site of the N-domain of sACE-1. Finally, the combined effect of binding selectivity and inactivation selectivity was assessed for each catalyst (double-filter selectivity factors), and several catalysts were found to cause domain-selective catalytic inactivation. The results of this study demonstrate the ability to optimize the target selectivity of catalytic metallopeptides through both binding and orientation factors (double-filter effect). PMID:24228790

  9. Nucleotide-oligomerizing domain-1 (NOD1) receptor activation induces pro-inflammatory responses and autophagy in human alveolar macrophages.

    Science.gov (United States)

    Juárez, Esmeralda; Carranza, Claudia; Hernández-Sánchez, Fernando; Loyola, Elva; Escobedo, Dante; León-Contreras, Juan Carlos; Hernández-Pando, Rogelio; Torres, Martha; Sada, Eduardo

    2014-09-25

    Nucleotide-binding oligomerizing domain-1 (NOD1) is a cytoplasmic receptor involved in recognizing bacterial peptidoglycan fragments that localize to the cytosol. NOD1 activation triggers inflammation, antimicrobial mechanisms and autophagy in both epithelial cells and murine macrophages. NOD1 mediates intracellular pathogen clearance in the lungs of mice; however, little is known about NOD1's role in human alveolar macrophages (AMs) or its involvement in Mycobacterium tuberculosis (Mtb) infection. AMs, monocytes (MNs), and monocyte-derived macrophages (MDMs) from healthy subjects were assayed for NOD1 expression. Cells were stimulated with the NOD1 ligand Tri-DAP and cytokine production and autophagy were assessed. Cells were infected with Mtb and treated with Tri-DAP post-infection. CFUs counting determined growth control, and autophagy protein recruitment to pathogen localization sites was analyzed by immunoelectron microscopy. NOD1 was expressed in AMs, MDMs and to a lesser extent MNs. Tri-DAP stimulation induced NOD1 up-regulation and a significant production of IL1β, IL6, IL8, and TNFα in AMs and MDMs; however, the level of NOD1-dependent response in MNs was limited. Autophagy activity determined by expression of proteins Atg9, LC3, IRGM and p62 degradation was induced in a NOD1-dependent manner in AMs and MDMs but not in MNs. Infected AMs could be activated by stimulation with Tri-DAP to control the intracellular growth of Mtb. In addition, recruitment of NOD1 and the autophagy proteins IRGM and LC3 to the Mtb localization site was observed in infected AMs after treatment with Tri-DAP. NOD1 is involved in AM and MDM innate responses, which include proinflammatory cytokines and autophagy, with potential implications in the killing of Mtb in humans.

  10. Iodine and freeze-drying enhanced high-resolution MicroCT imaging for reconstructing 3D intraneural topography of human peripheral nerve fascicles.

    Science.gov (United States)

    Yan, Liwei; Guo, Yongze; Qi, Jian; Zhu, Qingtang; Gu, Liqiang; Zheng, Canbin; Lin, Tao; Lu, Yutong; Zeng, Zitao; Yu, Sha; Zhu, Shuang; Zhou, Xiang; Zhang, Xi; Du, Yunfei; Yao, Zhi; Lu, Yao; Liu, Xiaolin

    2017-08-01

    The precise annotation and accurate identification of the topography of fascicles to the end organs are prerequisites for studying human peripheral nerves. In this study, we present a feasible imaging method that acquires 3D high-resolution (HR) topography of peripheral nerve fascicles using an iodine and freeze-drying (IFD) micro-computed tomography (microCT) method to greatly increase the contrast of fascicle images. The enhanced microCT imaging method can facilitate the reconstruction of high-contrast HR fascicle images, fascicle segmentation and extraction, feature analysis, and the tracing of fascicle topography to end organs, which define fascicle functions. The complex intraneural aggregation and distribution of fascicles is typically assessed using histological techniques or MR imaging to acquire coarse axial three-dimensional (3D) maps. However, the disadvantages of histological techniques (static, axial manual registration, and data instability) and MR imaging (low-resolution) limit these applications in reconstructing the topography of nerve fascicles. Thus, enhanced microCT is a new technique for acquiring 3D intraneural topography of the human peripheral nerve fascicles both to improve our understanding of neurobiological principles and to guide accurate repair in the clinic. Additionally, 3D microstructure data can be used as a biofabrication model, which in turn can be used to fabricate scaffolds to repair long nerve gaps. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Effects of acute tryptophan depletion on central processing of CT-targeted and discriminatory touch in humans.

    Science.gov (United States)

    Trotter, Paula Diane; McGlone, Francis; McKie, Shane; McFarquhar, Martyn; Elliott, Rebecca; Walker, Susannah Claire; Deakin, John Francis William

    2016-08-01

    C-tactile afferents (CTs) are slowly conducting nerve fibres, present only in hairy skin. They are optimally activated by slow, gentle stroking touch, such as those experienced during a caress. CT stimulation activates affective processing brain regions, alluding to their role in affective touch perception. We tested a theory that CT-activating touch engages the pro-social functions of serotonin, by determining whether reducing serotonin, through acute tryptophan depletion, diminishes subjective pleasantness and affective brain responses to gentle touch. A tryptophan depleting amino acid drink was administered to 16 healthy females, with a further 14 receiving a control drink. After 4 h, participants underwent an fMRI scan, during which time CT-innervated forearm skin and CT non-innervated finger skin was stroked with three brushes of differing texture, at CT-optimal force and velocity. Pleasantness ratings were obtained post scanning. The control group showed a greater response in ipsilateral orbitofrontal cortex to CT-activating forearm touch compared to touch to the finger where CTs are absent. This differential response was not present in the tryptophan depleted group. This interaction effect was significant. In addition, control participants showed a differential primary somatosensory cortex response to brush texture applied to the finger, a purely discriminatory touch response, which was not observed in the tryptophan depleted group. This interaction effect was also significant. Pleasantness ratings were similar across treatment groups. These results implicate serotonin in the differentiation between CT-activating and purely discriminatory touch responses. Such effects could contribute to some of the social abnormalities seen in psychiatric disorders associated with abnormal serotonin function. © 2016 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

  12. Human pro. cap alpha. 1)(I) collagen: cDNA sequence for the C-propeptide domain

    Energy Technology Data Exchange (ETDEWEB)

    Maekelae, J K; Raassina, M; Virta, A; Vuorio, E

    1988-01-11

    The authors have previously constructed a cDNA clone pHCAL1, covering most of the C-terminal propeptide domain of human pro..cap alpha..1(I) collagen mRNA,by inserting a 678 bp EcoRI-XhoI fragment of cDNA into pBR322. Since the XhoI/SalI ligation prevented removal of the insert, they used the same strategy to obtain a similar clone in pUC8. RNA was isolated from fetal calvarial bones. The cDNA was digested with EcoRI and XhoI and fractionated on a 1 % agarose gel. Fragments of 650-700 bp were cloned in pUC8 at the polylinker site, which now permits easy removal of the insert. The new clone was named pHCAL1U since the RNA was isolated from another individual. The approach outlined is useful for studies on individual variation which is important to recognize when searching for disease-related mutations in type I collagen.

  13. High-resolution whole-genome sequencing reveals that specific chromatin domains from most human chromosomes associate with nucleoli.

    Science.gov (United States)

    van Koningsbruggen, Silvana; Gierlinski, Marek; Schofield, Pietá; Martin, David; Barton, Geoffey J; Ariyurek, Yavuz; den Dunnen, Johan T; Lamond, Angus I

    2010-11-01

    The nuclear space is mostly occupied by chromosome territories and nuclear bodies. Although this organization of chromosomes affects gene function, relatively little is known about the role of nuclear bodies in the organization of chromosomal regions. The nucleolus is the best-studied subnuclear structure and forms around the rRNA repeat gene clusters on the acrocentric chromosomes. In addition to rDNA, other chromatin sequences also surround the nucleolar surface and may even loop into the nucleolus. These additional nucleolar-associated domains (NADs) have not been well characterized. We present here a whole-genome, high-resolution analysis of chromatin endogenously associated with nucleoli. We have used a combination of three complementary approaches, namely fluorescence comparative genome hybridization, high-throughput deep DNA sequencing and photoactivation combined with time-lapse fluorescence microscopy. The data show that specific sequences from most human chromosomes, in addition to the rDNA repeat units, associate with nucleoli in a reproducible and heritable manner. NADs have in common a high density of AT-rich sequence elements, low gene density and a statistically significant enrichment in transcriptionally repressed genes. Unexpectedly, both the direct DNA sequencing and fluorescence photoactivation data show that certain chromatin loci can specifically associate with either the nucleolus, or the nuclear envelope.

  14. Expression, purification, and refolding of active recombinant human E-selectin lectin and EGF domains in Escherichia coli.

    Science.gov (United States)

    Kawano, Susumu; Iyaguchi, Daisuke; Okada, Chiaki; Sasaki, Yusuke; Toyota, Eiko

    2013-06-01

    Attempts to obtain active E-selectin from Escherichia coli (E. coli) have not yet been successful. In this study, we succeeded in expressing the recombinant lectin and epidermal growth factor domain fragments of human E-selectin (rh-ESLE) in E. coli on a large-scale. The rh-ESLE protein was expressed as an inactive form in the inclusion bodies. The inactive form of rh-ESLE was denatured and solubilized by 6 M guanidine hydrochloride and then purified by Ni(2+) affinity chromatography under denaturing conditions. Denatured rh-ESLE was then refolded by a rapid-dilution method using a large amount of refolding buffer, which contained arginine and cysteine/cystine. The refolded rh-ESLE showed binding affinity for sLe(X) (K(d) = 321 nM, B(max) = 1.9 pmol/μg protein). This result suggests that the refolded rh-ESLE recovered its native and functional structure.

  15. Peptide array-based screening of human mesenchymal stem cell-adhesive peptides derived from fibronectin type III domain

    International Nuclear Information System (INIS)

    Okochi, Mina; Nomura, Shigeyuki; Kaga, Chiaki; Honda, Hiroyuki

    2008-01-01

    Human mesenchymal stem cell-adhesive peptides were screened based on the amino acid sequence of fibronectin type III domain 8-11 (FN-III 8-11 ) using a peptide array synthesized by the Fmoc-chemistry. Using hexameric peptide library of FN-III 8-11 scan, we identified the ALNGR (Ala-Leu-Asn-Gly-Arg) peptide that induced cell adhesion as well as RGDS (Arg-Gly-Asp-Ser) peptide. After incubation for 2 h, approximately 68% of inoculated cells adhere to the ALNGR peptide disk. Adhesion inhibition assay with integrin antibodies showed that the ALNGR peptide interacts with integrin β1 but not with αvβ3, indicating that the receptors for ALNGR are different from RGDS. Additionally, the ALNGR peptide expressed cell specificities for adhesion: cell adhesion was promoted for fibroblasts but not for keratinocytes or endotherial cells. The ALNGR peptide induced cell adhesion and promoted cell proliferation without changing its property. It is therefore useful for the construction of functional biomaterials

  16. Quantitative analysis of the interaction between the envelope protein domains and the core protein of human hepatitis B virus

    International Nuclear Information System (INIS)

    Choi, Kyoung-Jae; Lim, Chun-Woo; Yoon, Moon-Young; Ahn, Byung-Yoon; Yu, Yeon Gyu

    2004-01-01

    Interaction between preformed nucleocapsids and viral envelope proteins is critical for the assembly of virus particles in infected cells. The pre-S1 and pre-S2 and cytosolic regions of the human hepatitis B virus envelope protein had been implicated in the interaction with the core protein of nucleocapsids. The binding affinities of specific subdomains of the envelope protein to the core protein were quantitatively measured by both ELISA and BIAcore assay. While a marginal binding was detected with the pre-S1 or pre-S2, the core protein showed high affinities to pre-S with apparent dissociation constants (K D app ) of 7.3 ± 0.9 and 8.2 ± 0.4 μM by ELISA and BIAcore assay, respectively. The circular dichroism analysis suggested that conformational change occurs in pre-S through interaction with core protein. These results substantiate the importance of specific envelope domains in virion assembly, and demonstrate that the interaction between viral proteins can be quantitatively measured in vitro

  17. An optical color image watermarking scheme by using compressive sensing with human visual characteristics in gyrator domain

    Science.gov (United States)

    Liansheng, Sui; Bei, Zhou; Zhanmin, Wang; Ailing, Tian

    2017-05-01

    A novel optical color image watermarking scheme considering human visual characteristics is presented in gyrator transform domain. Initially, an appropriate reference image is constructed of significant blocks chosen from the grayscale host image by evaluating visual characteristics such as visual entropy and edge entropy. Three components of the color watermark image are compressed based on compressive sensing, and the corresponding results are combined to form the grayscale watermark. Then, the frequency coefficients of the watermark image are fused into the frequency data of the gyrator-transformed reference image. The fused result is inversely transformed and partitioned, and eventually the watermarked image is obtained by mapping the resultant blocks into their original positions. The scheme can reconstruct the watermark with high perceptual quality and has the enhanced security due to high sensitivity of the secret keys. Importantly, the scheme can be implemented easily under the framework of double random phase encoding with the 4f optical system. To the best of our knowledge, it is the first report on embedding the color watermark into the grayscale host image which will be out of attacker's expectation. Simulation results are given to verify the feasibility and its superior performance in terms of noise and occlusion robustness.

  18. Ultrashort echo-time MRI versus CT for skull aberration correction in MR-guided transcranial focused ultrasound: In vitro comparison on human calvaria.

    Science.gov (United States)

    Miller, G Wilson; Eames, Matthew; Snell, John; Aubry, Jean-François

    2015-05-01

    Transcranial magnetic resonance-guided focused ultrasound (TcMRgFUS) brain treatment systems compensate for skull-induced beam aberrations by adjusting the phase and amplitude of individual ultrasound transducer elements. These corrections are currently calculated based on a preacquired computed tomography (CT) scan of the patient's head. The purpose of the work presented here is to demonstrate the feasibility of using ultrashort echo-time magnetic resonance imaging (UTE MRI) instead of CT to calculate and apply aberration corrections on a clinical TcMRgFUS system. Phantom experiments were performed in three ex-vivo human skulls filled with tissue-mimicking hydrogel. Each skull phantom was imaged with both CT and UTE MRI. The MR images were then segmented into "skull" and "not-skull" pixels using a computationally efficient, threshold-based algorithm, and the resulting 3D binary skull map was converted into a series of 2D virtual CT images. Each skull was mounted in the head transducer of a clinical TcMRgFUS system (ExAblate Neuro, Insightec, Israel), and transcranial sonications were performed using a power setting of approximately 750 acoustic watts at several different target locations within the electronic steering range of the transducer. Each target location was sonicated three times: once using aberration corrections calculated from the actual CT scan, once using corrections calculated from the MRI-derived virtual CT scan, and once without applying any aberration correction. MR thermometry was performed in conjunction with each 10-s sonication, and the highest single-pixel temperature rise and surrounding-pixel mean were recorded for each sonication. The measured temperature rises were ∼ 45% larger for aberration-corrected sonications than for noncorrected sonications. This improvement was highly significant (p skull-induced ultrasound aberration corrections. Their results suggest that UTE MRI could be used instead of CT to implement such corrections on

  19. Radiation dosimetry of the α4β2 nicotinic receptor ligand (+-[18F]flubatine, comparing preclinical PET/MRI and PET/CT to first-in-human PET/CT results

    Directory of Open Access Journals (Sweden)

    Mathias Kranz

    2016-10-01

    Full Text Available Abstract Background Both enantiomers of [18F]flubatine are new radioligands for neuroimaging of α4β2 nicotinic acetylcholine receptors with positron emission tomography (PET exhibiting promising pharmacokinetics which makes them attractive for different clinical questions. In a previous preclinical study, the main advantage of (+-[18F]flubatine compared to (−-[18F]flubatine was its higher binding affinity suggesting that (+-[18F]flubatine might be able to detect also slight reductions of α4β2 nAChRs and could be more sensitive than (−-[18F]flubatine in early stages of Alzheimer’s disease. To support the clinical translation, we investigated a fully image-based internal dosimetry approach for (+-[18F]flubatine, comparing mouse data collected on a preclinical PET/MRI system to piglet and first-in-human data acquired on a clinical PET/CT system. Time-activity curves (TACs were obtained from the three species, the animal data extrapolated to human scale, exponentially fitted and the organ doses (OD, and effective dose (ED calculated with OLINDA. Results The excreting organs (urinary bladder, kidneys, and liver receive the highest organ doses in all species. Hence, a renal/hepatobiliary excretion pathway can be assumed. In addition, the ED conversion factors of 12.1 μSv/MBq (mice, 14.3 μSv/MBq (piglets, and 23.0 μSv/MBq (humans were calculated which are well within the order of magnitude as known from other 18F-labeled radiotracers. Conclusions Although both enantiomers of [18F]flubatine exhibit different binding kinetics in the brain due to the respective affinities, the effective dose revealed no enantiomer-specific differences among the investigated species. The preclinical dosimetry and biodistribution of (+-[18F]flubatine was shown and the feasibility of a dose assessment based on image data acquired on a small animal PET/MR and a clinical PET/CT was demonstrated. Additionally, the first-in-human study confirmed the tolerability

  20. Metabolic liver function in humans measured by 2-(18)F-fluoro-2-deoxy-D-galactose PET/CT-reproducibility and clinical potential

    DEFF Research Database (Denmark)

    Bak-Fredslund, Kirstine P; Lykke Eriksen, Peter; Munk, Ole L

    2017-01-01

    Background: PET/CT with the radioactively labelled galactose analogue 2-18F-fluoro-2-deoxy-D-galactose (18F-FDGal) can be used to quantify the hepatic metabolic function and visualise regional metabolic heterogeneity. We determined the day-to-day variation in humans with and without liver disease....... Furthermore, we examined whether the standardised uptake value (SUV) of 18F-FDGal from static scans can substitute the hepatic systemic clearance of 18F- FDGal (Kmet, mL blood/min/mL liver tissue/) quantified from dynamic scans as measure of metabolic function. Four patients with cirrhosis and six healthy...... subjects underwent two 18F-FDGal PET/CT scans within a median interval of 15 days for determination of day-to-day variation. The correlation between Kmet and SUV was examined using scan data and measured arterial blood concentrations of 18F-FDGal (blood samples) from 14 subjects from previous studies...

  1. PET/CT presentation of primary effusion lymphoma-like lymphoma unrelated to human herpes virus 8, a rare NHL subtype

    International Nuclear Information System (INIS)

    Patil, Vivek V; Sideras, Panagiotis; Machac, Josef

    2014-01-01

    We present a 71-year-old female with human herpes virus 8 (HHV8)-unrelated primary effusion lymphoma (PEL)-like lymphoma. Dyspnea and pericardial effusion led to pericardiocentesis, diagnosing diffuse large B-cell lymphoma. She underwent positron emission tomography/computed tomography (PET/CT), which demonstrated hypermetabolic pericardial, pleural, and ascites fluid without lymphadenopathy elsewhere. Malignant fluid in the absence of lymphadenopathy is a hallmark of PEL. PEL is associated with immunodeficiency states such as acquired immunodeficiency syndrome (AIDS) and infectious agents such as HHV8. Our patient had no such history and had not received immunosuppressive chemotherapy. We present the PET/CT findings of this rare case of HHV8-unrelated PEL-like lymphoma

  2. Checklist and scoring system for the assessment of soft tissue preservation in CT examinations of human mummies. Application to the Tyrolean Iceman

    Energy Technology Data Exchange (ETDEWEB)

    Panzer, Stephanie [Trauma Center Murnau (Germany). Dept. of Radiology; Trauma Center Murnau and Paracelsus Medical Univ. Salzburg, Murnau (Germany). Inst. of Biomechanics; Pernter, Patrizia [Central Hospital, Bozen (Italy). Dept. of Radiodiagnostics; Piombino-Mascali, Dario; Jankauskas, Rimantas [Vilnius Univ. (Lithuania). Dept. of Anatomy, Histology and Anthropology; Zesch, Stephanie; Rosendahl, Wilfried [Reiss-Engelhorn Museen, Mannheim (Germany). German Mummy Project; Hotz, Gerhard [Natural History Museum of Basel (Switzerland). Anthropology; Zink, Albert R. [EURAC-Institute for Mummies and the Iceman, Bolzano (Italy)

    2017-12-15

    Soft tissues make a skeleton into a mummy and they allow for a diagnosis beyond osteology. Following the approach of structured reporting in clinical radiology, a recently developed checklist was used to evaluate the soft tissue preservation status of the Tyrolean Iceman using computed tomography (CT). The purpose of this study was to apply the ''Checklist and Scoring System for the Assessment of Soft Tissue Preservation in CT Examinations of Human Mummies'' to the Tyrolean Iceman, and to compare the Iceman's soft tissue preservation score to the scores calculated for other mummies. A whole-body (CT) (SOMATOM Definition Flash, Siemens, Forchheim, Germany) consisting of five scans, performed in January 2013 in the Department of Radiodiagnostics, Central Hospital, Bolzano, was used (slice thickness 0.6mm; kilovolt ranging from80 to 140). For standardized evaluation the ''CT Checklist and Scoring System for the Assessment of Soft Tissue Preservation in Human Mummies'' was used. All checkpoints under category ''A. Soft Tissues of Head and Musculoskeletal System'' and more than half in category ''B. Organs and Organ Systems'' were observed. The scoring system accounted for a total score of 153 (out of 200). The comparison of the scores between the Iceman and three mummy collections from Vilnius, Lithuania, and Palermo, Sicily, as well as one Egyptian mummy resulted in overall higher soft tissue preservation scores for the Iceman. Application of the checklist allowed for standardized assessment and documentation of the Iceman's soft tissue preservation status. The scoring system allowed for a quantitative comparison between the Iceman and other mummies. The Iceman showed remarkable soft tissue preservation.

  3. Optimizing CT technique to reduce radiation dose: effect of changes in kVp, iterative reconstruction, and noise index on dose and noise in a human cadaver.

    Science.gov (United States)

    Chang, Kevin J; Collins, Scott; Li, Baojun; Mayo-Smith, William W

    2017-06-01

    For assessment of the effect of varying the peak kilovoltage (kVp), the adaptive statistical iterative reconstruction technique (ASiR), and automatic dose modulation on radiation dose and image noise in a human cadaver, a cadaver torso underwent CT scanning at 80, 100, 120 and 140 kVp, each at ASiR settings of 0, 30 and 50 %, and noise indices (NIs) of 5.5, 11 and 22. The volume CT dose index (CTDI vol ), image noise, and attenuation values of liver and fat were analyzed for 20 data sets. Size-specific dose estimates (SSDEs) and liver-to-fat contrast-to-noise ratios (CNRs) were calculated. Values for different combinations of kVp, ASiR, and NI were compared. The CTDI vol varied by a power of 2 with kVp values between 80 and 140 without ASiR. Increasing ASiR levels allowed a larger decrease in CTDI vol and SSDE at higher kVp than at lower kVp while image noise was held constant. In addition, CTDI vol and SSDE decreased with increasing NI at each kVp, but the decrease was greater at higher kVp than at lower kVp. Image noise increased with decreasing kVp despite a fixed NI; however, this noise could be offset with the use of ASiR. The CT number of the liver remained unchanged whereas that of fat decreased as the kVp decreased. Image noise and dose vary in a complicated manner when the kVp, ASiR, and NI are varied in a human cadaver. Optimization of CT protocols will require balancing of the effects of each of these parameters to maximize image quality while minimizing dose.

  4. Checklist and scoring system for the assessment of soft tissue preservation in CT examinations of human mummies. Application to the Tyrolean Iceman

    International Nuclear Information System (INIS)

    Panzer, Stephanie

    2017-01-01

    Soft tissues make a skeleton into a mummy and they allow for a diagnosis beyond osteology. Following the approach of structured reporting in clinical radiology, a recently developed checklist was used to evaluate the soft tissue preservation status of the Tyrolean Iceman using computed tomography (CT). The purpose of this study was to apply the ''Checklist and Scoring System for the Assessment of Soft Tissue Preservation in CT Examinations of Human Mummies'' to the Tyrolean Iceman, and to compare the Iceman's soft tissue preservation score to the scores calculated for other mummies. A whole-body (CT) (SOMATOM Definition Flash, Siemens, Forchheim, Germany) consisting of five scans, performed in January 2013 in the Department of Radiodiagnostics, Central Hospital, Bolzano, was used (slice thickness 0.6mm; kilovolt ranging from80 to 140). For standardized evaluation the ''CT Checklist and Scoring System for the Assessment of Soft Tissue Preservation in Human Mummies'' was used. All checkpoints under category ''A. Soft Tissues of Head and Musculoskeletal System'' and more than half in category ''B. Organs and Organ Systems'' were observed. The scoring system accounted for a total score of 153 (out of 200). The comparison of the scores between the Iceman and three mummy collections from Vilnius, Lithuania, and Palermo, Sicily, as well as one Egyptian mummy resulted in overall higher soft tissue preservation scores for the Iceman. Application of the checklist allowed for standardized assessment and documentation of the Iceman's soft tissue preservation status. The scoring system allowed for a quantitative comparison between the Iceman and other mummies. The Iceman showed remarkable soft tissue preservation.

  5. Characterization of the single transmembrane domain of human receptor activity-modifying protein 3 in adrenomedullin receptor internalization

    International Nuclear Information System (INIS)

    Kuwasako, Kenji; Kitamura, Kazuo; Nagata, Sayaka; Nozaki, Naomi; Kato, Johji

    2012-01-01

    Highlights: ► RAMP3 mediates CLR internalization much less effectively than does RAMP2. ► The RAMP3 TMD participates in the negative regulation of CLR/RAMP3 internalization. ► A new strategy of promoting internalization and resensitization of the receptor was found. -- Abstract: Two receptor activity-modifying proteins (RAMP2 and RAMP3) enable calcitonin receptor-like receptor (CLR) to function as two heterodimeric receptors (CLR/RAMP2 and CLR/RAMP3) for adrenomedullin (AM), a potent cardiovascular protective peptide. Following AM stimulation, both receptors undergo rapid internalization through a clathrin-dependent pathway, after which CLR/RAMP3, but not CLR/RAMP2, can be recycled to the cell surface for resensitization. However, human (h)RAMP3 mediates CLR internalization much less efficiently than does hRAMP2. Therefore, the molecular basis of the single transmembrane domain (TMD) and the intracellular domain of hRAMP3 during AM receptor internalization was investigated by transiently transfecting various RAMP chimeras and mutants into HEK-293 cells stably expressing hCLR. Flow cytometric analysis revealed that substituting the RAMP3 TMD with that of RAMP2 markedly enhanced AM-induced internalization of CLR. However, this replacement did not enhance the cell surface expression of CLR, [ 125 I]AM binding affinity or AM-induced cAMP response. More detailed analyses showed that substituting the Thr 130 –Val 131 sequence in the RAMP3 TMD with the corresponding sequence (Ile 157 –Pro 158 ) from RAMP2 significantly enhanced AM-mediated CLR internalization. In contrast, substituting the RAMP3 target sequence with Ala 130 –Ala 131 did not significantly affect CLR internalization. Thus, the RAMP3 TMD participates in the negative regulation of CLR/RAMP3 internalization, and the aforementioned introduction of the Ile–Pro sequence into the RAMP3 TMD may be a strategy for promoting receptor internalization/resensitization.

  6. Specificity and versatility of substrate binding sites in four catalytic domains of human N-terminal acetyltransferases.

    Directory of Open Access Journals (Sweden)

    Cédric Grauffel

    Full Text Available Nt-acetylation is among the most common protein modifications in eukaryotes. Although thought for a long time to protect proteins from degradation, the role of Nt-acetylation is still debated. It is catalyzed by enzymes called N-terminal acetyltransferases (NATs. In eukaryotes, several NATs, composed of at least one catalytic domain, target different substrates based on their N-terminal sequences. In order to better understand the substrate specificity of human NATs, we investigated in silico the enzyme-substrate interactions in four catalytic subunits of human NATs (Naa10p, Naa20p, Naa30p and Naa50p. To date hNaa50p is the only human subunit for which X-ray structures are available. We used the structure of the ternary hNaa50p/AcCoA/MLG complex and a structural model of hNaa10p as a starting point for multiple molecular dynamics simulations of hNaa50p/AcCoA/substrate (substrate=MLG, EEE, MKG, hNaa10p/AcCoA/substrate (substrate=MLG, EEE. Nine alanine point-mutants of the hNaa50p/AcCoA/MLG complex were also simulated. Homology models of hNaa20p and hNaa30p were built and compared to hNaa50p and hNaa10p. The simulations of hNaa50p/AcCoA/MLG reproduce the interactions revealed by the X-ray data. We observed strong hydrogen bonds between MLG and tyrosines 31, 138 and 139. Yet the tyrosines interacting with the substrate's backbone suggest that their role in specificity is limited. This is confirmed by the simulations of hNaa50p/AcCoA/EEE and hNaa10p/AcCoA/MLG, where these hydrogen bonds are still observed. Moreover these tyrosines are all conserved in hNaa20p and hNaa30p. Other amino acids tune the specificity of the S1' sites that is different for hNaa10p (acidic, hNaa20p (hydrophobic/basic, hNaa30p (basic and hNaa50p (hydrophobic. We also observe dynamic correlation between the ligand binding site and helix [Formula: see text] that tightens under substrate binding. Finally, by comparing the four structures we propose maps of the peptide

  7. Does protecting humans protect the environment? A crude examination for UK nuclear power plants and the marine environment using information in the public domain

    International Nuclear Information System (INIS)

    Brownless, G P

    2008-01-01

    Current activity around radiological protection of the environment implies concerns over the effectiveness of the current approach to this-namely if humans are adequately protected, then so are non-human species. This study uses models and data currently available in the public domain to carry out a 'quick and dirty' examination of whether protecting humans does indeed imply that other species are well protected. Using marine discharges and human habits data for operational coastal UK nuclear power stations, this study compares doses to humans and a set of reference non-human species. The study concludes that the availability of data and models, and consequent ease of studying potential effects on non-humans (as well as humans), vindicates recent efforts in this area, and that these imply a high level of protection, in general, for non-human biota from UK nuclear power station marine discharges. In general terms, the study finds that protection of non-human biota relies on taking ingestion and external exposure doses to humans into account; where only one of these pathways is considered, the level of protection of non-human biota through protection of humans would depend on the radionuclide(s) in question.

  8. Three-dimensional structure of a Streptomyces sviceus GNAT acetyltransferase with similarity to the C-terminal domain of the human GH84 O-GlcNAcase

    International Nuclear Information System (INIS)

    He, Yuan; Roth, Christian; Turkenburg, Johan P.; Davies, Gideon J.

    2013-01-01

    The crystal structure of a bacterial acetyltransferase with 27% sequence identity to the C-terminal domain of human O-GlcNAcase has been solved at 1.5 Å resolution. This S. sviceus protein is compared with known GCN5-related acetyltransferases, adding to the diversity observed in this superfamily. The mammalian O-GlcNAc hydrolysing enzyme O-GlcNAcase (OGA) is a multi-domain protein with glycoside hydrolase activity in the N-terminus and with a C-terminal domain that has low sequence similarity to known acetyltransferases, prompting speculation, albeit controversial, that the C-terminal domain may function as a histone acetyltransferase (HAT). There are currently scarce data available regarding the structure and function of this C-terminal region. Here, a bacterial homologue of the human OGA C-terminal domain, an acetyltransferase protein (accession No. ZP-05014886) from Streptomyces sviceus (SsAT), was cloned and its crystal structure was solved to high resolution. The structure reveals a conserved protein core that has considerable structural homology to the acetyl-CoA (AcCoA) binding site of GCN5-related acetyltransferases (GNATs). Calorimetric data further confirm that SsAT is indeed able to bind AcCoA in solution with micromolar affinity. Detailed structural analysis provided insight into the binding of AcCoA. An acceptor-binding cavity was identified, indicating that the physiological substrate of SsAT may be a small molecule. Consistent with recently published work, the SsAT structure further questions a HAT function for the human OGA domain

  9. Three-dimensional structure of a Streptomyces sviceus GNAT acetyltransferase with similarity to the C-terminal domain of the human GH84 O-GlcNAcase

    Energy Technology Data Exchange (ETDEWEB)

    He, Yuan [Northwest University, Xi’an 710069 (China); The University of York, York YO10 5DD (United Kingdom); Roth, Christian; Turkenburg, Johan P.; Davies, Gideon J., E-mail: gideon.davies@york.ac.uk [The University of York, York YO10 5DD (United Kingdom); Northwest University, Xi’an 710069 (China)

    2014-01-01

    The crystal structure of a bacterial acetyltransferase with 27% sequence identity to the C-terminal domain of human O-GlcNAcase has been solved at 1.5 Å resolution. This S. sviceus protein is compared with known GCN5-related acetyltransferases, adding to the diversity observed in this superfamily. The mammalian O-GlcNAc hydrolysing enzyme O-GlcNAcase (OGA) is a multi-domain protein with glycoside hydrolase activity in the N-terminus and with a C-terminal domain that has low sequence similarity to known acetyltransferases, prompting speculation, albeit controversial, that the C-terminal domain may function as a histone acetyltransferase (HAT). There are currently scarce data available regarding the structure and function of this C-terminal region. Here, a bacterial homologue of the human OGA C-terminal domain, an acetyltransferase protein (accession No. ZP-05014886) from Streptomyces sviceus (SsAT), was cloned and its crystal structure was solved to high resolution. The structure reveals a conserved protein core that has considerable structural homology to the acetyl-CoA (AcCoA) binding site of GCN5-related acetyltransferases (GNATs). Calorimetric data further confirm that SsAT is indeed able to bind AcCoA in solution with micromolar affinity. Detailed structural analysis provided insight into the binding of AcCoA. An acceptor-binding cavity was identified, indicating that the physiological substrate of SsAT may be a small molecule. Consistent with recently published work, the SsAT structure further questions a HAT function for the human OGA domain.

  10. Computer-aided polyp detection on CT colonography: Comparison of three systems in a high-risk human population

    International Nuclear Information System (INIS)

    Park, Hee Sun; Kim, Se Hyung; Kim, Jong Hyo; Lee, June-Goo; Kim, Sang Gyun; Lee, Jeong Min; Lee, Jae Young; Han, Joon Koo; Choi, Byung Ihn

    2010-01-01

    Purpose: To compare the detection performances of two commercial and one academic computer-aided diagnosis (CAD) systems for polyp detection on CT colonography (CTC) in a high-risk human population and to assess their detection characteristics. Materials and methods: This retrospective study had institutional review board approval, but informed consent was waived. Sixty-eight patients who were suspected of having colonic polyps and scheduled for colonoscopic polyp removal were included. After CTC was performed using a 64-row MDCT, two commercial (PEV, CAR) and one academic (Hessian matrix-based) CAD systems were applied to each CTC dataset. Colonoscopy using the segmental unblinded technique was performed as a standard of reference. Per-polyp and per-patient sensitivities were calculated and compared for each CAD system. The mean number of false-positives (FPs) and false-negatives (FNs) was computed and the causes of all FPs and FNs were analyzed. Results: A total of 151 polyps in 61 patients were detected (77 polyps <6 mm, 48 6-9.9 mm, 26 ≥ 10 mm). Per-polyp sensitivity for PEV, CAR, and Hessian matrix-based CAD were: 71.6%, 78.4%, and 83.8% for polyps ≥6 mm, and 88.5%, 96.2%, and 96.2% for polyps ≥10 mm. Per-patient sensitivity for polyps ≥6 mm was 80.4%, 89.1%, and 93.5%, and 87%, 95.7%, and 95.7% for polyps ≥10 mm, respectively. Per-polyp and per-patient sensitivities were not significantly different among the three CAD systems regardless of size threshold. Mean number of FPs was 6.9 for PEV, 7.3 for CAR, and 14 for Hessian matrix-based CAD. The most common cause of FPs were feces, followed by extracolonic findings, prominent folds and ileocecal valve, and rectal tube. The distribution of the causes of FPs was significantly different among the three systems. Conclusion: Sensitivity of the three CAD systems for polyp detection was comparable regardless of the polyp size threshold; however, the number of FPs was higher in the Hessian matrix-based CAD

  11. Crystallization and preliminary X-ray analysis of Na-ASP-1, a multi-domain pathogenesis-related-1 protein from the human hookworm parasite Necator americanus

    International Nuclear Information System (INIS)

    Asojo, Oluwatoyin A.; Loukas, Alex; Inan, Mehmet; Barent, Rick; Huang, Jicai; Plantz, Brad; Swanson, Amber; Gouthro, Mark; Meagher, Michael M.; Hotez, Peter J.

    2005-01-01

    In order to clarify the structural basis of the pathogenesis-related-1 domain, Na-ASP-1, the first multi-domain ASP from the human hookworm parasite N. americanus, has been crystallized. 2.2 Å resolution data have been collected from a crystal belonging to the monoclinic space group P2 1 . Human hookworm infection is a major cause of anemia and malnutrition in the developing world. In an effort to control hookworm infection, the Human Hookworm Vaccine Initiative has identified candidate vaccine antigens from the infective larval stage (L3) of the parasite, including a family of pathogenesis-related-1 (PR-1) proteins known as the ancylostoma-secreted proteins (ASPs). The functions of the ASPs are unknown. In addition, it is unclear why some ASPs have one while others have multiple PR-1 domains. There are no known structures of a multi-domain ASP and in an effort to remedy this situation, recombinant Na-ASP-1 has been expressed, purified and crystallized. Na-ASP-1 is a 406-amino-acid multi-domain ASP from the prevalent human hookworm parasite Necator americanus. Useful X-ray data to 2.2 Å have been collected from a crystal that belongs to the monoclinic space group P2 1 with unit-cell parameters a = 67.7, b = 74.27, c = 84.60 Å, β = 112.12°. An initial molecular-replacement solution has been obtained with one monomer in the asymmetric unit

  12. Crystallization and preliminary X-ray analysis of Na-ASP-1, a multi-domain pathogenesis-related-1 protein from the human hookworm parasite Necator americanus

    Energy Technology Data Exchange (ETDEWEB)

    Asojo, Oluwatoyin A., E-mail: oasojo@unmc.edu [Eppley Institute for Research in Cancer and Allied Diseases, 987696 Nebraska Medical Center, Omaha, NE 68198-7696 (United States); Loukas, Alex [Department of Microbiology and Tropical Medicine, The George Washington University Medical Center, Washington DC 20037 (United States); Division of Infectious Diseases and Immunology, Queensland Institute of Medical Research, Brisbane, QLD 4006 (Australia); Inan, Mehmet; Barent, Rick; Huang, Jicai; Plantz, Brad; Swanson, Amber; Gouthro, Mark; Meagher, Michael M. [Department of Chemical Engineering, The University of Nebraska-Lincoln, Lincoln, NE 68588-0643 (United States); Hotez, Peter J. [Department of Microbiology and Tropical Medicine, The George Washington University Medical Center, Washington DC 20037 (United States); Eppley Institute for Research in Cancer and Allied Diseases, 987696 Nebraska Medical Center, Omaha, NE 68198-7696 (United States)

    2005-04-01

    In order to clarify the structural basis of the pathogenesis-related-1 domain, Na-ASP-1, the first multi-domain ASP from the human hookworm parasite N. americanus, has been crystallized. 2.2 Å resolution data have been collected from a crystal belonging to the monoclinic space group P2{sub 1}. Human hookworm infection is a major cause of anemia and malnutrition in the developing world. In an effort to control hookworm infection, the Human Hookworm Vaccine Initiative has identified candidate vaccine antigens from the infective larval stage (L3) of the parasite, including a family of pathogenesis-related-1 (PR-1) proteins known as the ancylostoma-secreted proteins (ASPs). The functions of the ASPs are unknown. In addition, it is unclear why some ASPs have one while others have multiple PR-1 domains. There are no known structures of a multi-domain ASP and in an effort to remedy this situation, recombinant Na-ASP-1 has been expressed, purified and crystallized. Na-ASP-1 is a 406-amino-acid multi-domain ASP from the prevalent human hookworm parasite Necator americanus. Useful X-ray data to 2.2 Å have been collected from a crystal that belongs to the monoclinic space group P2{sub 1} with unit-cell parameters a = 67.7, b = 74.27, c = 84.60 Å, β = 112.12°. An initial molecular-replacement solution has been obtained with one monomer in the asymmetric unit.

  13. Quantum Mechanics and Molecular Mechanics Study of the Catalytic Mechanism of Human AMSH-LP Domain Deubiquitinating Enzymes.

    Science.gov (United States)

    Zhu, Wenyou; Liu, Yongjun; Ling, Baoping

    2015-08-25

    Deubiquitinating enzymes (DUBs) catalyze the cleavage of the isopeptide bond in polyubiquitin chains to control and regulate the deubiquitination process in all known eukaryotic cells. The human AMSH-LP DUB domain specifically cleaves the isopeptide bonds in the Lys63-linked polyubiquitin chains. In this article, the catalytic mechanism of AMSH-LP has been studied using a combined quantum mechanics and molecular mechanics method. Two possible hydrolysis processes (Path 1 and Path 2) have been considered. Our calculation results reveal that the activation of Zn(2+)-coordinated water molecule is the essential step for the hydrolysis of isopeptide bond. In Path 1, the generated hydroxyl first attacks the carbonyl group of Gly76, and then the amino group of Lys63 is protonated, which is calculated to be the rate limiting step with an energy barrier of 13.1 kcal/mol. The energy barrier of the rate limiting step and the structures of intermediate and product are in agreement with the experimental results. In Path 2, the protonation of amino group of Lys63 is prior to the nucleophilic attack of activated hydroxyl. The two proton transfer processes in Path 2 correspond to comparable overall barriers (33.4 and 36.1 kcal/mol), which are very high for an enzymatic reaction. Thus, Path 2 can be ruled out. During the reaction, Glu292 acts as a proton transfer mediator, and Ser357 mainly plays a role in stabilizing the negative charge of Gly76. Besides acting as a Lewis acid, Zn(2+) also influences the reaction by coordinating to the reaction substrates (W1 and Gly76).

  14. Human TRPA1 is intrinsically cold- and chemosensitive with and without its N-terminal ankyrin repeat domain.

    Science.gov (United States)

    Moparthi, Lavanya; Survery, Sabeen; Kreir, Mohamed; Simonsen, Charlotte; Kjellbom, Per; Högestätt, Edward D; Johanson, Urban; Zygmunt, Peter M

    2014-11-25

    We have purified and reconstituted human transient receptor potential (TRP) subtype A1 (hTRPA1) into lipid bilayers and recorded single-channel currents to understand its inherent thermo- and chemosensory properties as well as the role of the ankyrin repeat domain (ARD) of the N terminus in channel behavior. We report that hTRPA1 with and without its N-terminal ARD (Δ1-688 hTRPA1) is intrinsically cold-sensitive, and thus, cold-sensing properties of hTRPA1 reside outside the N-terminal ARD. We show activation of hTRPA1 by the thiol oxidant 2-((biotinoyl)amino)ethyl methanethiosulfonate (MTSEA-biotin) and that electrophilic compounds activate hTRPA1 in the presence and absence of the N-terminal ARD. The nonelectrophilic compounds menthol and the cannabinoid Δ(9)-tetrahydrocannabiorcol (C16) directly activate hTRPA1 at different sites independent of the N-terminal ARD. The TRPA1 antagonist HC030031 inhibited cold and chemical activation of hTRPA1 and Δ1-688 hTRPA1, supporting a direct interaction with hTRPA1 outside the N-terminal ARD. These findings show that hTRPA1 is an intrinsically cold- and chemosensitive ion channel. Thus, second messengers, including Ca(2+), or accessory proteins are not needed for hTRPA1 responses to cold or chemical activators. We suggest that conformational changes outside the N-terminal ARD by cold, electrophiles, and nonelectrophiles are important in hTRPA1 channel gating and that targeting chemical interaction sites outside the N-terminal ARD provides possibilities to fine tune TRPA1-based drug therapies (e.g., for treatment of pain associated with cold hypersensitivity and cardiovascular disease).

  15. Aerobic exercise training differentially affects ACE C- and N-domain activities in humans: Interactions with ACE I/D polymorphism and association with vascular reactivity.

    Science.gov (United States)

    Alves, Cléber Rene; Fernandes, Tiago; Lemos, José Ribeiro; Magalhães, Flávio de Castro; Trombetta, Ivani Credidio; Alves, Guilherme Barreto; Mota, Glória de Fátima Alves da; Dias, Rodrigo Gonçalves; Pereira, Alexandre Costa; Krieger, José Eduardo; Negrão, Carlos Eduardo; Oliveira, Edilamar Menezes

    2018-01-01

    Previous studies have linked angiotensin-converting enzyme ( ACE) insertion (I)/deletion (D) polymorphism (II, ID and DD) to physical performance. Moreover, ACE has two catalytic domains: NH2 (N) and COOH (C) with distinct functions, and their activity has been found to be modulated by ACE polymorphism. The aim of the present study is to investigate the effects of the interaction between aerobic exercise training (AET) and ACE I/D polymorphism on ACE N- and C-domain activities and vascular reactivity in humans. A total of 315 pre-selected healthy males were genotyped for II, ID and DD genotypes. Fifty completed the full AET (II, n = 12; ID, n = 25; and DD, n = 13), performed in three 90-minute sessions weekly, in the four-month exercise protocol. Pre- and post-training resting heart rate (HR), peak O 2 consumption (VO 2 peak), mean blood pressure (MBP), forearm vascular conduction (FVC), total circulating ACE and C- and N-domain activities were assessed. One-way ANOVA and two -way repeated-measures ANOVA were used. In pre-training, all variables were similar among the three genotypes. In post-training, a similar increase in FVC (35%) was observed in the three genotypes. AET increased VO 2 peak similarly in II, ID and DD (49±2 vs. 57±1; 48±1 vs. 56±3; and 48±5 vs. 58±2 ml/kg/min, respectively). Moreover, there were no changes in HR and MBP. The DD genotype was also associated with greater ACE and C-domain activities at pre- and post-training when compared to II. AET decreased similarly the total ACE and C-domain activities in all genotypes, while increasing the N-domain activity in the II and DD genotypes. However, interestingly, the measurements of N-domain activity after training indicate a greater activity than the other genotypes. These results suggest that the vasodilation in response to AET may be associated with the decrease in total ACE and C-domain activities, regardless of genotype, and that the increase in N-domain activity is dependent on the DD

  16. Domain analysis

    DEFF Research Database (Denmark)

    Hjørland, Birger

    2017-01-01

    The domain-analytic approach to knowledge organization (KO) (and to the broader field of library and information science, LIS) is outlined. The article reviews the discussions and proposals on the definition of domains, and provides an example of a domain-analytic study in the field of art studies....... Varieties of domain analysis as well as criticism and controversies are presented and discussed....

  17. Crystallization and preliminary X-ray diffraction studies of the tetramerization domain derived from the human potassium channel Kv1.3

    International Nuclear Information System (INIS)

    Winklmeier, Andreas; Weyand, Michael; Schreier, Christina; Kalbitzer, Hans Robert; Kremer, Werner

    2009-01-01

    The tetramerization domain of human Kv1.3 was cloned, expressed, purified and crystallized. The crystals belonged to space group I4 and diffracted to 1.2 Å resolution using synchrotron radiation. The tetramerization domain (T1 domain) derived from the voltage-dependent potassium channel Kv1.3 of Homo sapiens was recombinantly expressed in Escherichia coli and purified. The crystals were first grown in an NMR tube in 150 mM potassium phosphate pH 6.5 in the absence of additional precipitants. The crystals showed I4 symmetry characteristic of the naturally occurring tetrameric assembly of the single subunits. A complete native data set was collected to 1.2 Å resolution at 100 K using synchrotron radiation

  18. Observation of the human fetal corpses with maxillofacial malformations. 1. CT and MRI examinations of the fetal cleft lip and/or palate

    International Nuclear Information System (INIS)

    Saito, Chikara; Nakano, Yoko; Shigematsu, Shiro

    1999-01-01

    Of the various types of congenital malformations, the cleft lip and/or palate is one of the most frequent. Observation of human fetal corpses exhibiting cleft lip and palate is very important to research on its onset of its mechanism and development. In recent years, some of researchers have performed clinical studies on prenatal diagnosis and surgical treatment for the entirety. However, there have hardly been any reports on detailed observations of the maxillofacial structure of a fetus with cleft lip and palate. We seized an opportunity of observing the maxillofacial structure of fetuses with cleft lip and/or palate using three-dimensional CT (3D-CT) and MR imaging as non-disjunctive methods. In the present study, nine fetal corpses having cleft lip and/or palate were examined. The results were as follows: CT and MRI were useful for non-invasive observation of the maxillofacial structure, including soft tissues. Because the osseous tissues of young fetus tissue is not fully mature, observation of bone structures was slightly difficult. When corpses were immersed in formalin for a long time, osseous tissue was decalcified, thus making it difficult to obtain clear images. We could observe the details of the maxillofacial structures such as the alveolar process, the hard palate, the maxillary sinus, the nasal cavity, the nasal bone, and the vomer, in some of the cases. 3D-CT and MR findings observed in the fetuses with cleft lip and/or palate should provide some basement of the imaging diagnosis of congenital disorder. (author))

  19. Prognostic value of pretreatment 18F-FDG PET/CT and human papillomavirus type 16 testing in locally advanced oropharyngeal squamous cell carcinoma

    International Nuclear Information System (INIS)

    Cheng, Nai-Ming; Yen, Tzu-Chen; Chang, Joseph Tung-Chieh; Tsan, Din-Li; Lin, Chien-Yu; Huang, Chung-Guei; Ng, Shu-Hang; Wang, Hung-Ming; Hsu, Cheng-Lung; Liao, Chun-Ta

    2012-01-01

    Human papillomavirus type 16 (HPV-16) positivity is associated with favourable survival in oropharyngeal squamous cell carcinoma (OPSCC). We report here a study of the prognostic significance of 18 F-FDG PET/CT functional parameters and HPV-16 infection in OPSCC patients. We retrospectively analysed 60 patients with stage III or IV OPSCC who had had a pretherapy 18 F-FDG PET/CT scan and had completed concurrent chemoradiotherapy (n = 58) or curative radiotherapy (n = 2). All patients were followed up for ≥24 months or until death. We determined total lesion glycolysis (TLG) and the maximal standardized uptake values (SUV max ) of the primary tumour and neck lymph nodes from the pretherapy 18 F-FDG PET/CT scan. Optimal cut-offs of the 18 F-FDG PET/CT parameters were obtained by receiver operating characteristic (ROC) curve analyses. Pretherapy tumour biopsies were studied by polymerase chain reaction to determine HPV infection status. The pretherapy tumour biopsies were positive for HPV-16 in 12 patients (20.0 %). Cox regression analyses revealed HPV-16 positivity and tumour TLG >135.3 g to be independently associated with overall survival (p = 0.027 and 0.011, respectively). However, only tumour TLG >135.3 g was independently associated with progression-free survival, disease-free survival and locoregional control (p = 0.011, 0.001 and 0.034, respectively). A scoring system was formulated to define distinct overall survival groups using tumour TLG and HPV-16 status. Patients positive for HPV-16 and with tumour TLG ≤135.3 g experienced better survival than those with tumour TLG >135.3 g and no HPV infection (p = 0.001). Tumour TLG was an independent predictor of survival in patients with locally advanced OPSCC. A scoring system was developed and may serve as a risk stratification strategy for guiding therapy. (orig.)

  20. Observation of the human fetal corpses with maxillofacial malformations. 1. CT and MRI examinations of the fetal cleft lip and/or palate

    Energy Technology Data Exchange (ETDEWEB)

    Saito, Chikara; Nakano, Yoko; Shigematsu, Shiro [Tokyo Dental Coll., Chiba (Japan)] (and others)

    1999-06-01

    Of the various types of congenital malformations, the cleft lip and/or palate is one of the most frequent. Observation of human fetal corpses exhibiting cleft lip and palate is very important to research on its onset of its mechanism and development. In recent years, some of researchers have performed clinical studies on prenatal diagnosis and surgical treatment for the entirey. However, there have hardly been any reports on detailed observations of the maxillofacial structure of a fetus with cleft lip and palate. We seized an opportunity of observing the maxillofacial structure of fetuses with cleft lip and/or palate using three-dimensional CT (3D-CT) and MR imaging as non-disjunctive methods. In the present study, nine fetal corpses having cleft lip and/or palate were examined. The results were as follows: CT and MRI were useful for non-invasive observation of the maxillofacial structure, including soft tissues. Because the osseous tissues of young fetus tissue is not fully mature, observation of bone structures was slightly difficult. When corpses were immersed in formalin for a long time, osseous tissue was decalcified, thus making it difficult to obtain clear images. We could observe the details of the maxillofacial structures such as the alveolar process, the hard palate, the maxillary sinus, the nasal cavity, the nasal bone, and the vomer, in some of the cases. 3D-CT and MR findings observed in the fetuses with cleft lip and/or palate should provide some basement of the imaging diagnosis of congenital disorder. (author)

  1. Prognostic value of pretreatment {sup 18}F-FDG PET/CT and human papillomavirus type 16 testing in locally advanced oropharyngeal squamous cell carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Cheng, Nai-Ming; Yen, Tzu-Chen [Chang Gung University College of Medicine, Department of Nuclear Medicine and Molecular Imaging Center, Chang Gung Memorial Hospital, Taipei (China); Chang, Joseph Tung-Chieh; Tsan, Din-Li; Lin, Chien-Yu [Chang Gung University College of Medicine, Department of Radiation Oncology, Chang Gung Memorial Hospital, Taipei (China); Huang, Chung-Guei [Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Department of Laboratory Medicine, Taipei (China); Ng, Shu-Hang [Chang Gung University College of Medicine, Department of Diagnostic Radiology, Chang Gung Memorial Hospital, Taipei (China); Wang, Hung-Ming; Hsu, Cheng-Lung [Chang Gung University College of Medicine, Division of Hematology/Oncology, Department of Internal Medicine, Chang Gung Memorial Hospital, Taipei (China); Liao, Chun-Ta [Chang Gung University College of Medicine, Department of Otolaryngology-Head and Neck Surgery, Chang Gung Memorial Hospital, Taipei (China)

    2012-11-15

    Human papillomavirus type 16 (HPV-16) positivity is associated with favourable survival in oropharyngeal squamous cell carcinoma (OPSCC). We report here a study of the prognostic significance of {sup 18}F-FDG PET/CT functional parameters and HPV-16 infection in OPSCC patients. We retrospectively analysed 60 patients with stage III or IV OPSCC who had had a pretherapy {sup 18}F-FDG PET/CT scan and had completed concurrent chemoradiotherapy (n = 58) or curative radiotherapy (n = 2). All patients were followed up for {>=}24 months or until death. We determined total lesion glycolysis (TLG) and the maximal standardized uptake values (SUV{sub max}) of the primary tumour and neck lymph nodes from the pretherapy {sup 18}F-FDG PET/CT scan. Optimal cut-offs of the {sup 18}F-FDG PET/CT parameters were obtained by receiver operating characteristic (ROC) curve analyses. Pretherapy tumour biopsies were studied by polymerase chain reaction to determine HPV infection status. The pretherapy tumour biopsies were positive for HPV-16 in 12 patients (20.0 %). Cox regression analyses revealed HPV-16 positivity and tumour TLG >135.3 g to be independently associated with overall survival (p = 0.027 and 0.011, respectively). However, only tumour TLG >135.3 g was independently associated with progression-free survival, disease-free survival and locoregional control (p = 0.011, 0.001 and 0.034, respectively). A scoring system was formulated to define distinct overall survival groups using tumour TLG and HPV-16 status. Patients positive for HPV-16 and with tumour TLG {<=}135.3 g experienced better survival than those with tumour TLG >135.3 g and no HPV infection (p = 0.001). Tumour TLG was an independent predictor of survival in patients with locally advanced OPSCC. A scoring system was developed and may serve as a risk stratification strategy for guiding therapy. (orig.)

  2. A Human Antibody That Binds to the Sixth Ig-Like Domain of VCAM-1 Blocks Lung Cancer Cell Migration In Vitro

    Directory of Open Access Journals (Sweden)

    Mi Ra Kim

    2017-03-01

    Full Text Available Vascular cell adhesion molecule-1 (VCAM-1 is closely associated with tumor progression and metastasis. However, the relevance and role of VCAM-1 in lung cancer have not been clearly elucidated. In this study, we found that VCAM-1 was highly overexpressed in lung cancer tissue compared with that of normal lung tissue, and high VCAM-1 expression correlated with poor survival in lung cancer patients. VCAM-1 knockdown reduced migration of A549 human lung cancer cells into Matrigel, and competitive blocking experiments targeting the Ig-like domain 6 of VCAM-1 (VCAM-1-D6 demonstrated that the VCAM-1-D6 domain was critical for VCAM-1 mediated A549 cell migration into Matrigel. Next, we developed a human monoclonal antibody specific to human and mouse VCAM-1-D6 (VCAM-1-D6 huMab, which was isolated from a human synthetic antibody library using phage display technology. Finally, we showed that VCAM-1-D6 huMab had a nanomolar affinity for VCAM-1-D6 and that it potently suppressed the migration of A549 and NCI-H1299 lung cancer cell lines into Matrigel. Taken together, these results suggest that VCAM-1-D6 is a key domain for regulating VCAM-1-mediated lung cancer invasion and that our newly developed VCAM-1-D6 huMab will be a useful tool for inhibiting VCAM-1-expressing lung cancer cell invasion.

  3. Hum-mPLoc 3.0: prediction enhancement of human protein subcellular localization through modeling the hidden correlations of gene ontology and functional domain features.

    Science.gov (United States)

    Zhou, Hang; Yang, Yang; Shen, Hong-Bin

    2017-03-15

    Protein subcellular localization prediction has been an important research topic in computational biology over the last decade. Various automatic methods have been proposed to predict locations for large scale protein datasets, where statistical machine learning algorithms are widely used for model construction. A key step in these predictors is encoding the amino acid sequences into feature vectors. Many studies have shown that features extracted from biological domains, such as gene ontology and functional domains, can be very useful for improving the prediction accuracy. However, domain knowledge usually results in redundant features and high-dimensional feature spaces, which may degenerate the performance of machine learning models. In this paper, we propose a new amino acid sequence-based human protein subcellular location prediction approach Hum-mPLoc 3.0, which covers 12 human subcellular localizations. The sequences are represented by multi-view complementary features, i.e. context vocabulary annotation-based gene ontology (GO) terms, peptide-based functional domains, and residue-based statistical features. To systematically reflect the structural hierarchy of the domain knowledge bases, we propose a novel feature representation protocol denoted as HCM (Hidden Correlation Modeling), which will create more compact and discriminative feature vectors by modeling the hidden correlations between annotation terms. Experimental results on four benchmark datasets show that HCM improves prediction accuracy by 5-11% and F 1 by 8-19% compared with conventional GO-based methods. A large-scale application of Hum-mPLoc 3.0 on the whole human proteome reveals proteins co-localization preferences in the cell. www.csbio.sjtu.edu.cn/bioinf/Hum-mPLoc3/. hbshen@sjtu.edu.cn. Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com

  4. Transfer of C-terminal residues of human apolipoprotein A-I to insect apolipophorin III creates a two-domain chimeric protein with enhanced lipid binding activity.

    Science.gov (United States)

    Horn, James V C; Ellena, Rachel A; Tran, Jesse J; Beck, Wendy H J; Narayanaswami, Vasanthy; Weers, Paul M M

    2017-08-01

    Apolipophorin III (apoLp-III) is an insect apolipoprotein (18kDa) that comprises a single five-helix bundle domain. In contrast, human apolipoprotein A-I (apoA-I) is a 28kDa two-domain protein: an α-helical N-terminal domain (residues 1-189) and a less structured C-terminal domain (residues 190-243). To better understand the apolipoprotein domain organization, a novel chimeric protein was engineered by attaching residues 179 to 243 of apoA-I to the C-terminal end of apoLp-III. The apoLp-III/apoA-I chimera was successfully expressed and purified in E. coli. Western blot analysis and mass spectrometry confirmed the presence of the C-terminal domain of apoA-I within the chimera. While parent apoLp-III did not self-associate, the chimera formed oligomers similar to apoA-I. The chimera displayed a lower α-helical content, but the stability remained similar compared to apoLp-III, consistent with the addition of a less structured domain. The chimera was able to solubilize phospholipid vesicles at a significantly higher rate compared to apoLp-III, approaching that of apoA-I. The chimera was more effective in protecting phospholipase C-treated low density lipoprotein from aggregation compared to apoLp-III. In addition, binding interaction of the chimera with phosphatidylglycerol vesicles and lipopolysaccharides was considerably improved compared to apoLp-III. Thus, addition of the C-terminal domain of apoA-I to apoLp-III created a two-domain protein, with self-association, lipid and lipopolysaccharide binding properties similar to apoA-I. The apoA-I like behavior of the chimera indicate that these properties are independent from residues residing in the N-terminal domain of apoA-I, and that they can be transferred from apoA-I to apoLp-III. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of the central zinc-binding domain of the human Mcm10 DNA-replication factor

    International Nuclear Information System (INIS)

    Jung, Nam Young; Bae, Won Jin; Chang, Jeong Ho; Kim, Young Chang; Cho, Yunje

    2008-01-01

    Mcm10 is a highly conserved nuclear protein that plays a key role in the initiation and elongation processes of DNA replication by providing a physical link between the Mcm2–7 complex and DNA polymerases. In this study, the central domain of human Mcm10 was crystallized using the hanging-drop vapour-diffusion method in the presence of PEG 3350. The initiation of eukaryotic DNA replication requires the tightly controlled assembly of a set of replication factors. Mcm10 is a highly conserved nuclear protein that plays a key role in the initiation and elongation processes of DNA replication by providing a physical link between the Mcm2–7 complex and DNA polymerases. The central domain, which contains the CCCH zinc-binding motif, is most conserved within Mcm10 and binds to DNA and several proteins, including proliferative cell nuclear antigen. In this study, the central domain of human Mcm10 was crystallized using the hanging-drop vapour-diffusion method in the presence of PEG 3350. An X-ray diffraction data set was collected to a resolution of 2.6 Å on a synchrotron beamline. The crystals formed belonged to space group R3, with unit-cell parameters a = b = 99.5, c = 133.0 Å. According to Matthews coefficient calculations, the crystals were predicted to contain six MCM10 central domain molecules in the asymmetric unit

  6. Identification of a novel splice variant of human PD-L1 mRNA encoding an isoform-lacking Igv-like domain.

    Science.gov (United States)

    He, Xian-hui; Xu, Li-hui; Liu, Yi

    2005-04-01

    To investigate the expression and regulation of PD-1 ligand 1 (PD-L1) in peripheral blood mononuclear cells (PBMC). The cDNA encoding human PD-L1 precursor was cloned from the total RNA extracted from the resting and phorbol dibutyrate plus ionomycin- or phytohemagglutinin-activated PBMC, by reverse transcription polymerase chain reaction (RT-PCR), and independent clones were sequenced and analyzed. The expression and subcellular localization were examined in transiently transfected cells. The PD-L1 gene expression in different PBMC was also analyzed by RT-PCR. A novel human PD-L1 splice variant was identified from the activated PBMC. It was generated by splicing out exon? encoding an immunoglobulin variable domain (Igv)-like domain but retaining all other exons without a frame-shift. Consequently, the putative translated protein contained all other domains including the transmembrane region except for the Igv-like domain. Furthermore, the conventional isoform was expressed on the plasma surface whereas the novel isoform showed a pattern of intracellular membrane distribution in transiently transfected K562 cells. In addition, the expression pattern of the PD-L1 splice variant was variable in different individuals and in different cellular status. PD-L1 expression may be regulated at the posttranscriptional level through alternative splicing, and modulation of the PD-L1 isoform expression may influence the outcome of specific immune responses in the peripheral tissues.

  7. Effect of calcium ions on structure and stability of the C1q-like domain of otolin-1 from human and zebrafish.

    Science.gov (United States)

    Hołubowicz, Rafał; Wojtas, Magdalena; Taube, Michał; Kozak, Maciej; Ożyhar, Andrzej; Dobryszycki, Piotr

    2017-12-01

    Otolin-1 is a collagen-like protein expressed in the inner ear of vertebrates. It provides an organic scaffold for otoliths in fish and otoconia in land vertebrates. In this study, the expression and purification procedure of C1q-like domain of otolin-1 from human and zebrafish was developed. The structure and stability of the proteins were investigated. The results of sedimentation velocity analytical ultracentrifugation and small-angle X-ray scattering indicated that the C1q-like domain of otolin-1 forms stable trimers in solution in the presence of calcium ions. It was also observed that calcium ions influenced the secondary structure of the proteins. C1q-like domains were stabilized by the calcium ions. The human variant was especially affected by the calcium ions. The results indicate the importance of the C1q-like domain for the assembly of the organic matrix of otoliths and otoconia. © 2017 Federation of European Biochemical Societies.

  8. Correlation between a 2D channelized Hotelling observer and human observers in a low-contrast detection task with multislice reading in CT.

    Science.gov (United States)

    Yu, Lifeng; Chen, Baiyu; Kofler, James M; Favazza, Christopher P; Leng, Shuai; Kupinski, Matthew A; McCollough, Cynthia H

    2017-08-01

    Model observers have been successfully developed and used to assess the quality of static 2D CT images. However, radiologists typically read images by paging through multiple 2D slices (i.e., multislice reading). The purpose of this study was to correlate human and model observer performance in a low-contrast detection task performed using both 2D and multislice reading, and to determine if the 2D model observer still correlate well with human observer performance in multislice reading. A phantom containing 18 low-contrast spheres (6 sizes × 3 contrast levels) was scanned on a 192-slice CT scanner at five dose levels (CTDI vol = 27, 13.5, 6.8, 3.4, and 1.7 mGy), each repeated 100 times. Images were reconstructed using both filtered-backprojection (FBP) and an iterative reconstruction (IR) method (ADMIRE, Siemens). A 3D volume of interest (VOI) around each sphere was extracted and placed side-by-side with a signal-absent VOI to create a 2-alternative forced choice (2AFC) trial. Sixteen 2AFC studies were generated, each with 100 trials, to evaluate the impact of radiation dose, lesion size and contrast, and reconstruction methods on object detection. In total, 1600 trials were presented to both model and human observers. Three medical physicists acted as human observers and were allowed to page through the 3D volumes to make a decision for each 2AFC trial. The human observer performance was compared with the performance of a multislice channelized Hotelling observer (CHO_MS), which integrates multislice image data, and with the performance of previously validated CHO, which operates on static 2D images (CHO_2D). For comparison, the same 16 2AFC studies were also performed in a 2D viewing mode by the human observers and compared with the multislice viewing performance and the two CHO models. Human observer performance was well correlated with the CHO_2D performance in the 2D viewing mode [Pearson product-moment correlation coefficient R = 0.972, 95% confidence

  9. Correlation between a 2D Channelized Hotelling Observer and Human Observers in a Low-contrast Detection Task with Multi-slice Reading in CT

    Science.gov (United States)

    Yu, Lifeng; Chen, Baiyu; Kofler, James M.; Favazza, Christopher P.; Leng, Shuai; Kupinski, Matthew A.; McCollough, Cynthia H.

    2017-01-01

    Purpose Model observers have been successfully developed and used to assess the quality of static 2D CT images. However, radiologists typically read images by paging through multiple 2D slices (i.e. multi-slice reading). The purpose of this study was to correlate human and model observer performance in a low-contrast detection task performed using both 2D and multi-slice reading, and to determine if the 2D model observer still correlate well with human observer performance in multi-slice reading. Methods A phantom containing 18 low-contrast spheres (6 sizes × 3 contrast levels) was scanned on a 192-slice CT scanner at 5 dose levels (CTDIvol = 27, 13.5, 6.8, 3.4, and 1.7 mGy), each repeated 100 times. Images were reconstructed using both filtered-backprojection (FBP) and an iterative reconstruction (IR) method (ADMIRE, Siemens). A 3D volume of interest (VOI) around each sphere was extracted and placed side-by-side with a signal-absent VOI to create a 2-alternative forced choice (2AFC) trial. Sixteen 2AFC studies were generated, each with 100 trials, to evaluate the impact of radiation dose, lesion size and contrast, and reconstruction methods on object detection. In total, 1600 trials were presented to both model and human observers. Three medical physicists acted as human observers and were allowed to page through the 3D volumes to make a decision for each 2AFC trial. The human observer performance was compared with the performance of a multi-slice channelized Hotelling observer (CHO_MS), which integrates multi-slice image data, and with the performance of previously validated CHO, which operates on static 2D images (CHO_2D). For comparison, the same 16 2AFC studies were also performed in a 2D viewing mode by the human observers and compared with the multi-slice viewing performance and the two CHO models. Results Human observer performance was well correlated with the CHO_2D performance in the 2D viewing mode (Pearson product-moment correlation coefficient R=0

  10. The human TREM gene cluster at 6p21.1 encodes both activating and inhibitory single IgV domain receptors and includes NKp44.

    Science.gov (United States)

    Allcock, Richard J N; Barrow, Alexander D; Forbes, Simon; Beck, Stephan; Trowsdale, John

    2003-02-01

    We have characterized a cluster of single immunoglobulin variable (IgV) domain receptors centromeric of the major histocompatibility complex (MHC) on human chromosome 6. In addition to triggering receptor expressed on myeloid cells (TREM)-1 and TREM2, the cluster contains NKp44, a triggering receptor whose expression is limited to NK cells. We identified three new related genes and two gene fragments within a cluster of approximately 200 kb. Two of the three new genes lack charged residues in their transmembrane domain tails. Further, one of the genes contains two potential immunotyrosine Inhibitory motifs in its cytoplasmic tail, suggesting that it delivers inhibitory signals. The human and mouse TREM clusters appear to have diverged such that there are unique sequences in each species. Finally, each gene in the TREM cluster was expressed in a different range of cell types.

  11. Spatial Distribution of Iron Within the Normal Human Liver Using Dual-Source Dual-Energy CT Imaging.

    Science.gov (United States)

    Abadia, Andres F; Grant, Katharine L; Carey, Kathleen E; Bolch, Wesley E; Morin, Richard L

    2017-11-01

    Explore the potential of dual-source dual-energy (DSDE) computed tomography (CT) to retrospectively analyze the uniformity of iron distribution and establish iron concentration ranges and distribution patterns found in healthy livers. Ten mixtures consisting of an iron nitrate solution and deionized water were prepared in test tubes and scanned using a DSDE 128-slice CT system. Iron images were derived from a 3-material decomposition algorithm (optimized for the quantification of iron). A conversion factor (mg Fe/mL per Hounsfield unit) was calculated from this phantom study as the quotient of known tube concentrations and their corresponding CT values. Retrospective analysis was performed of patients who had undergone DSDE imaging for renal stones. Thirty-seven patients with normal liver function were randomly selected (mean age, 52.5 years). The examinations were processed for iron concentration. Multiple regions of interest were analyzed, and iron concentration (mg Fe/mL) and distribution was reported. The mean conversion factor obtained from the phantom study was 0.15 mg Fe/mL per Hounsfield unit. Whole-liver mean iron concentrations yielded a range of 0.0 to 2.91 mg Fe/mL, with 94.6% (35/37) of the patients exhibiting mean concentrations below 1.0 mg Fe/mL. The most important finding was that iron concentration was not uniform and patients exhibited regionally high concentrations (36/37). These regions of higher concentration were observed to be dominant in the middle-to-upper part of the liver (75%), medially (72.2%), and anteriorly (83.3%). Dual-source dual-energy CT can be used to assess the uniformity of iron distribution in healthy subjects. Applying similar techniques to unhealthy livers, future research may focus on the impact of hepatic iron content and distribution for noninvasive assessment in diseased subjects.

  12. The thyroxine-binding site of human apolipoprotein-A-I: Location in the N-terminal domain

    International Nuclear Information System (INIS)

    Benvenga, S.; Cahnmann, H.J.; Robbins, J.

    1991-01-01

    We tested the ability of nine monoclonal antibodies (MAb) against human apolipoprotein-A-I (apoA-I), the 28.3-kDa major apoprotein of high density lipoproteins (HDL), to inhibit its photoaffinity labeling with [125I]T4. Two forms were evaluated: isolated lipid-free apoA-I (Sigma or Calbiochem) and lipid-complexed apoA-I [HDL2, (density, 1.063-1.125 g/ml) and HDL3 (density, 1.125-1.210 g/ml)]. After labeling with 0.5 nM [125I]T4 in the presence of MAb or normal mouse IgG, the products were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and subsequent densitometric quantitation of radioactivity associated with the 28.3-kDa band. Group I MAbs, namely those having epitopes in the N-terminal portion of apoA-I, include MAb 16 (epitopes at residues 1-16), 4 and 14 (residues 1-86), and 18 (residues 98-105); group II includes MAbs 7,10, 15, and 17 (epitopes at residues 87-148); group III includes MAb 9 (residues 149-243). All group I MAbs inhibited [125I]T4 binding to isolated apoA-I with this order of potency: MAb 16 greater than MAb 14 greater than MAb 4 greater than MAb 18. In the case of lipid-associated apoA-I, the pattern of hierarchy was variable, presumably related to the known markedly polydisperse nature of HDL, but a constant feature, in contrast to the case of isolated apoA-I, was that MAb 4 was more potent than MAb 14. Group II MAbs gave less than 3% inhibition in both isolated and lipid-complexed apoA-I. Group III MAb 9 either failed to inhibit or gave 18-27% inhibition (one preparation each of HDL2 and HDL3). We conclude that the T4 site of apoA-I is in the N-terminal domain of apoA-I, closer to the epitope for MAb 16 than to that for MAb 18, and that conformational changes occurring when apoA-I is associated with lipids in the HDL particle alter the spatial relationship between some epitopes and the T4 site

  13. Acyl CoA Binding Domain Containing 3 (ACBD3) Protein in Huntington’s Disease 
Human Skin Fibroblasts

    Czech Academy of Sciences Publication Activity Database

    Kratochvílová, H.; Rodinová, M.; Sládková, J.; Klempíř, J.; Lišková, Irena; Motlík, Jan; Zeman, J.; Hansíková, H.; Tesařová, M.

    2015-01-01

    Roč. 78, Suppl. 2 (2015), s. 34-38 ISSN 1210-7859. [Conference on Animal Models for neurodegenerative Diseases /3./. Liblice, 08.11.2015-10.11.2015] R&D Projects: GA MŠk ED2.1.00/03.0124 Institutional support: RVO:67985904 Keywords : Huntington’s disease * Acyl-CoA binding domain containing 3 protein * human skin fibroblasts Subject RIV: FH - Neurology Impact factor: 0.209, year: 2015

  14. Analysis of the Varicella-Zoster Virus IE62 N-Terminal Acidic Transactivating Domain and Its Interaction with the Human Mediator Complex▿

    OpenAIRE

    Yamamoto, Shinobu; Eletsky, Alexander; Szyperski, Thomas; Hay, John; Ruyechan, William T.

    2009-01-01

    The varicella-zoster virus major transactivator, IE62, contains a potent N-terminal acidic transcriptional activation domain (TAD). Our experiments revealed that the minimal IE62 TAD encompasses amino acids (aa) 19 to 67. We showed that the minimal TAD interacts with the human Mediator complex. Site-specific mutations revealed residues throughout the minimal TAD that are important for both activation and Mediator interaction. The TAD interacts directly with aa 402 to 590 of the MED25 subunit,...

  15. Conserved Domain Database (CDD)

    Data.gov (United States)

    U.S. Department of Health & Human Services — CDD is a protein annotation resource that consists of a collection of well-annotated multiple sequence alignment models for ancient domains and full-length proteins.

  16. The combination of analytical and statistical method to define polygonal ship domain and reflect human experiences in estimating dangerous area

    Directory of Open Access Journals (Sweden)

    Gia Huy Dinh

    2016-06-01

    Full Text Available The paper suggests a new method of collision avoidance stemming from the concept of the polygonal target ship domain. Since the last century, we have witnessed the current typical ship domains classified and described. In this proposition, firstly, the domain is a geometrical manner which is used in both analytical and statistical method, resulting in the signification of practical application and simulation. Secondly, such domain will be applied to target ship under the combination of two separated parts: “Blocking area” and “Action area” in order to define the area where the ship must keep outside and how the actions to avoid collision can be generated. Thirdly, the concept has suggested the number of mathematical models for different approaching encounters, including head-on, overtaking and crossing situation. Finally, the parameters of turning circle of the ship can be proposed in determining the size of the domain. Statistical evidences indicate that this method reflects a crew's real habit and psychological in maneuvering. As the result, simple domain is shaped like imagination of sailors, but more accurate in calculating boundary. It promises an effective solution for automatic collision avoidance method. The next researches of this paper have achieved positive results in finding shortest route for avoiding collision. Moreover, while using statistical methods, classical researches face a serious problem in a wide application with different areas, this concept can make up a beneficial solution for the popular application. The numerous ship domains which are in previous researches will be carried out to compare and point out the simplification and effectiveness of the new method in practice.

  17. Epidermal Growth Factor Cytoplasmic Domain Affects ErbB Protein Degradation by the Lysosomal and Ubiquitin-Proteasome Pathway in Human Cancer Cells

    Directory of Open Access Journals (Sweden)

    Aleksandra Glogowska

    2012-05-01

    Full Text Available The cytoplasmic domains of EGF-like ligands, including EGF cytoplasmic domain (EGFcyt, have important biological functions. Using specific constructs and peptides of human EGF cytoplasmic domain, we demonstrate that EGFcyt facilitates lysosomal and proteasomal protein degradation, and this coincided with growth inhibition of human thyroid and glioma carcinoma cells. EGFcyt and exon 22–23-encoded peptide (EGF22.23 enhanced procathepsin B (procathB expression and procathB-mediated lysosomal degradation of EGFR/ErbB1 as determined by inhibitors for procathB and the lysosomal ATPase inhibitor BafA1. Presence of mbEGFctF, EGFcyt, EGF22.23, and exon 23-encoded peptides suppressed the expression of the deubiqitinating enzyme ubiquitin C-terminal hydrolase-L1 (UCH-L1. This coincided with hyperubiquitination of total cellular proteins and ErbB1/2 and reduced proteasome activity. Upon small interfering RNA-mediated silencing of endogenously expressed UCH-L1, a similar hyperubiquitinylation phenotype, reduced ErbB1/2 content, and attenuated growth was observed. The exon 23-encoded peptide region of EGFcyt was important for these biologic actions. Structural homology modeling of human EGFcyt showed that this molecular region formed an exposed surface loop. Peptides derived from this EGFcyt loop structure may aid in the design of novel peptide therapeutics aimed at inhibiting growth of cancer cells.

  18. Estimating the whole bone-marrow asset in humans by a computational approach to integrated PET/CT imaging

    Energy Technology Data Exchange (ETDEWEB)

    Sambuceti, Gianmario [University of Genoa, Nuclear Medicine, Department of Internal Medicine, Genova (Italy); CNR Institute of Bioimages and Molecular Physiology, Genova (Italy); Advanced Biotechnology Center, Genova (Italy); Brignone, Massimo [University of Genoa, Nuclear Medicine, Department of Internal Medicine, Genova (Italy); University of Genoa, Department of Mathematics, Genoa (Italy); Marini, Cecilia [CNR Institute of Bioimages and Molecular Physiology, Genova (Italy); Massollo, Michela; Fiz, Francesco; Morbelli, Silvia; Buschiazzo, Ambra; Piva, Roberta [University of Genoa, Nuclear Medicine, Department of Internal Medicine, Genova (Italy); Campi, Cristina [University of Helsinki, Department of Computer Science, Helsinki (Finland); Massone, Anna Maria [CNR-SPIN, Genova (Italy); Piana, Michele [University of Genoa, Department of Mathematics, Genoa (Italy); CNR-SPIN, Genova (Italy); Frassoni, Francesco [Istituto Giannina Gaslini, Genoa (Italy); Advanced Biotechnology Center, Genova (Italy)

    2012-08-15

    Despite their relevance in clinical medicine, the extension and activity of the bone marrow (BM) cannot be directly evaluated in vivo. We propose a new method to estimate these variables by combining structural and functional maps provided by CT and PET. BM extension and glucose uptake were estimated in 102 patients undergoing whole-body PET/CT because of a history of nonmetastatic melanoma. Image analysis assumed that the BM is surrounded by compact bone. An iterative optimization scheme was applied to each CT slice to identify the external border of the bone. To identify compact bone, the algorithm measured the average Hounsfield coefficient within a two-pixel ring located just inside the bone contour. All intraosseous pixels with an attenuation coefficient lower than this cut-off were flagged as 1, while the remaining pixels were set at 0. Binary masks created from all CT slices were thus applied to the PET data to determine the metabolic activity of the intraosseous volume (IBV). Estimated whole-body IBV was 1,632 {+-} 587 cm{sup 3} and was higher in men than in women (2,004 {+-} 498 cm{sup 3} vs. 1,203 {+-} 354 cm{sup 3}, P < 0.001). Overall, it was strictly correlated with ideal body weight (r = 0.81, P = 0.001) but only loosely with measured body weight (r = 0.43, P = 0.01). The average FDG standardized uptake value (SUV) in the thoracic and lumbar vertebrae was 2.01 {+-} 0.36, Accordingly, intraosseous voxels with SUV {>=}1.11 (mean spine SUV - 2.5 x SD) were considered as active ''red'' BM and those with SUV <1.11 as ''yellow'' BM. Estimated red BM volume was 541 {+-} 195 ml, with a higher prevalence in the axial than in the appendicular skeleton (87 {+-} 8 % vs. 10 {+-} 8 %, P < 0.001). Again, red BM volume was higher in men than in women (7.8 {+-} 2.2 vs. 6.7 {+-} 2.1 ml/kg body weight, P < 0.05), but in women it occupied a greater fraction of the IBV (32 {+-} 7 % vs. 36 {+-} 10 %, P < 0.05). Patient age modestly

  19. Mitochondrial dysfunction enhances cisplatin resistance in human gastric cancer cells via the ROS-activated GCN2-eIF2α-ATF4-xCT pathway.

    Science.gov (United States)

    Wang, Sheng-Fan; Chen, Meng-Shian; Chou, Yueh-Ching; Ueng, Yune-Fang; Yin, Pen-Hui; Yeh, Tien-Shun; Lee, Hsin-Chen

    2016-11-08

    Mitochondrial DNA mutations and defects in mitochondrial enzymes have been identified in gastric cancers, and they might contribute to cancer progression. In previous studies, mitochondrial dysfunction was induced by oligomycin-enhanced chemoresistance to cisplatin. Herein, we dissected the regulatory mechanism for mitochondrial dysfunction-enhanced cisplatin resistance in human gastric cancer cells. Repeated cisplatin treatment-induced cisplatin-resistant cells exhibited high SLC7A11 (xCT) expression, and xCT inhibitors (sulfasalazine or erastin), xCT siRNA, or a GSH synthesis inhibitor (buthionine sulphoximine, BSO) could sensitize these cells to cisplatin. Clinically, the high expression of xCT was associated with a poorer prognosis for gastric cancer patients under adjuvant chemotherapy. Moreover, we found that mitochondrial dysfunction enhanced cisplatin resistance and up-regulated xCT expression, as well as intracellular glutathione (GSH). The xCT inhibitors, siRNA against xCT or BSO decreased mitochondrial dysfunction-enhanced cisplatin resistance. We further demonstrated that the upregulation of the eIF2α-ATF4 pathway contributed to mitochondrial dysfunction-induced xCT expression, and activated eIF2α kinase GCN2, but not PERK, stimulated the eIF2α-ATF4-xCT pathway in response to mitochondrial dysfunction-increased reactive oxygen species (ROS) levels. In conclusion, our results suggested that the ROS-activated GCN2-eIF2α-ATF4-xCT pathway might contribute to mitochondrial dysfunction-enhanced cisplatin resistance and could be a potential target for gastric cancer therapy.

  20. Gene transfer and expression in human neutrophils. The phox homology domain of p47phox translocates to the plasma membrane but not to the membrane of mature phagosomes

    Directory of Open Access Journals (Sweden)

    Brzezinska Agnieszka A

    2006-12-01

    Full Text Available Abstract Background Neutrophils are non-dividing cells with poor survival after isolation. Consequently, exogenous gene expression in neutrophils is challenging. We report here the transfection of genes and expression of active proteins in human primary peripheral neutrophils using nucleofection. Results Exogenous gene expression in human neutrophils was achieved 2 h post-transfection. We show that neutrophils transfected by nucleofection are functional cells, able to respond to soluble and particulate stimuli. They conserved the ability to undergo physiological processes including phagocytosis. Using this technique, we were able to show that the phox homology (PX domain of p47phox localizes to the plasma membrane in human neutrophils. We also show that RhoB, but not the PX domain of p47phox, is translocated to the membrane of mature phagosomes. Conclusion We demonstrated that cDNA transfer and expression of exogenous protein in human neutrophils is compatible with cell viability and is no longer a limitation for the study of protein function in human neutrophils.

  1. Characterization of the in vitro expressed autoimmune rippling muscle disease immunogenic domain of human titin encoded by TTN exons 248-249

    Energy Technology Data Exchange (ETDEWEB)

    Zelinka, L. [Biomedical Sciences Program, Kent State University, Kent, OH (United States); McCann, S.; Budde, J.; Sethi, S.; Guidos, M.; Giles, R. [Center for Applied Chemical Biology, Department of Biological Sciences, Youngstown State University, One University Plaza, Youngstown, OH 44555 (United States); Walker, G.R., E-mail: grwalker@ysu.edu [Center for Applied Chemical Biology, Department of Biological Sciences, Youngstown State University, One University Plaza, Youngstown, OH 44555 (United States); Biomedical Sciences Program, Kent State University, Kent, OH (United States)

    2011-08-05

    Highlights: {yields} Affinity purification of the autoimmune rippling muscle disease immunogenic domain of titin. {yields} Partial sequence analysis confirms that the peptides is in the I band region of titin. {yields} This region of the human titin shows high degree of homology to mouse titin N2-A. -- Abstract: Autoimmune rippling muscle disease (ARMD) is an autoimmune neuromuscular disease associated with myasthenia gravis (MG). Past studies in our laboratory recognized a very high molecular weight skeletal muscle protein antigen identified by ARMD patient antisera as the titin isoform. These past studies used antisera from ARMD and MG patients as probes to screen a human skeletal muscle cDNA library and several pBluescript clones revealed supporting expression of immunoreactive peptides. This study characterizes the products of subcloning the titin immunoreactive domain into pGEX-3X and the subsequent fusion protein. Sequence analysis of the fusion gene indicates the cloned titin domain (GenBank ID: (EU428784)) is in frame and is derived from a sequence of N2-A spanning the exons 248-250 an area that encodes the fibronectin III domain. PCR and EcoR1 restriction mapping studies have demonstrated that the inserted cDNA is of a size that is predicted by bioinformatics analysis of the subclone. Expression of the fusion protein result in the isolation of a polypeptide of 52 kDa consistent with the predicted inferred amino acid sequence. Immunoblot experiments of the fusion protein, using rippling muscle/myasthenia gravis antisera, demonstrate that only the titin domain is immunoreactive.

  2. Characterization of the in vitro expressed autoimmune rippling muscle disease immunogenic domain of human titin encoded by TTN exons 248-249

    International Nuclear Information System (INIS)

    Zelinka, L.; McCann, S.; Budde, J.; Sethi, S.; Guidos, M.; Giles, R.; Walker, G.R.

    2011-01-01

    Highlights: → Affinity purification of the autoimmune rippling muscle disease immunogenic domain of titin. → Partial sequence analysis confirms that the peptides is in the I band region of titin. → This region of the human titin shows high degree of homology to mouse titin N2-A. -- Abstract: Autoimmune rippling muscle disease (ARMD) is an autoimmune neuromuscular disease associated with myasthenia gravis (MG). Past studies in our laboratory recognized a very high molecular weight skeletal muscle protein antigen identified by ARMD patient antisera as the titin isoform. These past studies used antisera from ARMD and MG patients as probes to screen a human skeletal muscle cDNA library and several pBluescript clones revealed supporting expression of immunoreactive peptides. This study characterizes the products of subcloning the titin immunoreactive domain into pGEX-3X and the subsequent fusion protein. Sequence analysis of the fusion gene indicates the cloned titin domain (GenBank ID: (EU428784)) is in frame and is derived from a sequence of N2-A spanning the exons 248-250 an area that encodes the fibronectin III domain. PCR and EcoR1 restriction mapping studies have demonstrated that the inserted cDNA is of a size that is predicted by bioinformatics analysis of the subclone. Expression of the fusion protein result in the isolation of a polypeptide of 52 kDa consistent with the predicted inferred amino acid sequence. Immunoblot experiments of the fusion protein, using rippling muscle/myasthenia gravis antisera, demonstrate that only the titin domain is immunoreactive.

  3. Concrete domains

    OpenAIRE

    Kahn, G.; Plotkin, G.D.

    1993-01-01

    This paper introduces the theory of a particular kind of computation domains called concrete domains. The purpose of this theory is to find a satisfactory framework for the notions of coroutine computation and sequentiality of evaluation.

  4. Structure and function of the Juxta membrane domain of the human epidermal growth factor receptor by NMR spectroscopy

    International Nuclear Information System (INIS)

    Choowongkomon, Kiattawee; Carlin, Cathleen; Sonnichsen, Frank D.

    2005-10-01

    The epidermal growth factor receptor (EGFR) is a member of the receptor tyrosine kinase family involved in the regulation of cellular proliferation and differentiation. Its juxta membrane domain (JX), the region located between the transmembrane and kinase domains, plays important roles in receptor trafficking since both basolateral sorting in polarized epithelial cells and lysosomal sorting signals are identified in this region. In order to understand the regulation of these signals, we characterized the structural properties of recombinant JX domain in dodecyl phosphocholine detergent (DPC) by nuclear magnetic resonance (NMR) spectroscopy. In DPC micelles, structures derived from NMR data showed three amphipathic, helical segments. Two equivalent average structural models on the surface of micelles were obtained that differ only in the relative orientation between the first and second helices. Our data suggests that the activity of sorting signals may be regulated by their membrane association and restricted accessibility in the intact receptor

  5. Purification, crystallization and preliminary X-ray crystallographic analysis of the ETS domain of human Ergp55 in complex with the cfos promoter DNA sequence

    International Nuclear Information System (INIS)

    Gangwar, Shanti P.; Meena, Sita R.; Saxena, Ajay K.

    2012-01-01

    The ETS domain of human Ergp55 was purified and crystallized in native, complexes with E74, and cfos promoter DNA sequences. The X-ray intensity data set was collected on ETS–cfos promoter DNA complex crystal at 3.1 Å resolution to analyze the structure by molecular replacement technique. The Ergp55 protein belongs to the Ets family of transciption factors. The Ets transcription factors are involved in various developmental processes and the regulation of cancer metabolism. They contain a highly similar DNA-binding domain known as the ETS domain and have diverse functions in oncogenesis and physiology. The Ets transcription factors differ in their DNA-binding preference at the ETS site and the mechanisms by which they target genes are not clearly understood. To understand its DNA-binding mechanism, the ETS domain of Ergp55 was expressed and purified. The ETS domain was crystallized in the native form and in complex forms with DNA sequences from the E74 and cfos promoters. An X-ray diffraction data set was collected from an ETS–cfos DNA complex crystal at a wavelength of 0.9725 Å on the BM14 synchrotron beamline at the ESRF, France. The ETS–cfos DNA complex crystal belonged to space group C222 1 , with four molecules in the asymmetric unit. For structure analysis, initial phases for the ETS–cfos DNA complex were obtained by the molecular-replacement technique with Phaser in the CCP4 suite using the coordinates of Fli-1 protein and cfos DNA as search models. Structure analysis of the ETS–cfos DNA complex may possibly explain the DNA-binding specificity and its mechanism of interaction with the ETS domain of Ergp55

  6. CT pelvimetry

    International Nuclear Information System (INIS)

    Vicens, J.L.; Charles, G.; Le Berre, J.; Flageat, J.

    1989-01-01

    Pelvimetry aims at providing the obstetrician with information on the size and morphology of the female pelvis, thus enabling him to assess the obstetrical prognosis as accurately as possible. Computed tomography (CT) has several advantages, in particular the lower mother and fetus radiation dose. After a brief description of obstetrical anatomy, the usual CT exploration method is presented and its results are considered in terms of measurements, indices and morphological findings. If performed with a very strict exploration procedure, CT pelvimetry is a very reliable, fact and comfortable method of examination. All these advantages, combined with the minimized patient dose, should lead to an increasingly widespread use of this technique [fr

  7. Characterization of trabecular bone plate-rod microarchitecture using multirow detector CT and the tensor scale: Algorithms, validation, and applications to pilot human studies

    Science.gov (United States)

    Saha, Punam K.; Liu, Yinxiao; Chen, Cheng; Jin, Dakai; Letuchy, Elena M.; Xu, Ziyue; Amelon, Ryan E.; Burns, Trudy L.; Torner, James C.; Levy, Steven M.; Calarge, Chadi A.

    2015-01-01

    Purpose: Osteoporosis is a common bone disease associated with increased risk of low-trauma fractures leading to substantial morbidity, mortality, and financial costs. Clinically, osteoporosis is defined by low bone mineral density (BMD); however, increasing evidence suggests that trabecular bone (TB) microarchitectural quality is an important determinant of bone strength and fracture risk. A tensor scale based algorithm for in vivo characterization of TB plate-rod microarchitecture at the distal tibia using multirow detector CT (MD-CT) imaging is presented and its performance and applications are examined. Methods: The tensor scale characterizes individual TB on the continuum between a perfect plate and a perfect rod and computes their orientation using optimal ellipsoidal representation of local structures. The accuracy of the method was evaluated using computer-generated phantom images at a resolution and signal-to-noise ratio achievable in vivo. The robustness of the method was examined in terms of stability across a wide range of voxel sizes, repeat scan reproducibility, and correlation between TB measures derived by imaging human ankle specimens under ex vivo and in vivo conditions. Finally, the application of the method was evaluated in pilot human studies involving healthy young-adult volunteers (age: 19 to 21 yr; 51 females and 46 males) and patients treated with selective serotonin reuptake inhibitors (SSRIs) (age: 19 to 21 yr; six males and six females). Results: An error of (3.2% ± 2.0%) (mean ± SD), computed as deviation from known measures of TB plate-width, was observed for computer-generated phantoms. An intraclass correlation coefficient of 0.95 was observed for tensor scale TB measures in repeat MD-CT scans where the measures were averaged over a small volume of interest of 1.05 mm diameter with limited smoothing effects. The method was found to be highly stable at different voxel sizes with an error of (2.29% ± 1.56%) at an in vivo voxel size

  8. Domain Engineering

    Science.gov (United States)

    Bjørner, Dines

    Before software can be designed we must know its requirements. Before requirements can be expressed we must understand the domain. So it follows, from our dogma, that we must first establish precise descriptions of domains; then, from such descriptions, “derive” at least domain and interface requirements; and from those and machine requirements design the software, or, more generally, the computing systems.

  9. Purification, crystallization and preliminary X-ray crystallographic analysis of the ATPase domain of human TAP in nucleotide-free and ADP-, vanadate- and azide-complexed forms

    International Nuclear Information System (INIS)

    Meena, Sita R.; Gangwar, Shanti P.; Saxena, Ajay K.

    2012-01-01

    The ATPase domain of human TAP in nucleotide free, ADP, vanadate and azide complexed forms were purified and crystallized. The X-ray diffraction data sets were collected for all crystals in the resolution range of 2.8–3.0 Å. The human transporter associated with antigen processing (TAP) protein belongs to the ATP-binding cassette (ABC) transporter superfamily and is formed by the heterodimerization of TAP1 and TAP2 subunits. TAP selectively pumps cytosolic peptides into the lumen of the endoplasmic reticulum in an ATP-dependent manner. The catalytic cycle of the ATPase domain of TAP is not understood at the molecular level. The structures of catalytic intermediates of the ATPase domain of TAP will contribute to the understanding of the chemical mechanism of ATP hydrolysis. In order to understand this mechanism, the ATPase domain of human TAP1 (NBD1) was expressed and purified, crystallized in nucleotide-free and transition-state complex forms and X-ray crystallographic studies were performed. The NBD1 protein was crystallized (i) in the nucleotide-free apo form; (ii) in complex with ADP–Mg 2+ , mimicking the product-bound state; (iii) in complex with vanadate–ADP–Mg 2+ , mimicking the ATP-bound state; and (iv) in complex with azide–ADP–Mg 2+ , also mimicking the ATP-bound state. X-ray diffraction data sets were collected for apo and complexed NBD1 using an in-house X-ray diffraction facility at a wavelength of 1.5418 Å. The apo and complexed NBD1 crystals belonged to the primitive hexagonal space group P6 2 , with one monomer in the asymmetric unit. Here, the crystallization, data collection and preliminary crystallographic analysis of apo and complexed NBD1 are reported

  10. Context-dependent social evaluation in 4.5-month-old human infants: the role of domain-general versus domain-specific processes in the development of social evaluation.

    Science.gov (United States)

    Hamlin, J K

    2014-01-01

    The ability to distinguish friends from foes allows humans to engage in mutually beneficial cooperative acts while avoiding the costs associated with cooperating with the wrong individuals. One way to do so effectively is to observe how unknown individuals behave toward third parties, and to selectively cooperate with those who help others while avoiding those who harm others. Recent research suggests that a preference for prosocial over antisocial individuals emerges by the time that infants are 3 months of age, and by 8 months, but not before, infants evaluate others' actions in context: they prefer those who harm, rather than help, individuals who have previously harmed others. Currently there are at least two reasons for younger infants' failure to show context-dependent social evaluations. First, this failure may reflect fundamental change in infants' social evaluation system over the first year of life, in which infants first prefer helpers in any situation and only later evaluate prosocial and antisocial actors in context. On the other hand, it is possible that this developmental change actually reflects domain-general limitations of younger infants, such as limited memory and processing capacities. To distinguish between these possibilities, 4.5-month-olds in the current studies were habituated, rather than familiarized as in previous work, to one individual helping and another harming a third party, greatly increasing infants' exposure to the characters' actions. Following habituation, 4.5-month-olds displayed context-dependent social preferences, selectively reaching for helpers of prosocial and hinderers of antisocial others. Such results suggest that younger infants' failure to display global social evaluation in previous work reflected domain-general rather than domain-specific limitations.

  11. Context-dependent social evaluation in 4.5-month-old human infants: The role of domain-general versus domain-specific processes in the development of social evaluation

    Directory of Open Access Journals (Sweden)

    J Kiley eHamlin

    2014-06-01

    Full Text Available The ability to distinguish friends from foes allows humans to engage in mutually beneficial cooperative acts while avoiding the costs associated with cooperating with the wrong individuals. One way to do so effectively is to observe how unknown individuals behave toward third parties, and to selectively cooperate with those who help others while avoiding those who harm others. Recent research suggests that a preference for prosocial over antisocial individuals emerges by the time that infants are 3 months of age, and by 8 months, but not before, infants evaluate others’ actions in context: they prefer those who harm, rather than help, individuals who have previously harmed others. Currently there are at least two reasons for younger infants’ failure to show context-dependent social evaluations. First, this failure may reflect fundamental change in infants’ social evaluation system over the first year of life, in which infants first prefer helpers in any situation and only later evaluate prosocial and antisocial actors in context. On the other hand, it is possible that this developmental change actually reflects domain-general limitations of younger infants, such as limited memory and processing capacities. To distinguish between these possibilities, 4.5-month-olds in the current studies were habituated, rather than familiarized as in previous work, to one individual helping and another harming a third party, greatly increasing infants’ exposure to the characters’ actions. Following habituation, 4.5-month-olds displayed context-dependent social preferences, selectively reaching for helpers of prosocial and hinderers of antisocial others. Such results suggest that younger infants’ failure to display global social evaluation in previous work reflected domain-general rather than domain-specific limitations.

  12. NMR Structure and Action on Nicotinic Acetylcholine Receptors of Water-soluble Domain of Human LYNX1

    Czech Academy of Sciences Publication Activity Database

    Lyukmanova, E. N.; Shenkarev, Z. O.; Shulepko, M. A.; Mineev, K. S.; D´Hoedt, D.; Kasheverov, I. E.; Filkin, S. Yu.; Krivolapova, A. P.; Janíčková, Helena; Doležal, Vladimír; Dolgikh, D. A.; Arseniev, A. S.; Bertrand, D.; Tsetlin, V.I.; Kirpichnikov, M. P.

    2011-01-01

    Roč. 286, č. 12 (2011), s. 10618-10627 ISSN 0021-9258 R&D Projects: GA ČR(CZ) GA305/09/0681 Institutional research plan: CEZ:AV0Z50110509 Keywords : NMR structure * nicotinic acetylcholine receptor * water-soluble domain Subject RIV: FH - Neurology Impact factor: 4.773, year: 2011

  13. Structures of the human Pals1 PDZ domain with and without ligand suggest gated access of Crb to the PDZ peptide-binding groove

    Energy Technology Data Exchange (ETDEWEB)

    Ivanova, Marina E.; Fletcher, Georgina C.; O’Reilly, Nicola; Purkiss, Andrew G.; Thompson, Barry J. [Cancer Research UK, 44 Lincoln’s Inn Fields, London WC2A 3LY (United Kingdom); McDonald, Neil Q., E-mail: neil.mcdonald@cancer.org.uk [Cancer Research UK, 44 Lincoln’s Inn Fields, London WC2A 3LY (United Kingdom); Birkbeck College, University of London, Malet Street, London WC1E 7HX (United Kingdom)

    2015-03-01

    This study characterizes the interaction between the carboxy-terminal (ERLI) motif of the essential polarity protein Crb and the Pals1/Stardust PDZ-domain protein. Structures of human Pals1 PDZ with and without a Crb peptide are described, explaining the highly conserved nature of the ERLI motif and revealing a sterically blocked peptide-binding groove in the absence of ligand. Many components of epithelial polarity protein complexes possess PDZ domains that are required for protein interaction and recruitment to the apical plasma membrane. Apical localization of the Crumbs (Crb) transmembrane protein requires a PDZ-mediated interaction with Pals1 (protein-associated with Lin7, Stardust, MPP5), a member of the p55 family of membrane-associated guanylate kinases (MAGUKs). This study describes the molecular interaction between the Crb carboxy-terminal motif (ERLI), which is required for Drosophila cell polarity, and the Pals1 PDZ domain using crystallography and fluorescence polarization. Only the last four Crb residues contribute to Pals1 PDZ-domain binding affinity, with specificity contributed by conserved charged interactions. Comparison of the Crb-bound Pals1 PDZ structure with an apo Pals1 structure reveals a key Phe side chain that gates access to the PDZ peptide-binding groove. Removal of this side chain enhances the binding affinity by more than fivefold, suggesting that access of Crb to Pals1 may be regulated by intradomain contacts or by protein–protein interaction.

  14. Novel human mutation and CRISPR/Cas genome-edited mice reveal the importance of C-terminal domain of MSX1 in tooth and palate development.

    Science.gov (United States)

    Mitsui, Silvia Naomi; Yasue, Akihiro; Masuda, Kiyoshi; Naruto, Takuya; Minegishi, Yoshiyuki; Oyadomari, Seiichi; Noji, Sumihare; Imoto, Issei; Tanaka, Eiji

    2016-12-05

    Several mutations, located mainly in the MSX1 homeodomain, have been identified in non-syndromic tooth agenesis predominantly affecting premolars and third molars. We identified a novel frameshift mutation of the highly conserved C-terminal domain of MSX1, known as Msx homology domain 6 (MH6), in a Japanese family with non-syndromic tooth agenesis. To investigate the importance of MH6 in tooth development, Msx1 was targeted in mice with CRISPR/Cas system. Although heterozygous MH6 disruption did not alter craniofacial development, homozygous mice exhibited agenesis of lower incisors with or without cleft palate at E16.5. In addition, agenesis of the upper third molars and the lower second and third molars were observed in 4-week-old mutant mice. Although the upper second molars were present, they were abnormally small. These results suggest that the C-terminal domain of MSX1 is important for tooth and palate development, and demonstrate that that CRISPR/Cas system can be used as a tool to assess causality of human disorders in vivo and to study the importance of conserved domains in genes.

  15. Human I-mfa domain proteins specifically interact with KSHV LANA and affect its regulation of Wnt signaling-dependent transcription

    Energy Technology Data Exchange (ETDEWEB)

    Kusano, Shuichi, E-mail: skusano@m2.kufm.kagoshima-u.ac.jp [Division of Persistent and Oncogenic Viruses, Center for Chronic Viral Diseases, Graduate School of Medical and Dental Sciences, Kagoshima University, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan); Eizuru, Yoshito [Division of Persistent and Oncogenic Viruses, Center for Chronic Viral Diseases, Graduate School of Medical and Dental Sciences, Kagoshima University, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan)

    2010-06-04

    Kaposi's sarcoma-associated herpes virus (KSHV)-encoded latency-associated nuclear antigen (LANA) protein has been reported to interact with glycogen synthase kinase 3{beta} (GSK-3{beta}) and to negatively regulate its activity, leading to stimulation of GSK-3{beta}-dependent {beta}-catenin degradation. We show here that the I-mfa domain proteins, HIC (human I-mfa domain-containing protein) and I-mfa (inhibitor of MyoD family a), interacted in vivo with LANA through their C-terminal I-mfa domains. This interaction affected the intracellular localization of HIC, inhibited the LANA-dependent transactivation of a {beta}-catenin-regulated reporter construct, and decreased the level of the LANA.GSK-3{beta} complex. These data reveal for the first time that I-mfa domain proteins interact with LANA and negatively regulate LANA-mediated activation of Wnt signaling-dependent transcription by inhibiting the formation of the LANA.GSK-3{beta} complex.

  16. CT Scan

    Science.gov (United States)

    ... disease, lung nodules and liver masses Monitor the effectiveness of certain treatments, such as cancer treatment Detect ... scan done in a hospital or an outpatient facility. CT scans are painless and, with newer machines, ...

  17. Thoracic CT

    Science.gov (United States)

    ... through a vein (IV) in your hand or forearm. It may be given through the rectum using ... CT scan Vertebra, thoracic (mid back) Normal lung anatomy Thoracic organs References Gotway MB, Panse PM, Gruden ...

  18. CT Enterography

    Science.gov (United States)

    ... a CT scan can be reformatted in multiple planes, and can even generate three-dimensional images. These ... other medical conditions, and if you have a history of heart disease, asthma, diabetes, kidney disease or ...

  19. Humane metapneumovirus (HMPV) associated pulmonary infections in immunocompromised adults—Initial CT findings, disease course and comparison to respiratory-syncytial-virus (RSV) induced pulmonary infections

    International Nuclear Information System (INIS)

    Syha, R.; Beck, R.; Hetzel, J.; Ketelsen, D.; Grosse, U.; Springer, F.; Horger, M.

    2012-01-01

    Aim: To describe computed tomography (CT)-imaging findings in human metapneumovirus (HMPV)-related pulmonary infection as well as their temporal course and to analyze resemblances/differences to pulmonary infection induced by the closely related respiratory-syncytial-virus (RSV) in immunocompromised patients. Materials and methods: Chest-CT-scans of 10 HMPV PCR-positive patients experiencing pulmonary symptoms were evaluated retrospectively with respect to imaging findings and their distribution and results were then compared with data acquired in 13 patients with RSV pulmonary infection. Subsequently, we analyzed the course of chest-findings in HMPV patients. Results: In HMPV, 8/10 patients showed asymmetric pulmonary findings, whereas 13/13 patients with RSV-pneumonia presented more symmetrical bilateral pulmonary infiltrates. Image analysis yielded in HMPV patients following results: ground-glass-opacity (GGO) (n = 6), parenchymal airspace consolidations (n = 5), ill-defined nodular-like centrilobular opacities (n = 9), bronchial wall thickening (n = 8). In comparison, results in RSV patients were: GGO (n = 10), parenchymal airspace consolidations (n = 9), ill-defined nodular-like centrilobular opacities (n = 10), bronchial wall thickening (n = 4). In the course of the disease, signs of acute HMPV interstitial pneumonia regressed transforming temporarily in part into findings compatible with bronchitis/bronchiolitis. Conclusions: Early chest-CT findings in patients with HMPV-related pulmonary symptoms are compatible with asymmetric acute interstitial pneumonia accompanied by signs of bronchitis; the former transforming with time into bronchitis and bronchiolitis before they resolve. On the contrary, RSV-induced pulmonary infection exhibits mainly symmetric acute interstitial pneumonia.

  20. Humane metapneumovirus (HMPV) associated pulmonary infections in immunocompromised adults—Initial CT findings, disease course and comparison to respiratory-syncytial-virus (RSV) induced pulmonary infections

    Energy Technology Data Exchange (ETDEWEB)

    Syha, R., E-mail: roland.syha@med.uni-tuebingen.de [Department of Diagnostic Radiology, Eberhard-Karls-University, Hoppe-Seyler-Str.3, 72076 Tübingen (Germany); Beck, R. [Institute of Medical Virology, Eberhard-Karls-University, Elfriede-Authorn-Str. 6, 72076 Tübingen (Germany); Hetzel, J. [Department of Medical Oncology and Hematology, Eberhard-Karls-University, Otfried-Müller-Str. 10, 72070 Tübingen (Germany); Ketelsen, D.; Grosse, U.; Springer, F.; Horger, M. [Department of Diagnostic Radiology, Eberhard-Karls-University, Hoppe-Seyler-Str.3, 72076 Tübingen (Germany)

    2012-12-15

    Aim: To describe computed tomography (CT)-imaging findings in human metapneumovirus (HMPV)-related pulmonary infection as well as their temporal course and to analyze resemblances/differences to pulmonary infection induced by the closely related respiratory-syncytial-virus (RSV) in immunocompromised patients. Materials and methods: Chest-CT-scans of 10 HMPV PCR-positive patients experiencing pulmonary symptoms were evaluated retrospectively with respect to imaging findings and their distribution and results were then compared with data acquired in 13 patients with RSV pulmonary infection. Subsequently, we analyzed the course of chest-findings in HMPV patients. Results: In HMPV, 8/10 patients showed asymmetric pulmonary findings, whereas 13/13 patients with RSV-pneumonia presented more symmetrical bilateral pulmonary infiltrates. Image analysis yielded in HMPV patients following results: ground-glass-opacity (GGO) (n = 6), parenchymal airspace consolidations (n = 5), ill-defined nodular-like centrilobular opacities (n = 9), bronchial wall thickening (n = 8). In comparison, results in RSV patients were: GGO (n = 10), parenchymal airspace consolidations (n = 9), ill-defined nodular-like centrilobular opacities (n = 10), bronchial wall thickening (n = 4). In the course of the disease, signs of acute HMPV interstitial pneumonia regressed transforming temporarily in part into findings compatible with bronchitis/bronchiolitis. Conclusions: Early chest-CT findings in patients with HMPV-related pulmonary symptoms are compatible with asymmetric acute interstitial pneumonia accompanied by signs of bronchitis; the former transforming with time into bronchitis and bronchiolitis before they resolve. On the contrary, RSV-induced pulmonary infection exhibits mainly symmetric acute interstitial pneumonia.

  1. Kinetics of intravenous radiographic contrast medium injections as used on CT: simulation with time delay differential equations in a basic human cardiovascular multicompartment model.

    Science.gov (United States)

    Violon, D

    2012-12-01

    To develop a multicompartment model of only essential human body components that predicts the contrast medium concentration vs time curve in a chosen compartment after an intravenous injection. Also to show that the model can be used to time adequately contrast-enhanced CT series. A system of linked time delay instead of ordinary differential equations described the model and was solved with a Matlab program (Matlab v. 6.5; The Mathworks, Inc., Natick, MA). All the injection and physiological parameters were modified to cope with normal or pathological situations. In vivo time-concentration curves from the literature were recalculated to validate the model. The recalculated contrast medium time-concentration curves and parameters are given. The results of the statistical analysis of the study findings are expressed as the median prediction error and the median absolute prediction error values for both the time delay and ordinary differential equation systems; these are situated well below the generally accepted maximum 20% limit. The presented program correctly predicts the time-concentration curve of an intravenous contrast medium injection and, consequently, allows an individually tailored approach of CT examinations with optimised use of the injected contrast medium volume, as long as time delay instead of ordinary differential equations are used. The presented program offers good preliminary knowledge of the time-contrast medium concentration curve after any intravenous injection, allowing adequate timing of a CT examination, required by the short scan time of present-day scanners. The injected volume of contrast medium can be tailored to the individual patient with no more contrast medium than is strictly needed.

  2. Identification of a domain within human TAF(I)48, a subunit of Selectivity Factor 1, that interacts with helix 2 of TBP.

    Science.gov (United States)

    Xu, Shuping; Hori, Roderick T

    2004-09-01

    RNA polymerase I transcription in human cells requires Selectivity Factor 1, a multisubunit complex composed of the TATA-box-binding protein (TBP) and three TBP-associated factors (TAFs) called TAF(I)48, TAF(I)63 and TAF(I)110. Each of the Selectivity Factor 1 subunits binds directly to the other three components, but these interactions have not been characterized. This study is the initial identification and analysis of a TBP-binding domain within a Selectivity Factor 1 TAF. The interaction between human TBP and human TAF(I)48 was initially examined using the yeast two-hybrid assay, and a TBP-binding domain was identified in the carboxyl-terminus of human (h)TAF(I)48. Consistent with this result, the hTAF(I)48 carboxyl-terminus was able to bind directly to TBP in protein-protein interaction assays. When mutations were introduced into the hTAF(I)48 carboxyl-terminus, we identified changes in uncharged and positive residues that affect its interaction with TBP. By examining TBP mutants, residues within and adjacent to helix 2 of TBP, previously demonstrated to interact with subunits of other TBP-containing complexes [Transcription Factor IID (TFIID) and TFIIIB] were also found to diminish its affinity for the carboxyl-terminus of hTAF(I)48. The regions of hTAF(I)48 and TBP that interact are compared to those identified within other complexes containing TBP.

  3. Normal cranial CT anatomy

    International Nuclear Information System (INIS)

    Gado, M.H.; Rao, K.C.V.G.

    1987-01-01

    The human brain consists of well-known anatomical components. Some parts of these components have been shown to be concerned with certain functions. A complete cranial CT examination consists of a series of several slices obtained in a sequence usually from the base to the vertex of the cranial vault, in the axial mode. The ultimate goal of this chapter is to pinpoint those slices that depict a given anatomical structure or several structures that deal with a given function. To achieve this goal, the discussion of CT cranial anatomy is presented in three sections

  4. Classification of micro-CT images using 3D characterization of bone canal patterns in human osteogenesis imperfecta

    Science.gov (United States)

    Abidin, Anas Z.; Jameson, John; Molthen, Robert; Wismüller, Axel

    2017-03-01

    Few studies have analyzed the microstructural properties of bone in cases of Osteogenenis Imperfecta (OI), or `brittle bone disease'. Current approaches mainly focus on bone mineral density measurements as an indirect indicator of bone strength and quality. It has been shown that bone strength would depend not only on composition but also structural organization. This study aims to characterize 3D structure of the cortical bone in high-resolution micro CT images. A total of 40 bone fragments from 28 subjects (13 with OI and 15 healthy controls) were imaged using micro tomography using a synchrotron light source (SRµCT). Minkowski functionals - volume, surface, curvature, and Euler characteristics - describing the topological organization of the bone were computed from the images. The features were used in a machine learning task to classify between healthy and OI bone. The best classification performance (mean AUC - 0.96) was achieved with a combined 4-dimensional feature of all Minkowski functionals. Individually, the best feature performance was seen using curvature (mean AUC - 0.85), which characterizes the edges within a binary object. These results show that quantitative analysis of cortical bone microstructure, in a computer-aided diagnostics framework, can be used to distinguish between healthy and OI bone with high accuracy.

  5. Transient Dynamics Simulation of Airflow in a CT-Scanned Human Airway Tree: More or Fewer Terminal Bronchi?

    Directory of Open Access Journals (Sweden)

    Shouliang Qi

    2017-01-01

    Full Text Available Using computational fluid dynamics (CFD method, the feasibility of simulating transient airflow in a CT-based airway tree with more than 100 outlets for a whole respiratory period is studied, and the influence of truncations of terminal bronchi on CFD characteristics is investigated. After an airway model with 122 outlets is extracted from CT images, the transient airflow is simulated. Spatial and temporal variations of flow velocity, wall pressure, and wall shear stress are presented; the flow pattern and lobar distribution of air are gotten as well. All results are compared with those of a truncated model with 22 outlets. It is found that the flow pattern shows lobar heterogeneity that the near-wall air in the trachea is inhaled into the upper lobe while the center flow enters the other lobes, and the lobar distribution of air is significantly correlated with the outlet area ratio. The truncation decreases airflow to right and left upper lobes and increases the deviation of airflow distributions between inspiration and expiration. Simulating the transient airflow in an airway tree model with 122 bronchi using CFD is feasible. The model with more terminal bronchi decreases the difference between the lobar distributions at inspiration and at expiration.

  6. Interaction of CtBP with adenovirus E1A suppresses immortalization of primary epithelial cells and enhances virus replication during productive infection.

    Science.gov (United States)

    Subramanian, T; Zhao, Ling-Jun; Chinnadurai, G

    2013-09-01

    Adenovirus E1A induces cell proliferation, oncogenic transformation and promotes viral replication through interaction with p300/CBP, TRRAP/p400 multi-protein complex and the retinoblastoma (pRb) family proteins through distinct domains in the E1A N-terminal region. The C-terminal region of E1A suppresses E1A/Ras co-transformation and interacts with FOXK1/K2, DYRK1A/1B/HAN11 and CtBP1/2 (CtBP) protein complexes. To specifically dissect the role of CtBP interaction with E1A, we engineered a mutation (DL→AS) within the CtBP-binding motif, PLDLS, and investigated the effect of the mutation on immortalization and Ras cooperative transformation of primary cells and viral replication. Our results suggest that CtBP-E1A interaction suppresses immortalization and Ras co-operative transformation of primary rodent epithelial cells without significantly influencing the tumorigenic activities of transformed cells in immunodeficient and immunocompetent animals. During productive infection, CtBP-E1A interaction enhances viral replication in human cells. Between the two CtBP family proteins, CtBP2 appears to restrict viral replication more than CtBP1 in human cells. Copyright © 2013 Elsevier Inc. All rights reserved.

  7. L-arginine mediated renaturation enhances yield of human, α6 Type IV collagen non-collagenous domain from bacterial inclusion bodies.

    Science.gov (United States)

    Gunda, Venugopal; Boosani, Chandra Shekhar; Verma, Raj Kumar; Guda, Chittibabu; Sudhakar, Yakkanti Akul

    2012-10-01

    The anti-angiogenic, carboxy terminal non-collagenous domain (NC1) derived from human Collagen type IV alpha 6 chain, [α6(IV)NC1] or hexastatin, was earlier obtained using different recombinant methods of expression in bacterial systems. However, the effect of L-arginine mediated renaturation in enhancing the relative yields of this protein from bacterial inclusion bodies has not been evaluated. In the present study, direct stirring and on-column renaturation methods using L-arginine and different size exclusion chromatography matrices were applied for enhancing the solubility in purifying the recombinant α6(IV)NC1 from bacterial inclusion bodies. This methodology enabled purification of higher quantities of soluble protein from inclusion bodies, which inhibited endothelial cell proliferation, migration and tube formation. Thus, the scope for L-arginine mediated renaturation in obtaining higher yields of soluble, biologically active NC1 domain from bacterial inclusion bodies was evaluated.

  8. Transformation and radiosensitivity of human diploid skin fibroblasts transfected with SV40 T-antigen mutants defective in RB and P53 binding domains

    International Nuclear Information System (INIS)

    LingNah Su; Little, J.B.

    1992-01-01

    A series of human diploid fibroblast cell clones were developed by DNA transfection with either wild-type SV40 T-antigen (SV40T) or T-antigen mutants defective in its various functional domains. Cell clones expressing the wild-type SV40 T were significantly radioresistant as compared with clones transfected with the neo gene only (D o 192 ± 13 vs 127 ± 19). This radioresistance persisted in post-crisis, immortalized cell lines. A series of mutants with point or deletion mutations within each functionally active domain of SV40 T were also examined for their ability to alter radiosensitivity and induce morphological transformation. Cell clones transfected with T-antigen mutants defective in nuclear localization or origin binding showed increased radioresistance similar to clones transfected with wild-type T-antigen, and expressed morphological changes characteristic of SV40 T-transfected cells. (author)

  9. Non-destructive, preclinical evaluation of root canal anatomy of human teeth with flat-panel detector volume CT (FD-VCT); Zerstoerungsfreie praeklinische Evaluation der Wurzelkanalanatomie menschlicher Zaehne mittels Flaechendetektor-Volumen-CT (FD-VCT)

    Energy Technology Data Exchange (ETDEWEB)

    Heidrich, G.; Hassepass, F.; Dullin, C.; Grabbe, E. [Universitaetsklinikum Goettingen, Abt. Diagnostische Radiologie (Germany); Attin, T.; Hannig, C. [Universitaetsklinikum Goettingen, Abt. fuer Zahnerhaltung, Praeventive Zahnheilkunde und Paradontologie (Germany)

    2005-12-15

    Purpose: Successful endodontic diagnostics and therapy call for adequate depiction of the root canal anatomy with multimodal diagnostic imaging. The aim of the present study is to evaluate visualization of the endodont with flat-panel detector volume CT (FD-VCT). Materials and methods: 13 human teeth were examined with the prototype of a FD-VCT. After data acquisition and generation of volume data sets in volume rendering technology (VRT), the findings obtained were compared to conventional X-rays and cross-section preparations of the teeth. Results: The anatomical structures of the endodont such as root canals, side canals and communications between different root canals as well as dentricles could be detected precisely with FD-VCT. The length of curved root canals was also determined accurately. The spatial resolution of the system is around 140 {mu}m. Only around 73% of the main root canals detected with FD-VCT and 87% of the roots could be visualized with conventional dental X-rays. None of the side canals, shown with FD-VCT, was detectable on conventional X-rays. In all cases the enamel and dentin of the teeth could be well delineated. No differences in image quality could be discerned between stored and freshly extracted teeth, or between primary and adult teeth. (orig.)

  10. Heteronuclear multidimensional NMR and homology modelling studies of the C-terminal nucleotide-binding domain of the human mitochondrial ABC transporter ABCB6

    Energy Technology Data Exchange (ETDEWEB)

    Kurashima-Ito, Kaori [RIKEN, Cellular and Molecular Biology Laboratory (Japan); Ikeya, Teppei [National Institute of Advanced Industrial Science and Technology (AIST), (Japan); Senbongi, Hiroshi [Mitochondrial Diseases Group, MRC Dunn Human NutritionUnit (United Kingdom); Tochio, Hidehito [International Graduate School of Arts and Sciences, Supramolecular Biology, Yokohama City University, Molecular Biophysics Laboratory (Japan); Mikawa, Tsutomu [RIKEN, Cellular and Molecular Biology Laboratory (Japan); Shibata, Takehiko [RIKEN, Shibata Distinguished Senior Scientist Laboratory (Japan); Ito, Yutaka [RIKEN, Cellular and Molecular Biology Laboratory (Japan)], E-mail: ito-yutaka@center.tmu.ac.jp

    2006-05-15

    Human ATP-binding cassette, sub-family B, member 6 (ABCB6) is a mitochondrial ABC transporter, and presumably contributes to iron homeostasis. Aimed at understanding the structural basis for the conformational changes accompanying the substrate-transportation cycle, we have studied the C-terminal nucleotide-binding domain of ABCB6 (ABCB6-C) in both the nucleotide-free and ADP-bound states by heteronuclear multidimensional NMR and homology modelling. A non-linear sampling scheme was utilised for indirectly acquired {sup 13}C and {sup 15}N dimensions of all 3D triple-resonance NMR experiments, in order to overcome the instability and the low solubility of ABCB6-C. The backbone resonances for approximately 25% of non-proline residues, which are mostly distributed around the functionally important loops and in the Helical domain, were not observed for nucleotide-free form of ABCB6-C. From the pH, temperature and magnetic field strength dependencies of the resonance intensities, we concluded that this incompleteness in the assignments is mainly due to the exchange between multiple conformations at an intermediate rate on the NMR timescale. These localised conformational dynamics remained in ADP-bound ABCB6-C except for the loops responsible for adenine base and {alpha}/{beta}-phosphate binding. These results revealed that the localised dynamic cooperativity, which was recently proposed for a prokaryotic ABC MJ1267, also exists in a higher eukaryotic ABC, and is presumably shared by all members of the ABC family. Since the Helical domain is the putative interface to the transmembrane domain, this cooperativity may explain the coupled functions between domains in the substrate-transportation cycle.

  11. Crystal-contact engineering to obtain a crystal form of the Kelch domain of human Keap1 suitable for ligand-soaking experiments

    International Nuclear Information System (INIS)

    Hörer, Stefan; Reinert, Dirk; Ostmann, Katja; Hoevels, Yvette; Nar, Herbert

    2013-01-01

    A mutant of the Kelch domain of the human Keap1 protein has been designed in order to enable the soaking of small-molecule ligands. The apo structure of this mutant is reported at 1.98 Å resolution and the suitability of the crystal system has been demonstrated by the structure of the mutated Keap1 Kelch domain in complex with a cyclic peptide derived from Nrf2. Keap1 is a substrate adaptor protein for a Cul3-dependent ubiquitin ligase complex and plays an important role in the cellular response to oxidative stress. It binds Nrf2 with its Kelch domain and thus triggers the ubiquitinylation and degradation of Nrf2. Oxidative stress prevents the degradation of Nrf2 and leads to the activation of cytoprotective genes. Therefore, Keap1 is an attractive drug target in inflammatory diseases. The support of a medicinal chemistry effort by structural research requires a robust crystallization system in which the crystals are preferably suited for performing soaking experiments. This facilitates the generation of protein–ligand complexes in a routine and high-throughput manner. The structure of human Keap1 has been described previously. In this crystal form, however, the binding site for Nrf2 was blocked by a crystal contact. This interaction was analysed and mutations were introduced to disrupt this crystal contact. One double mutation (E540A/E542A) crystallized in a new crystal form in which the binding site for Nrf2 was not blocked and was accessible to small-molecule ligands. The crystal structures of the apo form of the mutated Keap1 Kelch domain (1.98 Å resolution) and of the complex with an Nrf2-derived peptide obtained by soaking (2.20 Å resolution) are reported

  12. Imaging the Aqueous Humor Outflow Pathway in Human Eyes by Three-dimensional Micro-computed Tomography (3D micro-CT)

    Energy Technology Data Exchange (ETDEWEB)

    C Hann; M Bentley; A Vercnocke; E Ritman; M Fautsch

    2011-12-31

    The site of outflow resistance leading to elevated intraocular pressure in primary open-angle glaucoma is believed to be located in the region of Schlemm's canal inner wall endothelium, its basement membrane and the adjacent juxtacanalicular tissue. Evidence also suggests collector channels and intrascleral vessels may have a role in intraocular pressure in both normal and glaucoma eyes. Traditional imaging modalities limit the ability to view both proximal and distal portions of the trabecular outflow pathway as a single unit. In this study, we examined the effectiveness of three-dimensional micro-computed tomography (3D micro-CT) as a potential method to view the trabecular outflow pathway. Two normal human eyes were used: one immersion fixed in 4% paraformaldehyde and one with anterior chamber perfusion at 10 mmHg followed by perfusion fixation in 4% paraformaldehyde/2% glutaraldehyde. Both eyes were postfixed in 1% osmium tetroxide and scanned with 3D micro-CT at 2 {mu}m or 5 {mu}m voxel resolution. In the immersion fixed eye, 24 collector channels were identified with an average orifice size of 27.5 {+-} 5 {mu}m. In comparison, the perfusion fixed eye had 29 collector channels with a mean orifice size of 40.5 {+-} 13 {mu}m. Collector channels were not evenly dispersed around the circumference of the eye. There was no significant difference in the length of Schlemm's canal in the immersed versus the perfused eye (33.2 versus 35.1 mm). Structures, locations and size measurements identified by 3D micro-CT were confirmed by correlative light microscopy. These findings confirm 3D micro-CT can be used effectively for the non-invasive examination of the trabecular meshwork, Schlemm's canal, collector channels and intrascleral vasculature that comprise the distal outflow pathway. This imaging modality will be useful for non-invasive study of the role of the trabecular outflow pathway as a whole unit.

  13. Multi-detector row CT colonography: effect of collimation, pitch, and orientation on polyp detection in a human colectomy specimen.

    Science.gov (United States)

    Taylor, Stuart A; Halligan, Steve; Bartram, Clive I; Morgan, Paul R; Talbot, Ian C; Fry, Nicola; Saunders, Brian P; Khosraviani, Kirosh; Atkin, Wendy

    2003-10-01

    To investigate the effects of orientation, collimation, pitch, and tube current setting on polyp detection at multi-detector row computed tomographic (CT) colonography and to determine the optimal combination of scanning parameters for screening. A colectomy specimen containing 117 polyps of different sizes was insufflated and imaged with a multi-detector row CT scanner at various collimation (1.25 and 2.5 mm), pitch (3 and 6), and tube current (50, 100, and 150 mA) settings. Two-dimensional multiplanar reformatted images and three-dimensional endoluminal surface renderings from the 12 resultant data sets were examined by one observer for the presence and conspicuity of polyps. The results were analyzed with Poisson regression and logistic regression to determine the effects of scanning parameters and of specimen orientation on polyp detection. The percentage of polyps that were detected significantly increased when collimation (P =.008) and table feed (P =.03) were decreased. Increased tube current resulted in improved detection only of polyps with a diameter of less than 5 mm. Polyps of less than 5 mm were optimally depicted with a collimation of 1.25 mm, a pitch of 3, and a tube current setting of 150 mA; polyps with a diameter greater than 5 mm were adequately depicted with 1.25-mm collimation and with either pitch setting and any of the three tube current settings. Small polyps in the transverse segment (positioned at a 90 degrees angle to the z axis of scanning) were significantly less visible than those in parallel or oblique orientations (P detector row CT is highly dependent on collimation, pitch, and, to a lesser extent, tube current. Collimation of 1.25 mm, combined with pitch of 6 and tube current of 50 mA, provides for reliable detection of polyps 5 mm or larger while limiting the effective radiation dose. Polyps smaller than 5 mm, however, may be poorly depicted with use of these settings in the transverse colon. Copyright RSNA, 2003

  14. HER family kinase domain mutations promote tumor progression and can predict response to treatment in human breast cancer

    KAUST Repository

    Boulbes, Delphine R.

    2014-11-11

    Resistance to HER2-targeted therapies remains a major obstacle in the treatment of HER2-overexpressing breast cancer. Understanding the molecular pathways that contribute to the development of drug resistance is needed to improve the clinical utility of novel agents, and to predict the success of targeted personalized therapy based on tumor-specific mutations. Little is known about the clinical significance of HER family mutations in breast cancer. Because mutations within HER1/EGFR are predictive of response to tyrosine kinase inhibitors (TKI) in lung cancer, we investigated whether mutations in HER family kinase domains are predictive of response to targeted therapy in HER2-overexpressing breast cancer. We sequenced the HER family kinase domains from 76 HER2-overexpressing invasive carcinomas and identified 12 missense variants. Patients whose tumors carried any of these mutations did not respond to HER2 directed therapy in the metastatic setting. We developed mutant cell lines and used structural analyses to determine whether changes in protein conformation could explain the lack of response to therapy. We also functionally studied all HER2 mutants and showed that they conferred an aggressive phenotype and altered effects of the TKI lapatinib. Our data demonstrate that mutations in the finely tuned HER kinase domains play a critical function in breast cancer progression and may serve as prognostic and predictive markers.

  15. HER family kinase domain mutations promote tumor progression and can predict response to treatment in human breast cancer

    KAUST Repository

    Boulbes, Delphine R.; Arold, Stefan T.; Chauhan, Gaurav B.; Blachno, Korina V.; Deng, Nanfu; Chang, Wei-Chao; Jin, Quanri; Huang, Tzu-Hsuan; Hsu, Jung-Mao; Brady, Samuel W.; Bartholomeusz, Chandra; Ladbury, John E.; Stone, Steve; Yu, Dihua; Hung, Mien-Chie; Esteva, Francisco J.

    2014-01-01

    Resistance to HER2-targeted therapies remains a major obstacle in the treatment of HER2-overexpressing breast cancer. Understanding the molecular pathways that contribute to the development of drug resistance is needed to improve the clinical utility of novel agents, and to predict the success of targeted personalized therapy based on tumor-specific mutations. Little is known about the clinical significance of HER family mutations in breast cancer. Because mutations within HER1/EGFR are predictive of response to tyrosine kinase inhibitors (TKI) in lung cancer, we investigated whether mutations in HER family kinase domains are predictive of response to targeted therapy in HER2-overexpressing breast cancer. We sequenced the HER family kinase domains from 76 HER2-overexpressing invasive carcinomas and identified 12 missense variants. Patients whose tumors carried any of these mutations did not respond to HER2 directed therapy in the metastatic setting. We developed mutant cell lines and used structural analyses to determine whether changes in protein conformation could explain the lack of response to therapy. We also functionally studied all HER2 mutants and showed that they conferred an aggressive phenotype and altered effects of the TKI lapatinib. Our data demonstrate that mutations in the finely tuned HER kinase domains play a critical function in breast cancer progression and may serve as prognostic and predictive markers.

  16. A graph kernel approach for alignment-free domain–peptide interaction prediction with an application to human SH3 domains

    Science.gov (United States)

    Kundu, Kousik; Costa, Fabrizio; Backofen, Rolf

    2013-01-01

    Motivation: State-of-the-art experimental data for determining binding specificities of peptide recognition modules (PRMs) is obtained by high-throughput approaches like peptide arrays. Most prediction tools applicable to this kind of data are based on an initial multiple alignment of the peptide ligands. Building an initial alignment can be error-prone, especially in the case of the proline-rich peptides bound by the SH3 domains. Results: Here, we present a machine-learning approach based on an efficient graph-kernel technique to predict the specificity of a large set of 70 human SH3 domains, which are an important class of PRMs. The graph-kernel strategy allows us to (i) integrate several types of physico-chemical information for each amino acid, (ii) consider high-order correlations between these features and (iii) eliminate the need for an initial peptide alignment. We build specialized models for each human SH3 domain and achieve competitive predictive performance of 0.73 area under precision-recall curve, compared with 0.27 area under precision-recall curve for state-of-the-art methods based on position weight matrices. We show that better models can be obtained when we use information on the noninteracting peptides (negative examples), which is currently not used by the state-of-the art approaches based on position weight matrices. To this end, we analyze two strategies to identify subsets of high confidence negative data. The techniques introduced here are more general and hence can also be used for any other protein domains, which interact with short peptides (i.e. other PRMs). Availability: The program with the predictive models can be found at http://www.bioinf.uni-freiburg.de/Software/SH3PepInt/SH3PepInt.tar.gz. We also provide a genome-wide prediction for all 70 human SH3 domains, which can be found under http://www.bioinf.uni-freiburg.de/Software/SH3PepInt/Genome-Wide-Predictions.tar.gz. Contact: backofen@informatik.uni-freiburg.de Supplementary

  17. Domain structure of human complement C4b extends with increasing NaCl concentration: implications for its regulatory mechanism.

    Science.gov (United States)

    Fung, Ka Wai; Wright, David W; Gor, Jayesh; Swann, Marcus J; Perkins, Stephen J

    2016-12-01

    During the activation of complement C4 to C4b, the exposure of its thioester domain (TED) is crucial for the attachment of C4b to activator surfaces. In the C4b crystal structure, TED forms an Arg 104 -Glu 1032 salt bridge to tether its neighbouring macroglobulin (MG1) domain. Here, we examined the C4b domain structure to test whether this salt bridge affects its conformation. Dual polarisation interferometry of C4b immobilised at a sensor surface showed that the maximum thickness of C4b increased by 0.46 nm with an increase in NaCl concentration from 50 to 175 mM NaCl. Analytical ultracentrifugation showed that the sedimentation coefficient s 20,w of monomeric C4b of 8.41 S in 50 mM NaCl buffer decreased to 7.98 S in 137 mM NaCl buffer, indicating that C4b became more extended. Small angle X-ray scattering reported similar R G values of 4.89-4.90 nm for C4b in 137-250 mM NaCl. Atomistic scattering modelling of the C4b conformation showed that TED and the MG1 domain were separated by 4.7 nm in 137-250 mM NaCl and this is greater than that of 4.0 nm in the C4b crystal structure. Our data reveal that in low NaCl concentrations, both at surfaces and in solution, C4b forms compact TED-MG1 structures. In solution, physiologically relevant NaCl concentrations lead to the separation of the TED and MG1 domain, making C4b less capable of binding to its complement regulators. These conformational changes are similar to those seen previously for complement C3b, confirming the importance of this salt bridge for regulating both C4b and C3b. © 2016 The Author(s).

  18. 64Cu-PSMA-617 PET/CT Imaging of Prostate Adenocarcinoma: First In-Human Studies.

    Science.gov (United States)

    Grubmüller, Bernhard; Baum, Richard P; Capasso, Enza; Singh, Aviral; Ahmadi, Yasaman; Knoll, Peter; Floth, Andreas; Righi, Sergio; Zandieh, Shahin; Meleddu, Carlo; Shariat, Shahrokh F; Klingler, Hans Christoph; Mirzaei, Siroos

    2016-10-07

    The prostate-specific membrane antigen (PSMA) is a cell surface protein, which is overexpressed in nearly all cases of prostate cancer (PCa). PET imaging with 68 Ga-PSMA-HBED-CC has recently found widespread application in the diagnosis of recurrent PCa. In this study, the diagnostic potential of 64 Cu-labeled PSMA ligand (PSMA-617) PET in patients with PCa has been investigated. The study was conducted simultaneously at two nuclear medicine centers, Austria (Vienna, Center 1) and Germany (Bad Berka, Center 2). The patients (n = 29) included in this study were referred for PET (Center 1, 21 patients) or PET/CT (Center 2, 8 patients) imaging with either a high suspicion of recurrent disease or for possible surgical or PSMA radioligand therapy planning. PET images of the whole body were performed at 1 hour p.i. and additional images of the pelvis at 2 hours p.i. In 23 of 29 patients, at least one focus of pathological tracer uptake suspicious for primary disease in the prostate lobe or recurrent disease was detected. Among healthy organs, the salivary glands, kidneys, and liver showed the highest radiotracer uptake. Lesions suspicious for PCa were detected with excellent contrast as early as 1 hour p.i. with high detection rates even at low prostate-specific antigen (PSA) levels. The preliminary results of this study demonstrate the high potential of 64 Cu-PSMA ligand PET/CT imaging in patients with recurrent disease and in the primary staging of selected patients with progressive local disease. The acquired PET images showed an excellent resolution of the detected lesions with very high lesion-to- background contrast. Furthermore, the long half-life of 64 Cu allows distribution of the tracer to clinical PET centers that lack radiochemistry facilities for the preparation of 68 Ga-PSMA ligand (satellite concept).

  19. "Being an English Major, Being a Humanities Student": Connecting Academic Subject Identity in Literary Studies to Other Social Domains

    Science.gov (United States)

    Chan, Evelyn T. Y.

    2016-01-01

    This study examined students' construction of academic subject identity in a university humanities discipline, English literary studies. In so doing, the study aimed to provide an empirically grounded intervention in current debates on the value of the humanities in higher education. Eight students participated in interviews lasting 15-20 minutes…

  20. PDZ domain-binding motif of human T-cell leukemia virus type 1 Tax oncoprotein augments the transforming activity in a rat fibroblast cell line

    International Nuclear Information System (INIS)

    Hirata, Akira; Higuchi, Masaya; Niinuma, Akiko; Ohashi, Minako; Fukushi, Masaya; Oie, Masayasu; Akiyama, Tetsu; Tanaka, Yuetsu; Gejyo, Fumitake; Fujii, Masahiro

    2004-01-01

    While human T-cell leukemia virus type 1 (HTLV-1) is associated with the development of adult T-cell leukemia (ATL), HTLV-2 has not been reported to be associated with such malignant leukemias. HTLV-1 Tax1 oncoprotein transforms a rat fibroblast cell line (Rat-1) to form multiple large colonies in soft agar, and this activity is much greater than that of HTLV-2 Tax2. We have demonstrated here that the increased number of transformed colonies induced by Tax1 relative to Tax2 was mediated by a PDZ domain-binding motif (PBM) in Tax1, which is absent in Tax2. Tax1 PBM mediated the interaction of Tax1 with the discs large (Dlg) tumor suppressor containing PDZ domains, and the interaction correlated well with the transforming activities of Tax1 and the mutants. Through this interaction, Tax1 altered the subcellular localization of Dlg from the detergent-soluble to the detergent-insoluble fraction in a fibroblast cell line as well as in HTLV-1-infected T-cell lines. These results suggest that the interaction of Tax1 with PDZ domain protein(s) is critically involved in the transforming activity of Tax1, the activity of which may be a crucial factor in malignant transformation of HTLV-1-infected cells in vivo

  1. Overproduction, purification, crystallization and preliminary X-ray analysis of human Fe65-PTB2 in complex with the amyloid precursor protein intracellular domain

    Energy Technology Data Exchange (ETDEWEB)

    Radzimanowski, Jens [Heidelberg University Biochemistry Center, INF328, D-69120 Heidelberg (Germany); Beyreuther, Konrad [Center for Molecular Biology, University Heidelberg, INF282, D-69120 Heidelberg (Germany); Sinning, Irmgard; Wild, Klemens, E-mail: klemens.wild@bzh.uni-heidelberg.de [Heidelberg University Biochemistry Center, INF328, D-69120 Heidelberg (Germany)

    2008-05-01

    Alzheimer’s disease is characterized by proteolytic processing of the amyloid precursor protein (APP), which releases the aggregation-prone amyloid-β (Aβ) peptide and liberates the intracellular domain (AICD) that interacts with various adaptor proteins. The crystallized AICD–Fe65-PTB2 complex is of central importance for APP translocation, nuclear signalling, processing and Aβ generation. Alzheimer’s disease is associated with typical brain deposits (senile plaques) that mainly contain the neurotoxic amyloid β peptide. This peptide results from proteolytic processing of the type I transmembrane protein amyloid precursor protein (APP). During this proteolytic pathway the APP intracellular domain (AICD) is released into the cytosol, where it associates with various adaptor proteins. The interaction of the AICD with the C-terminal phosphotyrosine-binding domain of Fe65 (Fe65-PTB2) regulates APP translocation, signalling and processing. Human AICD and Fe65-PTB2 have been cloned, overproduced and purified in large amounts in Escherichia coli. A complex of Fe65-PTB2 with the C-terminal 32 amino acids of the AICD gave well diffracting hexagonal crystals and data have been collected to 2.1 Å resolution. Initial phases obtained by the molecular-replacement method are of good quality and revealed well defined electron density for the substrate peptide.

  2. The value of CT measurement of normal human mandible, hyoid and the airway area at the hyoid level

    International Nuclear Information System (INIS)

    Chen Changliang; Zhang Jia; Ge Pingjiang; Zhang Siyi

    2004-01-01

    Objective: To determine the normal value and clinical significance of the lengths and angles of both mandible and hyoid, and their relationship with cervical vertebra as well as the transverse area of the airway at the hyoid level using CT. Methods: Several lines and angles on the CT images were measured in 68 normal subjects. Line A was the length between both free ends of the mandible; line B was the distance from the body of the mandible to line A; Line C was the distance from line A to the anterior aspect of the cervical vertebra. α 1 was the angle between the middle of mandible body and its two free ends. Line a was the distance between two free ends of greater horn of hyoid bone. Line b was the distance from hyoid to line a. Line c was the distance from line a to the cervical vertebra. α 2 was the angle between the middle of hyoid body and its two free ends. S stood for the area of the airway at the hyoid level. SPSS 11.5 statistical analysis package was used to analyze the results. Results: The average and median distance/angle of various measurements were as follows: line A was (89.28±5.90) mm and 88.70 mm, line B was (62.61±5.78) mm and 62.50 mm, line C was (9.29±3.29) mm and 4.20 mm, α 1 was (71.25±6.77) degree and 71.05 degree, line a was (38.69 ± 6.07) mm and 39.90 mm, line b was (28.79 ± 4.37) mm and 28.50 mm, line c was (1.91 ± 3.03) mm and 1.75 mm, α 2 was (68.47 ± 15.71) degree and 66.95 degree. The average S was (436.14 ± 160.37) mm 2 and median was 431.75 mm 2 . Conclusion: It is easy to measure the three lines and the two angles of mandible and hyoid. The measurement is of vital importance in the diagnosis and treatment for obstructive sleep apnea syndrome

  3. Structure of the ligand-binding domain (LBD) of human androgen receptor in complex with a selective modulator LGD2226

    International Nuclear Information System (INIS)

    Wang, Feng; Liu, Xiao-qin; Li, He; Liang, Kai-ni; Miner, Jeffrey N.; Hong, Mei; Kallel, E. Adam; Oeveren, Arjan van; Zhi, Lin; Jiang, Tao

    2006-01-01

    Crystal structure of the ligand-binding domain of androgen receptor in complex with LGD2226. The androgen receptor (AR) is a ligand-inducible steroid hormone receptor that mediates androgen action, determining male sexual phenotypes and promoting spermatogenesis. As the androgens play a dominant role in male sexual development and function, steroidal androgen agonists have been used clinically for some years. However, there is a risk of potential side effects and most steroidal androgens cannot be dosed orally, which limits the use of these substances. 1,2-Dihydro-6-N,N-bis(2,2,2-trifluoroethyl) amino-4-trifluoromethyl-2-quinolinone (LGD2226) is a synthetic nonsteroidal ligand and a novel selective AR modulator. The crystal structure of the complex of LGD2226 with the androgen receptor ligand-binding domain (AR LBD) at 2.1 Å was solved and compared with the structure of the AR LBD–R1881 complex. It is hoped that this will aid in further explaining the selectivity of LGD2226 observed in in vitro and in vivo assays and in developing more selective and effective therapeutic agents

  4. Crystallization and preliminary X-ray analysis of the N-terminal domain of human thioredoxin-interacting protein

    International Nuclear Information System (INIS)

    Polekhina, Galina; Ascher, David Benjamin; Kok, Shie Foong; Waltham, Mark

    2011-01-01

    The N-terminal domain of thioredoxin-interacting protein has been expressed, purified and crystallized. The crystals belonged to a monoclinic space group and diffracted to 3 Å resolution using synchrotron radiation. Thioredoxin-interacting protein (TXNIP) is a negative regulator of thioredoxin and its roles in the pathologies of diabetes and cardiovascular diseases have marked it out as a potential drug target. Expression of TXNIP is robustly induced under various stress conditions such as high glucose, heat shock, UV, H 2 O 2 and mechanical stress amongst others. Elevated levels of TXNIP result in the sequestration and inactivation of thioredoxin, leading to cellular oxidative stress. For some time, this was the only known function of TXNIP; however, more recently the protein has been shown to play a role in regulation of glucose uptake and activation of the inflammasome. Based on the primary sequence, TXNIP is remotely related to β-arrestins, which include the visual arrestins. TXNIP has thus been classified as a member of the α-arrestin family, which to date includes five other members. None of the other α-arrestins are known to interact with thioredoxin, although curiously one has been implicated in glucose uptake. In order to gain insight into the structure–function relationships of the α-arrestin protein family, and particularly that of TXNIP, the N-terminal domain of TXNIP has been crystallized. The crystals belonged to a monoclinic space group and diffracted to 3 Å resolution using synchrotron radiation

  5. Characterization of five new mutants in the carboxyl-terminal domain of human apolipoprotein E: No cosegregation with severe hyperlipidemia

    Energy Technology Data Exchange (ETDEWEB)

    Maagdenberg, A.M.J.M. van den; Bruijn, I.H. de; Hofker, M.H.; Frants, R.R. (Leiden Univ. (Netherlands)); Knijff, P. de; Smelt, A.H.M.; Leuven, J.A.G.; van' t Hooft, F.; Assmann, G.; Havekes, L.M. (Univ. Hospital, Leiden (Netherlands)); Weng, Wei; Funke, H. (Westfalische Wilhelms-Universitaet, Muester (Germany))

    1993-05-01

    Assessment of the apolipoprotein E (apoE) phenotype by isoelectric focusing of both hyperlipidemic and normolipidemic individuals identified five new variants. All mutations were confined to the downstream part of the APOE gene by using denaturing gradient gel electrophoresis (DGGE). Sequence analysis revealed five new mutations causing unique amino acid substitutions in the carboxyl-terminal part of the protein containing the putative lipid-binding domain. Three hyperlipoproteinemic probands were carriers of the APOE*2(Va1236[r arrow]Glu) allele, the APOE*3(Cys112-Arg; Arg251[r arrow]Gly) allele, or the APOE*1(Arg158[r arrow]Cys; Leu252[r arrow]Glu) allele. DGGE of the region encoding the receptor-binding domain was useful for haplotyping the mutations at codons 112 and 158. Family studies failed to demonstrate cosegregation between the new mutations and severe hyperlipoproteinemia, although a number of carriers for the APOE*3(Cys112[r arrow]Arg; Arg251[r arrow]Gly) allele and the APOE*1(Arg158-Cys; Leu252[r arrow]Glu) allele expressed hypertriglyceridemia and/ or hypercholesterolemia. Two other mutant alleles, APOE*4[sup [minus

  6. Recombinant human B7.2 IgV-like domain expressed in bacteria maintains its co-stimulatory activity in vitro.

    Science.gov (United States)

    Yan, Xiaocai; Ma, Jun; Zheng, Jin; Lai, Baochang; Geng, Yiping; Wang, Yili; Si, Lüsheng

    2002-07-01

    To investigate which of the two immunoglobulin (Ig)-like domains, the immunoglobulin variable region homologous domain IgV (hB7.2 IgV) and the immunoglobulin constant region homologous domain IgC (hB7.2 IgC) on the human B7.2 molecule contains receptor binding sites, and to evaluate whether the B7.2 protein expressed in bacteria has biological activity in vitro. Three fragments of hB7.2 IgV,hB7.2 IgC and the complete extracellular region of human B7.2 containing both the IgV and IgC domains,hB7.2 Ig (V+C), were amplified by PCR and subcloned into pGEM-Teasy. Three recombinants,pGEX-4T-3-hB7.2 IgV,pGEX-4T-3-hB7.2 IgC and pGEX-4T-3-hB7.2 Ig (V+C), were generated by cloning the fragments into a prokaryote expression plasmid (pGEX-4T-3) and transformed into the host strain E. coli DH5alpha. The relevant target fusion proteins consisting of GST and hB7.2 IgV,hB7.2 IgC and hB7.2 Ig (V+C), were identified by SDS-PAGE and Western blotting. With the presence of the first signal imitated by anti-CD3 antibody, T cell activation was observed by exposing purified T lymphocytes to each soluble form of the three bacterially-produced human B7.2 fusion proteins by [(3)H]-TdR incorporation. Three recombinant fusion proteins of human B7.2, GST-hB7.2 IgV, GST-hB7.2 IgC and GST-hB7.2 Ig (V+C) were produced and detected in inclusion body form from engineered bacteria. With the first signal present,T lymphocytes proliferated when co-stimulated by bacterially-produced either GST-hB7.2 Ig (V+C) or GST-hB7.2 IgV fusion proteins, but not by GST-hB7.2 IgC. Functional human B7.2 fusion protein can be produced in bacteria. The IgV-like domain of human B7.2 is sufficient for B7.2 to interact with its counter-receptors and co-stimulate T lymphocytes.

  7. Chemical applicability domain of the local lymph node assay (LLNA) for skin sensitisation potency. Part 4. Quantitative correlation of LLNA potency with human potency.

    Science.gov (United States)

    Roberts, David W; Api, Anne Marie

    2018-07-01

    Prediction of skin sensitisation potential and potency by non-animal methods is the target of many active research programmes. Although the aim is to predict sensitisation potential and potency in humans, data from the murine local lymph node assay (LLNA) constitute much the largest source of quantitative data on in vivo skin sensitisation. The LLNA has been the preferred in vivo method for identification of skin sensitising chemicals and as such is potentially valuable as a benchmark for assessment of non-animal approaches. However, in common with all predictive test methods, the LLNA is subject to false positives and false negatives with an overall level of accuracy said variously to be approximately 80% or 90%. It is also necessary to consider the extent to which, for true positives, LLNA potency correlates with human potency. In this paper LLNA potency and human potency are compared so as to express quantitatively the correlation between them, and reasons for non-agreement between LLNA and human potency are analysed. This leads to a better definition of the applicability domain of the LLNA, within which LLNA data can be used confidently to predict human potency and as a benchmark to assess the performance of non-animal approaches. Copyright © 2018. Published by Elsevier Inc.

  8. A Sequence in the loop domain of hepatitis C virus E2 protein identified in silico as crucial for the selective binding to human CD81.

    Directory of Open Access Journals (Sweden)

    Chun-Chun Chang

    Full Text Available Hepatitis C virus (HCV is a species-specific pathogenic virus that infects only humans and chimpanzees. Previous studies have indicated that interactions between the HCV E2 protein and CD81 on host cells are required for HCV infection. To determine the crucial factors for species-specific interactions at the molecular level, this study employed in silico molecular docking involving molecular dynamic simulations of the binding of HCV E2 onto human and rat CD81s. In vitro experiments including surface plasmon resonance measurements and cellular binding assays were applied for simple validations of the in silico results. The in silico studies identified two binding regions on the HCV E2 loop domain, namely E2-site1 and E2-site2, as being crucial for the interactions with CD81s, with the E2-site2 as the determinant factor for human-specific binding. Free energy calculations indicated that the E2/CD81 binding process might follow a two-step model involving (i the electrostatic interaction-driven initial binding of human-specific E2-site2, followed by (ii changes in the E2 orientation to facilitate the hydrophobic and van der Waals interaction-driven binding of E2-site1. The sequence of the human-specific, stronger-binding E2-site2 could serve as a candidate template for the future development of HCV-inhibiting peptide drugs.

  9. Crystallization and initial X-ray diffraction study of the three PASTA domains of the Ser/Thr kinase Stk1 from the human pathogen Staphylococcus aureus

    International Nuclear Information System (INIS)

    Paracuellos, Patricia; Ballandras, Allison; Robert, Xavier; Cozzone, Alain J.; Duclos, Bertrand; Gouet, Patrice

    2009-01-01

    Crystallization conditions have been determined for an extracellular portion of the Ser/Thr kinase Stk1 from the human pathogen S. aureus that contains three PASTA subunits. Synchrotron data have been collected to a resolution of 2.9 Å. Phasing is in progress. PASTA subunits (∼70 amino acids) are specific to bacterial serine/threonine kinases and to penicillin-binding proteins (PBPs) and are involved in the synthesis of peptidoglycan. The human pathogen Staphylococcus aureus contains a serine/threonine kinase, Stk1, which plays a major role in virulence. A recombinant His-tagged portion of the extracellular domain of Stk1 containing three PASTA subunits has been crystallized using zinc sulfate as a crystallizing agent. The crystals belonged to the tetragonal space group P4 1 22, with unit-cell parameters a = 68.0, b = 68.0, c = 158.1 Å. Structure determination by the MAD method is now in progress

  10. Stability-Diversity Tradeoffs Impose Fundamental Constraints on Selection of Synthetic Human VH/VL Single-Domain Antibodies from In Vitro Display Libraries.

    Science.gov (United States)

    Henry, Kevin A; Kim, Dae Young; Kandalaft, Hiba; Lowden, Michael J; Yang, Qingling; Schrag, Joseph D; Hussack, Greg; MacKenzie, C Roger; Tanha, Jamshid

    2017-01-01

    Human autonomous V H /V L single-domain antibodies (sdAbs) are attractive therapeutic molecules, but often suffer from suboptimal stability, solubility and affinity for cognate antigens. Most commonly, human sdAbs have been isolated from in vitro display libraries constructed via synthetic randomization of rearranged V H /V L domains. Here, we describe the design and characterization of three novel human V H /V L sdAb libraries through a process of: (i) exhaustive biophysical characterization of 20 potential V H /V L sdAb library scaffolds, including assessment of expression yield, aggregation resistance, thermostability and tolerance to complementarity-determining region (CDR) substitutions; (ii) in vitro randomization of the CDRs of three V H /V L sdAb scaffolds, with tailored amino acid representation designed to promote solubility and expressibility; and (iii) systematic benchmarking of the three V H /V L libraries by panning against five model antigens. We isolated ≥1 antigen-specific human sdAb against four of five targets (13 V H s and 7 V L s in total); these were predominantly monomeric, had antigen-binding affinities ranging from 5 nM to 12 µM (average: 2-3 µM), but had highly variable expression yields (range: 0.1-19 mg/L). Despite our efforts to identify the most stable V H /V L scaffolds, selection of antigen-specific binders from these libraries was unpredictable (overall success rate for all library-target screens: ~53%) with a high attrition rate of sdAbs exhibiting false positive binding by ELISA. By analyzing V H /V L sdAb library sequence composition following selection for monomeric antibody expression (binding to protein A/L followed by amplification in bacterial cells), we found that some V H /V L sdAbs had marked growth advantages over others, and that the amino acid composition of the CDRs of this set of sdAbs was dramatically restricted (bias toward Asp and His and away from aromatic and hydrophobic residues). Thus, CDR sequence

  11. The PDZ Ligand Domain of the Human Papillomavirus Type 16 E6 Protein Is Required for E6's Induction of Epithelial Hyperplasia In Vivo

    Science.gov (United States)

    Nguyen, Marie L.; Nguyen, Minh M.; Lee, Denis; Griep, Anne E.; Lambert, Paul F.

    2003-01-01

    Human papillomaviruses (HPVs) are the causative agent of warts. Infections with high-risk HPVs are associated with anogenital and head and neck cancers. One of the viral genes responsible for HPV's oncogenic activity is E6. Mice expressing the HPV-16 E6 protein in their epidermis (K14E6WT) develop epithelial hyperplasia and squamous carcinomas. Numerous cellular proteins interact with E6, some of which can be grouped based on common amino acid motifs in their E6-binding domains. One such group, the PDZ partners, including hDLG, hSCRIBBLE, MUPP1, and MAGI, bind to the carboxy-terminal four amino acids of E6 through their PDZ domains. E6's interaction with the PDZ partners leads to their degradation. Additionally, E6's binding to PDZ proteins has been correlated with its ability to transform baby rat kidney cells in tissue culture and to confer tumorigenicity onto cells in xenograft experiments. To address whether the ability of E6 to bind PDZ domain partners is necessary for E6 to confer epithelial hyperproliferation in vivo, we generated transgenic mice that express in stratified squamous epithelia a mutant of E6 lacking the last six amino acids at its carboxyl terminus, E6Δ146-151, from the human keratin 14 (K14) promoter. The K14E6Δ146-151 mice exhibit a radiation response similar to that of the K14E6WT mice, demonstrating that this protein, as predicted, retains an ability to inactivate p53. However, the K14E6Δ146-151 mice fail to display epithelial hyperplasia. These results indicate that an interaction of E6 with PDZ partners is necessary for its induction of epithelial hyperplasia. PMID:12768014

  12. Computed Tomography (CT) -- Sinuses

    Medline Plus

    Full Text Available ... Physician Resources Professions Site Index A-Z Computed Tomography (CT) - Sinuses Computed tomography (CT) of the sinuses ... CT of the Sinuses? What is CT (Computed Tomography) of the Sinuses? Computed tomography, more commonly known ...

  13. Crystallization and preliminary X-ray diffraction analysis of the C-terminal domain of the human spliceosomal DExD/H-box protein hPrp22

    International Nuclear Information System (INIS)

    Kudlinzki, Denis; Nagel, Christian; Ficner, Ralf

    2009-01-01

    The cloning, purification and crystallization of the C-terminal domain of human hPrp22 are reported. This communication also contains data for the preliminary X-ray diffraction analysis. The Homo sapiens DExD/H-box protein hPrp22 is a crucial component of the eukaryotic pre-mRNA splicing machinery. Within the splicing cycle, it is involved in the ligation of exons and generation of the lariat and it additionally catalyzes the release of mature mRNA from the spliceosomal U5 snRNP. The yeast homologue of this protein, yPrp22, shows ATP-dependent RNA-helicase activity and is capable of unwinding RNA/RNA duplex molecules. A truncated construct coding for residues 950–1183 of human Prp22, comprising the structurally and functionally uncharacterized C-terminal domain, was cloned into an Escherichia coli expression vector. The protein was subsequently overproduced, purified and crystallized. The crystals obtained diffracted to 2.1 Å resolution, belonged to the tetragonal space group P4 1 2 1 2 or P4 3 2 1 2, with unit-cell parameters a = b = 78.2, c = 88.4 Å, and contained one molecule in the asymmetric unit

  14. Expression, purification, crystallization and preliminary X-ray diffraction studies of the human keratin 4-binding domain of serine-rich repeat protein 1 from Streptococcus agalactiae

    International Nuclear Information System (INIS)

    Sundaresan, Ramya; Samen, Ulrike; Ponnuraj, Karthe

    2011-01-01

    Expression, purification and crystallization of Srr-1-K4BD, a human keratin 4-binding domain of serine-rich repeat protein 1 from S. agalactiae, was carried out. Native crystals of Srr-1-K4BD diffracted to 3.8 Å resolution using synchrotron radiation. Serine-rich repeat protein 1 (Srr-1) is a surface protein from Streptococcus agalactiae. A 17 kDa region of this protein has been identified to bind to human keratin 4 (K4) and is termed the Srr-1 K4-binding domain (Srr-1-K4BD). Recombinant Srr-1-K4BD was overexpressed in Escherichia coli BL21 (DE3) cells. Native and selenomethionine-substituted proteins were prepared using Luria–Bertani (LB) and M9 minimal media, respectively. A two-step purification protocol was carried out to obtain a final homogenous sample of Srr-1-K4BD. Crystals of native Srr-1-K4BD were obtained using PEG 3350 as a precipitant. The crystals diffracted to 3.8 Å resolution using synchrotron radiation and belonged to space group P2 1 , with unit-cell parameters a = 47.56, b = 59.48, c = 94.71 Å, β = 93.95°

  15. The realistic consideration of human factors in model based simulation tools for the air traffic control domain.

    Science.gov (United States)

    Duca, Gabriella; Attaianese, Erminia

    2012-01-01

    Advanced Air Traffic Management (ATM) concepts related to automation, airspace organization and operational procedures are driven by the overall goal to increase ATM system performance. Independently on the nature and/or impact of envisaged changes (e.g. from a short term procedure adjustment to a very long term operational concept or aid tools completion), the preliminary assessment of possible gains in airspace/airport capacity, safety and cost-effectiveness is done by running Model Based Simulations (MBSs, also known as Fast Time Simulations - FTS). Being a not human-in-the-loop technique, the reliability of a MBS results depend on the accuracy and significance of modeled human factors. Despite that, it can be observed in the practice that modeling tools commonly assume a generalized standardization of human behaviors and tasks and consider a very few range of work environment factors that, in the reality, affect the actual human-system performance. The present paper is aimed at opening a discussion about the possibility to keep task description and related weight at a high/general level, suitable for an efficient use of MBSs and, at the same time, increasing simulations reliability adopting some adjustment coming from the elaboration of further variables related to the human aspects of controllers workload.

  16. Interaction of CtBP with adenovirus E1A suppresses immortalization of primary epithelial cells and enhances virus replication during productive infection

    Energy Technology Data Exchange (ETDEWEB)

    Subramanian, T.; Zhao, Ling-jun; Chinnadurai, G., E-mail: chinnag@slu.edu

    2013-09-01

    Adenovirus E1A induces cell proliferation, oncogenic transformation and promotes viral replication through interaction with p300/CBP, TRRAP/p400 multi-protein complex and the retinoblastoma (pRb) family proteins through distinct domains in the E1A N-terminal region. The C-terminal region of E1A suppresses E1A/Ras co-transformation and interacts with FOXK1/K2, DYRK1A/1B/HAN11 and CtBP1/2 (CtBP) protein complexes. To specifically dissect the role of CtBP interaction with E1A, we engineered a mutation (DL→AS) within the CtBP-binding motif, PLDLS, and investigated the effect of the mutation on immortalization and Ras cooperative transformation of primary cells and viral replication. Our results suggest that CtBP–E1A interaction suppresses immortalization and Ras co-operative transformation of primary rodent epithelial cells without significantly influencing the tumorigenic activities of transformed cells in immunodeficient and immunocompetent animals. During productive infection, CtBP–E1A interaction enhances viral replication in human cells. Between the two CtBP family proteins, CtBP2 appears to restrict viral replication more than CtBP1 in human cells. - Highlights: • Adenovirus E1A C-terminal region suppresses E1A/Ras co-transformation. • This E1A region binds with FOXK, DYRK1/HAN11 and CtBP cellular protein complexes. • We found that E1A–CtBP interaction suppresses immortalization and transformation. • The interaction enhances viral replication in human cells.

  17. Evaluating differential nuclear DNA yield rates and osteocyte numbers among human bone tissue types: A synchrotron radiation micro-CT approach.

    Science.gov (United States)

    Andronowski, Janna M; Mundorff, Amy Z; Pratt, Isaac V; Davoren, Jon M; Cooper, David M L

    2017-05-01

    Molecular human identification has conventionally focused on DNA sampling from dense, weight-bearing cortical bone tissue, typically from femora or tibiae. A comparison of skeletal elements from three contemporary individuals demonstrated that elements with high quantities of cancellous bone yielded nuclear DNA at the highest rates, suggesting that preferentially sampling cortical bone may be suboptimal (Mundorff & Davoren, 2014). Despite these findings, the reason for the differential DNA yields between cortical and cancellous bone tissues remains unknown. The primary goal of this work is to ascertain whether differences in bone microstructure can be used to explain differential nuclear DNA yield among bone tissue types observed by Mundorff and Davoren (2014), with a focus on osteocytes and the three-dimensional (3D) quantification of their associated lacunae. Osteocytes and other bone cells are recognized to house DNA in bone tissue, thus examining the density of their lacunae may explain why nuclear DNA yield rates differ among bone tissue types. Lacunae were visualized and quantified using synchrotron radiation-based micro-Computed Tomographic imaging (SR micro-CT). Volumes of interest (VOIs) from cortical and cancellous bone tissues (n=129) were comparatively analyzed from the three skeletons sampled for Mundorff and Davoren's (2014) study. Analyses tested the primary hypothesis that the abundance and density of osteocytes (inferred from their lacunar spaces) vary between cortical and cancellous bone tissue types. Results demonstrated that osteocyte lacunar abundance and density vary between cortical and cancellous bone tissue types, with cortical bone VOIs containing a higher lacunar abundance and density. We found that the osteocyte lacunar density values are independent of nuclear DNA yield, suggesting an alternative explanation for the higher nuclear DNA yields from bones with greater quantities of cancellous bone tissue. The use of SR micro-CT allowed for

  18. First-in-Human PET/CT Imaging of Metastatic Neuroendocrine Neoplasms with Cyclotron-Produced 44Sc-DOTATOC: A Proof-of-Concept Study.

    Science.gov (United States)

    Singh, Aviral; van der Meulen, Nicholas P; Müller, Cristina; Klette, Ingo; Kulkarni, Harshad R; Türler, Andreas; Schibli, Roger; Baum, Richard P

    2017-05-01

    44 Sc is a promising positron emission tomography (PET) radionuclide (T 1/2  = 4.04 hours, E β+average  = 632 keV) and can be made available, using a cyclotron production route, in substantial quantities as a highly pure product. Herein, the authors report on a first-in-human PET/CT study using 44 Sc-DOTATOC prepared with cyclotron-produced 44 Sc. The production of 44 Sc was carried out through the 44 Ca(p,n) 44 Sc nuclear reaction at Paul Scherrer Institut, Switzerland. After separation, 44 Sc was shipped to Zentralklinik Bad Berka, Germany, where radiolabeling was performed, yielding radiochemically pure 44 Sc-DOTATOC. Two patients, currently followed up after peptide receptor radionuclide therapy of metastatic neuroendocrine neoplasms, participated in this proof-of-concept study. Blood sampling was performed before and after application of 44 Sc-DOTATOC. PET/CT acquisitions, performed at different time points after injection of 44 Sc-DOTATOC, allowed detection of even very small lesions on delayed scans. No clinical adverse effects were observed and the laboratory hematological, renal, and hepatic profiles remained unchanged. In this study, cyclotron-produced 44 Sc was used in the clinic for the first time. It is attractive for theranostic application with 177 Lu, 90 Y, or 47 Sc as therapeutic counterparts. 44 Sc-based radiopharmaceuticals will be of particular value for PET facilities without radiopharmacy, to which they can be shipped from a centralized production site.

  19. The segmentation of the human brain; a message to the neuroimaging community from an adjacent domain of the neurosciences

    NARCIS (Netherlands)

    Nieuwenhuys, R.

    2018-01-01

    Morphological and genoarchitectonic studies have conclusively shown that the human brain (and that of all vertebrates) is segmented i. e. is fundamentally composed of a number of rostrocaudally arranged brain segments or neuromeres. However in the current neuroimaging literature the term

  20. Traces of pFc' in IVIG interact with human IgG Fc domains and counteract aggregation

    NARCIS (Netherlands)

    Rispens, Theo; Himly, Martin; Ooievaar-de Heer, Pleuni; den Bleker, Tamara H.; Aalberse, Rob C.

    2010-01-01

    To prevent multimer formation, intravenous immunoglobulin (IVIG) is often treated with traces of pepsin. So far, the mechanism behind this treatment has been unclear. Recently, we reported that human IgG4 binds other IgG molecules via Fc-Fc interactions. Here we show that IVIG treated with traces of

  1. The epidermal growth factor-like domains of the human EMR2 receptor mediate cell attachment through chondroitin sulfate glycosaminoglycans

    NARCIS (Netherlands)

    Stacey, Martin; Chang, Gin-Wen; Davies, John Q.; Kwakkenbos, Mark J.; Sanderson, Ralph D.; Hamann, Jörg; Gordon, Siamon; Lin, Hsi-Hsien

    2003-01-01

    Using multivalent protein probes, an evolutionarily conserved endogenous ligand for EMR2, a human myeloid cell-restricted EGF-TM7 receptor, was identified on the surface of a number of adherent cell lines. In addition, in situ staining of the ligand has revealed specific in vivo patterns consistent

  2. CT cystography

    International Nuclear Information System (INIS)

    Tsili, A.

    2012-01-01

    Full text: Cancer of the urinary bladder is one of the commonest urothelial neoplasms. Conventional cystoscopy remains the mainstay in the diagnosis of urinary bladder carcinomas. But it is an invasive and uncomfortable procedure, associated with complications. Computed tomographic (CT) virtual cystoscopy has been proposed as an alternative imaging modality with potential advantages in the detection of urinary bladder neoplasms and good patient acceptance. It is a minimally invasive technique, allowing imaging of the urinary bladder in multiple planes and a 360 0 view. CT cystoscopy provides information about the location, size and morphologic features of urinary bladder lesions, indicating appropriate areas for biopsy. It can be performed in cases in which conventional cystoscopy is not feasible, such as in the presence of urethral strictures, marked prostatic hypertrophy or active bleeding, and in cases in which cystoscopic findings are inconclusive. The technique can be used to evaluate areas of the urinary bladder difficult to evaluate with cystoscopy, such as the anterior bladder neck and narrowmouthed diverticula. Finally, virtual cystoscopy provides both intraluminal and extraluminal pathologic changes, so intravesical disease and exrtavesical extension can be evaluated in the same study. One of the limitations of this technique is the difficulty to demonstrate small-sized lesions. The introduction of multi-detector row CT (MDCT) scanners was a major technological advancement because among other things it substantially improves z-axis resolution by reducing section collimation and allowing the detection of very small lesions. The near isotropic or isotropic pixels achieved with a multidetector CT scanner, enable the creation of multiplanar reformatted images with a resolution very close to that of the axial images and three-dimensional (3D) renderings of outstanding quality. Transverse, multiplanar reformations and virtual endoscopic images are complementary

  3. Transfer of the amino-terminal nuclear envelope targeting domain of human MX2 converts MX1 into an HIV-1 resistance factor.

    Science.gov (United States)

    Goujon, Caroline; Moncorgé, Olivier; Bauby, Hélène; Doyle, Tomas; Barclay, Wendy S; Malim, Michael H

    2014-08-01

    The myxovirus resistance 2 (MX2) protein of humans has been identified recently as an interferon (IFN)-inducible inhibitor of human immunodeficiency virus type 1 (HIV-1) that acts at a late postentry step of infection to prevent the nuclear accumulation of viral cDNA (C. Goujon et al., Nature 502:559-562, 2013, http://dx.doi.org/10.1038/nature12542; M. Kane et al., Nature 502:563-566, 2013, http://dx.doi.org/10.1038/nature12653; Z. Liu et al., Cell Host Microbe 14:398-410, 2013, http://dx.doi.org/10.1016/j.chom.2013.08.015). In contrast, the closely related human MX1 protein, which suppresses infection by a range of RNA and DNA viruses (such as influenza A virus [FluAV]), is ineffective against HIV-1. Using a panel of engineered chimeric MX1/2 proteins, we demonstrate that the amino-terminal 91-amino-acid domain of MX2 confers full anti-HIV-1 function when transferred to the amino terminus of MX1, and that this fusion protein retains full anti-FluAV activity. Confocal microscopy experiments further show that this MX1/2 fusion, similar to MX2 but not MX1, can localize to the nuclear envelope (NE), linking HIV-1 inhibition with MX accumulation at the NE. MX proteins are dynamin-like GTPases, and while MX1 antiviral function requires GTPase activity, neither MX2 nor MX1/2 chimeras require this attribute to inhibit HIV-1. This key discrepancy between the characteristics of MX1- and MX2-mediated viral resistance, together with previous observations showing that the L4 loop of the stalk domain of MX1 is a critical determinant of viral substrate specificity, presumably reflect fundamental differences in the mechanisms of antiviral suppression. Accordingly, we propose that further comparative studies of MX proteins will help illuminate the molecular basis and subcellular localization requirements for implementing the noted diversity of virus inhibition by MX proteins. Interferon (IFN) elicits an antiviral state in cells through the induction of hundreds of IFN

  4. Solution Structure and Backbone Dynamics of the Pleckstrin Homology Domain of the Human Protein Kinase B (PKB/Akt). Interaction with Inositol Phosphates

    International Nuclear Information System (INIS)

    Auguin, Daniel; Barthe, Philippe; Auge-Senegas, Marie-Therese; Stern, Marc-Henri; Noguchi, Masayuki; Roumestand, Christian

    2004-01-01

    The programmed cell death occurs as part of normal mammalian development. The induction of developmental cell death is a highly regulated process and can be suppressed by a variety of extracellular stimuli. Recently, the ability of trophic factors to promote survival have been attributed, at least in part, to the phosphatidylinositide 3'-OH kinase (PI3K)/Protein Kinase B (PKB, also named Akt) cascade. Several targets of the PI3K/PKB signaling pathway have been identified that may underlie the ability of this regulatory cascade to promote cell survival. PKB possesses a N-terminal Pleckstrin Homology (PH) domain that binds specifically and with high affinity to PtIns(3,4,5)P 3 and PtIns(3,4)P 2 , the PI3K second messengers. PKB is then recruited to the plasma membrane by virtue of its interaction with 3'-OH phosphatidylinositides and activated. Recent evidence indicates that PKB is active in various types of human cancer; constitutive PKB signaling activation is believed to promote proliferation and increased cell survival, thereby contributing to cancer progression. Thus, it has been shown that induction of PKB activity is augmented by the TCL1/MTCP1 oncoproteins through a physical association requiring the PKB PH domain. Here we present the three-dimensional solution structure of the PH domain of the human protein PKB (isoform β). PKBβ-PH is an electrostatically polarized molecule that adopts the same fold and topology as other PH-domains, consisting of a β-sandwich of seven strands capped on one top by an α-helix. The opposite face presents three variable loops that appear poorly defined in the NMR structure. Measurements of 15 N spin relaxation times and heteronuclear 15 N{ 1 H}NOEs showed that this poor definition is due to intrinsic flexibility, involving complex motions on different time scales. Chemical shift mapping studies correctly defined the binding site of Ins(1,3,4,5)P 4 (the head group of PtIns(3,4,5)P 3 ), as was previously proposed from a

  5. Innate immune response of human plasmacytoid dendritic cells to poxvirus infection is subverted by vaccinia E3 via its Z-DNA/RNA binding domain.

    Directory of Open Access Journals (Sweden)

    Hua Cao

    Full Text Available Plasmacytoid dendritic cells (pDCs play important roles in antiviral innate immunity by producing type I interferon (IFN. In this study, we assess the immune responses of primary human pDCs to two poxviruses, vaccinia and myxoma virus. Vaccinia, an orthopoxvirus, was used for immunization against smallpox, a contagious human disease with high mortality. Myxoma virus, a Leporipoxvirus, causes lethal disease in rabbits, but is non-pathogenic in humans. We report that myxoma virus infection of human pDCs induces IFN-α and TNF production, whereas vaccinia infection does not. Co-infection of pDCs with myxoma virus plus vaccinia blocks myxoma induction effects. We find that heat-inactivated vaccinia (Heat-VAC; by incubating the virus at 55°C for 1 h gains the ability to induce IFN-α and TNF in primary human pDCs. Induction of IFN-α in pDCs by myxoma virus or Heat-VAC is blocked by chloroquine, which inhibits endosomal acidification required for TLR7/9 signaling, and by inhibitors of cellular kinases PI3K and Akt. Using purified pDCs from genetic knockout mice, we demonstrate that Heat-VAC-induced type I IFN production in pDCs requires the endosomal RNA sensor TLR7 and its adaptor MyD88, transcription factor IRF7 and the type I IFN feedback loop mediated by IFNAR1. These results indicate that (i vaccinia virus, but not myxoma virus, expresses inhibitor(s of the poxvirus sensing pathway(s in pDCs; and (ii Heat-VAC infection fails to produce inhibitor(s but rather produces novel activator(s, likely viral RNA transcripts that are sensed by the TLR7/MyD88 pathway. Using vaccinia gene deletion mutants, we show that the Z-DNA/RNA binding domain at the N-terminus of the vaccinia immunomodulatory E3 protein is an antagonist of the innate immune response of human pDCs to poxvirus infection and TLR agonists. The myxoma virus ortholog of vaccinia E3 (M029 lacks the N-terminal Z-DNA/RNA binding domain, which might contribute to the immunostimulating

  6. Innate Immune Response of Human Plasmacytoid Dendritic Cells to Poxvirus Infection Is Subverted by Vaccinia E3 via Its Z-DNA/RNA Binding Domain

    Science.gov (United States)

    Dai, Peihong; Wang, Weiyi; Li, Hao; Yuan, Jianda; Wang, Fangjin; Fang, Chee-Mun; Pitha, Paula M; Liu, Jia; Condit, Richard C; McFadden, Grant; Merghoub, Taha; Houghton, Alan N; Young, James W; Shuman, Stewart; Deng, Liang

    2012-01-01

    Plasmacytoid dendritic cells (pDCs) play important roles in antiviral innate immunity by producing type I interferon (IFN). In this study, we assess the immune responses of primary human pDCs to two poxviruses, vaccinia and myxoma virus. Vaccinia, an orthopoxvirus, was used for immunization against smallpox, a contagious human disease with high mortality. Myxoma virus, a Leporipoxvirus, causes lethal disease in rabbits, but is non-pathogenic in humans. We report that myxoma virus infection of human pDCs induces IFN-α and TNF production, whereas vaccinia infection does not. Co-infection of pDCs with myxoma virus plus vaccinia blocks myxoma induction effects. We find that heat-inactivated vaccinia (Heat-VAC; by incubating the virus at 55°C for 1 h) gains the ability to induce IFN-α and TNF in primary human pDCs. Induction of IFN-α in pDCs by myxoma virus or Heat-VAC is blocked by chloroquine, which inhibits endosomal acidification required for TLR7/9 signaling, and by inhibitors of cellular kinases PI3K and Akt. Using purified pDCs from genetic knockout mice, we demonstrate that Heat-VAC-induced type I IFN production in pDCs requires the endosomal RNA sensor TLR7 and its adaptor MyD88, transcription factor IRF7 and the type I IFN feedback loop mediated by IFNAR1. These results indicate that (i) vaccinia virus, but not myxoma virus, expresses inhibitor(s) of the poxvirus sensing pathway(s) in pDCs; and (ii) Heat-VAC infection fails to produce inhibitor(s) but rather produces novel activator(s), likely viral RNA transcripts that are sensed by the TLR7/MyD88 pathway. Using vaccinia gene deletion mutants, we show that the Z-DNA/RNA binding domain at the N-terminus of the vaccinia immunomodulatory E3 protein is an antagonist of the innate immune response of human pDCs to poxvirus infection and TLR agonists. The myxoma virus ortholog of vaccinia E3 (M029) lacks the N-terminal Z-DNA/RNA binding domain, which might contribute to the immunostimulating properties of

  7. Cardiac CT

    International Nuclear Information System (INIS)

    Dewey, Marc

    2011-01-01

    Computed tomography of the heart has become a highly accurate diagnostic modality that is attracting increasing attention. This extensively illustrated book aims to assist the reader in integrating cardiac CT into daily clinical practice, while also reviewing its current technical status and applications. Clear guidance is provided on the performance and interpretation of imaging using the latest technology, which offers greater coverage, better spatial resolution, and faster imaging. The specific features of scanners from all four main vendors, including those that have only recently become available, are presented. Among the wide range of applications and issues to be discussed are coronary artery bypass grafts, stents, plaques, and anomalies, cardiac valves, congenital and acquired heart disease, and radiation exposure. Upcoming clinical uses of cardiac CT, such as plaque imaging and functional assessment, are also explored. (orig.)

  8. Cardiac CT

    Energy Technology Data Exchange (ETDEWEB)

    Dewey, Marc [Charite - Universitaetsmedizin Berlin (Germany). Inst. fuer Radiologie

    2011-07-01

    Computed tomography of the heart has become a highly accurate diagnostic modality that is attracting increasing attention. This extensively illustrated book aims to assist the reader in integrating cardiac CT into daily clinical practice, while also reviewing its current technical status and applications. Clear guidance is provided on the performance and interpretation of imaging using the latest technology, which offers greater coverage, better spatial resolution, and faster imaging. The specific features of scanners from all four main vendors, including those that have only recently become available, are presented. Among the wide range of applications and issues to be discussed are coronary artery bypass grafts, stents, plaques, and anomalies, cardiac valves, congenital and acquired heart disease, and radiation exposure. Upcoming clinical uses of cardiac CT, such as plaque imaging and functional assessment, are also explored. (orig.)

  9. Identification of the DNA-Binding Domains of Human Replication Protein A That Recognize G-Quadruplex DNA

    Directory of Open Access Journals (Sweden)

    Aishwarya Prakash

    2011-01-01

    Full Text Available Replication protein A (RPA, a key player in DNA metabolism, has 6 single-stranded DNA-(ssDNA- binding domains (DBDs A-F. SELEX experiments with the DBDs-C, -D, and -E retrieve a 20-nt G-quadruplex forming sequence. Binding studies show that RPA-DE binds preferentially to the G-quadruplex DNA, a unique preference not observed with other RPA constructs. Circular dichroism experiments show that RPA-CDE-core can unfold the G-quadruplex while RPA-DE stabilizes it. Binding studies show that RPA-C binds pyrimidine- and purine-rich sequences similarly. This difference between RPA-C and RPA-DE binding was also indicated by the inability of RPA-CDE-core to unfold an oligonucleotide containing a TC-region 5′ to the G-quadruplex. Molecular modeling studies of RPA-DE and telomere-binding proteins Pot1 and Stn1 reveal structural similarities between the proteins and illuminate potential DNA-binding sites for RPA-DE and Stn1. These data indicate that DBDs of RPA have different ssDNA recognition properties.

  10. Nucleotide-binding oligomerization domain 2 (NOD2) activation induces apoptosis of human oral squamous cell carcinoma cells.

    Science.gov (United States)

    Yoon, Hyo-Eun; Ahn, Mee-Young; Kwon, Seong-Min; Kim, Dong-Jae; Lee, Jun; Yoon, Jung-Hoon

    2016-04-01

    Microbial Pattern-recognition receptors (PRRs), such as nucleotide-binding oligomerization domains (NODs), are essential for mammalian innate immune response. This study was designed to determine the effect of NOD1 and NOD2 agonist on innate immune responses and antitumor activity in oral squamous cell carcinoma (OSCC) cells. NODs expression was examined by RT-PCR, and IL-8 production by NODs agonist was examined by ELISA. Western blot analysis was performed to determine the MAPK activation in response to their agonist. Cell proliferation was determined by MTT assay. Flow cytometry and Western blot analysis were performed to determine the MDP-induced cell death. The levels of NODs were apparently expressed in OSCC cells. NODs agonist, Tri-DAP and MDP, led to the production of IL-8 and MAPK activation. NOD2 agonist, MDP, inhibited the proliferation of YD-10B cells in a dose-dependent manner. Also, the ratio of Annexin V-positive cells and cleaved PARP was increased by MDP treatment in YD-10B cells, suggesting that MDP-induced cell death in YD-10B cells may be owing to apoptosis. Our results indicate that NODs are functionally expressed in OSCC cells and can trigger innate immune responses. In addition, NOD2 agonist inhibited cell proliferation and induced apoptosis. These findings provide the potential value of MDP as novel candidates for antitumor agents of OSCC. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  11. A mutation in human VAP-B--MSP domain, present in ALS patients, affects the interaction with other cellular proteins.

    Science.gov (United States)

    Mitne-Neto, M; Ramos, C R R; Pimenta, D C; Luz, J S; Nishimura, A L; Gonzales, F A; Oliveira, C C; Zatz, M

    2007-09-01

    Amyotrophic Lateral Sclerosis (ALS) is the most common adult-onset Motor Neuron Disease (MND), characterized by motor neurons death in the cortex, brainstem and spinal cord. Ten loci linked to Familial ALS have been mapped. ALS8 is caused by a substitution of a proline by a serine in the Vesicle-Associated Membrane Protein-Associated protein-B/C (VAP-B/C). VAP-B belongs to a highly conserved family of proteins implicated in Endoplasmic Reticulum-Golgi and intra-Golgi transport and microtubules stabilization. Previous studies demonstrated that the P56S mutation disrupts the subcellular localization of VAP-B and that this position would be essential for Unfolded Protein Response (UPR) induced by VAP-B. In the present work we expressed and purified recombinant wild-type and P56S mutant VAP-B-MSP domain for the analysis of its interactions with other cellular proteins. Our findings suggest that the P56S mutation may lead to a less stable interaction of this endoplasmic reticulum protein with at least two other proteins: tubulin and GAPDH. These two proteins have been previously related to other forms of neurodegenerative diseases and are potential key points to understand ALS8 pathogenesis and other forms of MND. Understanding the role of these protein interactions may help the treatment of this devastating disease in the future.

  12. Skipping of exon 27 in C3 gene compromises TED domain and results in complete human C3 deficiency.

    Science.gov (United States)

    da Silva, Karina Ribeiro; Fraga, Tatiana Rodrigues; Lucatelli, Juliana Faggion; Grumach, Anete Sevciovic; Isaac, Lourdes

    2016-05-01

    Primary deficiency of complement C3 is rare and usually associated with increased susceptibility to bacterial infections. In this work, we investigated the molecular basis of complete C3 deficiency in a Brazilian 9-year old female patient with a family history of consanguinity. Hemolytic assays revealed complete lack of complement-mediated hemolytic activity in the patient's serum. While levels of the complement regulatory proteins Factor I, Factor H and Factor B were normal in the patient's and family members' sera, complement C3 levels were undetectable in the patient's serum and were reduced by at least 50% in the sera of the patient's parents and brother. Additionally, no C3 could be observed in the patient's plasma and cell culture supernatants by Western blot. We also observed that patient's skin fibroblasts stimulated with Escherichia coli LPS were unable to secrete C3, which might be accumulated within the cells before being intracellularly degraded. Sequencing analysis of the patient's C3 cDNA revealed a genetic mutation responsible for the complete skipping of exon 27, resulting in the loss of 99 nucleotides (3450-3549) located in the TED domain. Sequencing of the intronic region between the exons 26 and 27 of the C3 gene (nucleotides 6690313-6690961) showed a nucleotide exchange (T→C) at position 6690626 located in a splicing donor site, resulting in the complete skipping of exon 27 in the C3 mRNA. Copyright © 2016. Published by Elsevier GmbH.

  13. Conserved helicase domain of human RecQ4 is required for strand annealing-independent DNA unwinding

    DEFF Research Database (Denmark)

    Rossi, Marie L; Ghosh, Avik K; Kulikowicz, Tomasz

    2010-01-01

    Humans have five members of the well conserved RecQ helicase family: RecQ1, Bloom syndrome protein (BLM), Werner syndrome protein (WRN), RecQ4, and RecQ5, which are all known for their roles in maintaining genome stability. BLM, WRN, and RecQ4 are associated with premature aging and cancer...... provide the first evidence that human RecQ4's unwinding is independent of strand annealing, and that it does not require the presence of excess ssDNA. Moreover, we demonstrate that a point mutation of the conserved lysine in the Walker A motif abolished helicase activity, implying that not the N...... activities and protein partners of RecQ4 are conserved with those of the other RecQ helicases....

  14. Construction of an Interface Terminology on SNOMED CT Generic Approach and Its Application in Intensive Care

    NARCIS (Netherlands)

    Bakhshi-Raiez, F.; Ahmadian, L.; Cornet, R.; de Jonge, E.; de Keizer, N. F.

    2010-01-01

    Objective: To provide a generic approach for developing a domain-specific interface terminology on SNOMED CT and to apply this approach to the domain of intensive care. Methods: The process of developing an interface terminology on SNOMED CT can be regarded as six sequential phases: domain analysis,

  15. A simple formulation for deriving effective atomic numbers via electron density calibration from dual-energy CT data in the human body.

    Science.gov (United States)

    Saito, Masatoshi; Sagara, Shota

    2017-06-01

    The main objective of this study is to propose a simple formulation (which we called DEEDZ) for deriving effective atomic numbers (Z eff ) via electron density (ρ e ) calibration from dual-energy (DE) CT data. We carried out numerical analysis of this DEEDZ method for a large variety of materials with known elemental compositions and mass densities using an available photon cross sections database. The new conversion approach was also applied to previously published experimental DECT data to validate its practical feasibility. We performed numerical analysis of the DEEDZ conversion method for tissue surrogates that have the same chemical compositions and mass densities as a commercial tissue-characterization phantom in order to determine the parameters necessary for the ρ e and Z eff calibrations in the DEEDZ conversion. These parameters were then applied to the human-body-equivalent tissues of ICRU Report 46 as objects of interest with unknown ρ e and Z eff . The attenuation coefficients of these materials were calculated using the XCOM photon cross sections database. We also applied the DEEDZ conversion to experimental DECT data available in the literature, which was measured for two commercial phantoms of different shapes and sizes using a dual-source CT scanner at 80 kV and 140 kV/Sn. The simulated Z eff 's were in excellent agreement with the reference values for almost all of the ICRU-46 human tissues over the Z eff range from 5.83 (gallstones-cholesterol) to 16.11 (bone mineral-hydroxyapatite). The relative deviations from the reference Z eff were within ± 0.3% for all materials, except for one outlier that presented a -3.1% deviation, namely, the thyroid. The reason for this discrepancy is that the thyroid contains a small amount of iodine, an element with a large atomic number (Z = 53). In the experimental case, we confirmed that the simple formulation with less fit parameters enable to calibrate Z eff as accurately as the existing calibration

  16. CT-Urography; Urografia CT

    Energy Technology Data Exchange (ETDEWEB)

    Dalla Palma, Ludovico; Grotto, Maurizio [Trieste Univ., Trieste (Italy). Dipartimento di scienze cliniche, morfologiche e tecnologiche, UCO di radiologia; Morra, Aldo [CRO, Aviano (Italy). Reparto di radiologia

    2005-09-15

    In this paper we present an overview of CT-Urography. With the advent of multislice CT scanners and the evolution of image processing methods this technique now affords optimal urographic images comparable to those obtained with conventional techniques. We describe the acquisition techniques and protocols used by the various authors. Effective radiation dose has conditioned the use of CT-Urography so that the tendency today is to reduce the number of scans by performing, after the non enhanced scan, a single contrast-enhanced scan comprising both the nephrographic and urographic phase. With the use of multislice CT the quality of the urogram improves with the number of slices. We illustrate a variety of processing techniques, multiplanar reconstruction (MPR), maximum (MIP) and average intensity projection (AIP) and volume rendering (VR) and present a series of upper urinary tract tumours testifying to the superiority of the AIP technique over MIP. We then review the results of comparative studies of CT-Urography with conventional urography in upper urinary tract diagnostics. Finally, we describe the advantages and limitations of CT-Urography. [Italian] Gli Autori presentano una panoramica sulla Urografia TC (Uro TC). L'avvento della TC multistrato e l'evoluzione delle tecniche di elaborazione consentono di ottenere dei quadri urografici ottimali comparativi con quelli convenzionali. Vengono ricordate le varie tecniche di acquisizione e i protocolli usati dai vari Autori. La dose effettiva di radiazioni ha rappresentato uno dei fattori condizionanti per cui oggi prevale l'orientamento di ridurre il numero di scansioni, considerando dopo la scansione senza mezzo di contrasto un'unica scansione contrasto grafica comprendente sia la fase nefrografica che quella urografica. Con l'uso della TC multistrato la qualita dell'urogramma migliora con l'aumento del numero degli strati. Vengono descritte le varie tecniche di elaborazione, la

  17. The p2 domain of human immunodeficiency virus type 1 Gag regulates sequential proteolytic processing and is required to produce fully infectious virions.

    Science.gov (United States)

    Pettit, S C; Moody, M D; Wehbie, R S; Kaplan, A H; Nantermet, P V; Klein, C A; Swanstrom, R

    1994-12-01

    The proteolytic processing sites of the human immunodeficiency virus type 1 (HIV-1) Gag precursor are cleaved in a sequential manner by the viral protease. We investigated the factors that regulate sequential processing. When full-length Gag protein was digested with recombinant HIV-1 protease in vitro, four of the five major processing sites in Gag were cleaved at rates that differ by as much as 400-fold. Three of these four processing sites were cleaved independently of the others. The CA/p2 site, however, was cleaved approximately 20-fold faster when the adjacent downstream p2/NC site was blocked from cleavage or when the p2 domain of Gag was deleted. These results suggest that the presence of a C-terminal p2 tail on processing intermediates slows cleavage at the upstream CA/p2 site. We also found that lower pH selectively accelerated cleavage of the CA/p2 processing site in the full-length precursor and as a peptide primarily by a sequence-based mechanism rather than by a change in protein conformation. Deletion of the p2 domain of Gag results in released virions that are less infectious despite the presence of the processed final products of Gag. These findings suggest that the p2 domain of HIV-1 Gag regulates the rate of cleavage at the CA/p2 processing site during sequential processing in vitro and in infected cells and that p2 may function in the proper assembly of virions.

  18. TGFβ1-mediated PI3K/Akt and p38 MAP kinase dependent alternative splicing of fibronectin extra domain A in human podocyte culture.

    Science.gov (United States)

    Madne, Tarunkumar Hemraj; Dockrell, Mark Edward Carl

    2018-04-30

    Alternative splicing is an important gene regulation process to distribute proteins in health and diseases. Extra Domain A+ Fibronectin (EDA+Fn) is an alternatively spliced form of fibronectin (Fn) protein, present in the extra cellular matrix (ECM) and a recognised marker of various pathologies. TGFβ1 has been shown to induce alternative splicing of EDA+Fn in many cell types. Podocytes are spectacular cell type and play a key role in filtration and synthesise ECM proteins in renal physiology and pathology. In our previous study we have demonstrated expression and alternative splicing of EDA+Fn in basal condition in human podocytes culture. TGFβ1 further induced the basal expression and alternative splicing of EDA+Fn through Alk5 receptor and SR proteins. In this study, we have investigated TGFβ1 mediated signalling involved in alternative splicing of EDA+Fn in human podocytes. We have performed western blotting to characterise the expression of the EDA+Fn protein and other signalling proteins and RT-PCR to look for signalling pathways involved in regulation of alternative splicing of EDA+Fn in conditionally immortalised human podocytes culture.We have used TGFβ1 as a stimulator and SB431542, SB202190 and LY294002 for inhibitory studies. In this work, we have demonstrated in human podocytes culture TGFβ1 2.5ng/ml induced phosphorylation of Smad1/5/8, Smad2 and Smad3 via the ALK5 receptor. TGFβ1 significantly induced the PI3K/Akt pathway and the PI3K/Akt pathway inhibitor LY294002 significantly downregulated basal as well as TGFβ1 induced alternative splicing of EDA+Fn in human podocytes. In addition to this, TGFβ1 significantly induced the p38 MAP kinase signalling pathway and p38 MAP kinase signalling pathway inhibitor SB202190 downregulated the TGFβ1-mediated alternative splicing of EDA+Fn in human podocytes. The results with PI3K and p38 MAP kinase signalling pathway suggest that inhibiting PI3K signalling pathway downregulated the basal alternative

  19. Trafficking Dynamics of PCSK9-Induced LDLR Degradation: Focus on Human PCSK9 Mutations and C-Terminal Domain.

    Directory of Open Access Journals (Sweden)

    Steve Poirier

    Full Text Available PCSK9 is a secreted ligand and negative post-translational regulator of low-density lipoprotein receptor (LDLR in hepatocytes. Gain-of-function (GOF or loss-of-function (LOF mutations in PCSK9 are directly correlated with high or low plasma LDL-cholesterol levels, respectively. Therefore, PCSK9 is a prevailing lipid-lowering target to prevent coronary heart diseases and stroke. Herein, we fused monomeric fluorescent proteins to PCSK9 and LDLR to visualize their intra- and extracellular trafficking dynamics by live confocal microscopy. Fluorescence recovery after photobleaching (FRAP showed that PCSK9 LOF R46L mutant and GOF mutations S127R and D129G, but not the LDLR high-affinity mutant D374Y, significantly accelerate PCSK9 exit from the endoplasmic reticulum (ER. Quantitative analysis of inverse FRAP revealed that only R46L presented a much slower trafficking from the trans-Golgi network (TGN to the plasma membrane and a lower mobile fraction likely suggesting accumulation or delayed exit at the TGN as an underlying mechanism. While not primarily involved in LDLR binding, PCSK9 C-terminal domain (CTD was found to be essential to induce LDLR degradation both upon its overexpression in cells or via the extracellular pathway. Our data revealed that PCSK9 CTD is required for the localization of PCSK9 at the TGN and increases its LDLR-mediated endocytosis. Interestingly, intracellular lysosomal targeting of PCSK9-ΔCTD was able to rescue its capacity to induce LDLR degradation emphasizing a role of the CTD in the sorting of PCSK9-LDLR complex towards late endocytic compartments. Finally, we validated our dual fluorescence system as a cell based-assay by preventing PCSK9 internalization using a PCSK9-LDLR blocking antibody, which may be expended to identify protein, peptide or small molecule inhibitors of PCSK9.

  20. Human cytomegalovirus and Epstein-Barr virus infection impact on {sup 18}F-FDG PET/CT SUVmax, CT volumetric and KRAS-based parameters of patients with locally advanced rectal cancer treated with neoadjuvant therapy

    Energy Technology Data Exchange (ETDEWEB)

    Sole, Claudio V. [Instituto de Radiomedicina, Department of Radiation Oncology, Santiago (Chile); School of Medicine Complutense University, Madrid (Spain); Calvo, Felipe A. [Hospital General Universitario Gregorio Maranon, Department of Oncology, Madrid (Spain); School of Medicine Complutense University, Madrid (Spain); Hospital General Universitario Gregorio Maranon, Institute for Sanitary Research, Madrid (Spain); Ferrer, Carlos [Hospital Provincial de Castellon, Institute of Oncology, Castellon de la Plana (Spain); School of Medicine Cardenal Herrera-CEU University, Castellon de la Plana (Spain); Alvarez, Emilio [School of Medicine Complutense University, Madrid (Spain); Hospital General Universitario Gregorio Maranon, Department of Pathology, Madrid (Spain); Hospital General Universitario Gregorio Maranon, Institute for Sanitary Research, Madrid (Spain); Carreras, Jose L. [School of Medicine Complutense University, Madrid (Spain); Hospital General Universitario Gregorio Maranon, Department of Radiology and Medical Physics, Madrid (Spain); Ochoa, Enrique [Hospital Provincial de Castellon, Institute of Oncology, Castellon de la Plana (Spain)

    2014-10-01

    It has long been debated whether human cytomegalovirus (HCMV) and Epstein-Barr virus (EBV) are associated with rectal cancer. The gene products of HCMV and EBV contribute to cell-cycle progression, mutagenesis, angiogenesis and immune evasion. The aim of this prospective study was to analyse the association between infection of a tumour by HCMV and EBV and clinical, histological, metabolic ({sup 18}F-FDG uptake), volumetric (from CT) and molecular (KRAS status) features and long-term outcomes in a homogeneously treated group of patients with locally advanced rectal cancer. HCMV and EBV were detected in pretreatment biopsies using polymerase chain reaction (PCR). The Cox proportional hazards regression model was used to explore associations between viral infection and disease-free survival (DFS) and overall survival (OS). We analysed 37 patients with a median follow-up of 74 months (range 5-173 months). Locoregional control, OS and DFS at 5 years were 93 %, 74 % and 71 %, respectively. Patients with HCMV/EBV coinfection had a significantly higher maximum standardized uptake value than patients without viral coinfection (p = 0.02). Significant differences were also observed in staging and percentage relative reduction in tumour volume between patients with and without HCMV infection (p < 0.01) and EBV infection (p < 0.01). KRAS wildtype status was significantly more frequently observed in patients with EBV infection (p <0.01) and HCMV/EBV co-infection (p = 0.04). No significant differences were observed in OS or DFS between patients with and without EBV infection (p = 0.88 and 0.73), HCMV infection (p = 0.84 and 0.79), and EBV/CMV coinfection (p = 0.24 and 0.39). This pilot study showed that viral infections were associated with metabolic staging differences, and differences in the evolution of metabolic and volumetric parameters and KRAS mutations. Further findings of specific features will help determine the best candidates for metabolic and volumetric staging and

  1. Human cytomegalovirus and Epstein-Barr virus infection impact on 18F-FDG PET/CT SUVmax, CT volumetric and KRAS-based parameters of patients with locally advanced rectal cancer treated with neoadjuvant therapy

    International Nuclear Information System (INIS)

    Sole, Claudio V.; Calvo, Felipe A.; Ferrer, Carlos; Alvarez, Emilio; Carreras, Jose L.; Ochoa, Enrique

    2015-01-01

    It has long been debated whether human cytomegalovirus (HCMV) and Epstein-Barr virus (EBV) are associated with rectal cancer. The gene products of HCMV and EBV contribute to cell-cycle progression, mutagenesis, angiogenesis and immune evasion. The aim of this prospective study was to analyse the association between infection of a tumour by HCMV and EBV and clinical, histological, metabolic ( 18 F-FDG uptake), volumetric (from CT) and molecular (KRAS status) features and long-term outcomes in a homogeneously treated group of patients with locally advanced rectal cancer. HCMV and EBV were detected in pretreatment biopsies using polymerase chain reaction (PCR). The Cox proportional hazards regression model was used to explore associations between viral infection and disease-free survival (DFS) and overall survival (OS). We analysed 37 patients with a median follow-up of 74 months (range 5-173 months). Locoregional control, OS and DFS at 5 years were 93 %, 74 % and 71 %, respectively. Patients with HCMV/EBV coinfection had a significantly higher maximum standardized uptake value than patients without viral coinfection (p = 0.02). Significant differences were also observed in staging and percentage relative reduction in tumour volume between patients with and without HCMV infection (p < 0.01) and EBV infection (p < 0.01). KRAS wildtype status was significantly more frequently observed in patients with EBV infection (p <0.01) and HCMV/EBV co-infection (p = 0.04). No significant differences were observed in OS or DFS between patients with and without EBV infection (p = 0.88 and 0.73), HCMV infection (p = 0.84 and 0.79), and EBV/CMV coinfection (p = 0.24 and 0.39). This pilot study showed that viral infections were associated with metabolic staging differences, and differences in the evolution of metabolic and volumetric parameters and KRAS mutations. Further findings of specific features will help determine the best candidates for metabolic and volumetric staging and

  2. Alternative splicing targeting the hTAF4-TAFH domain of TAF4 represses proliferation and accelerates chondrogenic differentiation of human mesenchymal stem cells.

    Directory of Open Access Journals (Sweden)

    Jekaterina Kazantseva

    Full Text Available Transcription factor IID (TFIID activity can be regulated by cellular signals to specifically alter transcription of particular subsets of genes. Alternative splicing of TFIID subunits is often the result of external stimulation of upstream signaling pathways. We studied tissue distribution and cellular expression of different splice variants of TFIID subunit TAF4 mRNA and biochemical properties of its isoforms in human mesenchymal stem cells (hMSCs to reveal the role of different isoforms of TAF4 in the regulation of proliferation and differentiation. Expression of TAF4 transcripts with exons VI or VII deleted, which results in a structurally modified hTAF4-TAFH domain, increases during early differentiation of hMSCs into osteoblasts, adipocytes and chondrocytes. Functional analysis data reveals that TAF4 isoforms with the deleted hTAF4-TAFH domain repress proliferation of hMSCs and preferentially promote chondrogenic differentiation at the expense of other developmental pathways. This study also provides initial data showing possible cross-talks between TAF4 and TP53 activity and switching between canonical and non-canonical WNT signaling in the processes of proliferation and differentiation of hMSCs. We propose that TAF4 isoforms generated by the alternative splicing participate in the conversion of the cellular transcriptional programs from the maintenance of stem cell state to differentiation, particularly differentiation along the chondrogenic pathway.

  3. Structures of a minimal human CFTR first nucleotide-binding domain as a monomer, head-to-tail homodimer, and pathogenic mutant

    Energy Technology Data Exchange (ETDEWEB)

    Atwell, Shane; Brouillette, Christie G.; Conners, Kris; Emtage, Spencer; Gheyi, Tarun; Guggino, William B.; Hendle, Jorg; Hunt, John F.; Lewis, Hal A.; Lu, Frances; Protasevich, Irina I.; Rodgers, Logan A.; Romero, Rich; Wasserman, Stephen R.; Weber, Patricia C.; Wetmore, Diana; Zhang, Feiyu F.; Zhao, Xun (Cystic); (UAB); (JHU); (Columbia); (Lilly)

    2010-04-26

    Upon removal of the regulatory insert (RI), the first nucleotide binding domain (NBD1) of human cystic fibrosis transmembrane conductance regulator (CFTR) can be heterologously expressed and purified in a form that remains stable without solubilizing mutations, stabilizing agents or the regulatory extension (RE). This protein, NBD1 387-646({Delta}405-436), crystallizes as a homodimer with a head-to-tail association equivalent to the active conformation observed for NBDs from symmetric ATP transporters. The 1.7-{angstrom} resolution X-ray structure shows how ATP occupies the signature LSGGQ half-site in CFTR NBD1. The {Delta}F508 version of this protein also crystallizes as a homodimer and differs from the wild-type structure only in the vicinity of the disease-causing F508 deletion. A slightly longer construct crystallizes as a monomer. Comparisons of the homodimer structure with this and previously published monomeric structures show that the main effect of ATP binding at the signature site is to order the residues immediately preceding the signature sequence, residues 542-547, in a conformation compatible with nucleotide binding. These residues likely interact with a transmembrane domain intracellular loop in the full-length CFTR channel. The experiments described here show that removing the RI from NBD1 converts it into a well-behaved protein amenable to biophysical studies yielding deeper insights into CFTR function.

  4. HDJC9, a novel human type C DnaJ/HSP40 member interacts with and cochaperones HSP70 through the J domain

    International Nuclear Information System (INIS)

    Han Chaofeng; Chen Taoyong; Li Nan; Yang Mingjin; Wan Tao; Cao Xuetao

    2007-01-01

    HSP40s are a subfamily of heat shock proteins (HSPs) and play important roles in regulation of cell proliferation, survival and apoptosis by serving as chaperones for HSP70s. Up to date hundreds of HSP40 proteins derived from various species ranging from Escherichia coli to homo sapiens have been identified. Here we report the cloning and characterization of a novel human type C DnaJ homologue, HDJC9, containing a typical N-terminal J domain. HDJC9 is upregulated at both mRNA and protein levels upon various stress and mitogenic stimulations. HDJC9 is mainly localized in cell nuclei under normal culture conditions while it is transported into cytoplasm and plasma membrane upon heat shock stress through a non-classical and lipid-dependent pathway. HDJC9 can interact with HSP70s and activate the ATPase activity of HSP70s, both of which are dependent on the J domain. Our data suggest that HDJC9 is a novel cochaperone for HSP70s

  5. Regulatory domain or CpG site variation in SLC12A5, encoding the chloride transporter KCC2, in human autism and schizophrenia

    Directory of Open Access Journals (Sweden)

    Nancy D Merner

    2015-10-01

    Full Text Available Many encoded gene products responsible for neurodevelopmental disorders (NDs like autism spectrum disorders (ASD, schizophrenia (SCZ, intellectual disability (ID, and idiopathic generalized epilepsy (IGE converge on networks controlling synaptic function. An increase in KCC2 (SLC12A5 Cl- transporter activity drives the developmental GABA excitatory-inhibitory sequence, but the role of KCC2 in human NDs is essentially unknown. Here, we report two rare, non-synonymous (NS, functionally-impairing variants in the KCC2 C-terminal regulatory domain (CTRD in human ASD (R952H and R1049C and SCZ (R952H previously linked with IGE and familial febrile seizures, and another novel NS KCC2 variant in ASD (R1048W with highly-predicted pathogenicity. Exome data from 2517 simplex families in the ASD Simon Simplex Collection revealed significantly more KCC2 CTRD variants in ASD cases than controls, and interestingly, these were more often synonymous and predicted to disrupt or introduce a CpG site. Furthermore, full gene analysis showed ASD cases are more likely to contain rare KCC2 variants affecting CpG sites than controls. These data suggest genetically-encoded dysregulation of KCC2-dependent GABA signaling may contribute to multiple human NDs.

  6. Generalized resistance to thyroid hormone associated with a mutation in the ligand-binding domain of the human thyroid hormone receptor β

    International Nuclear Information System (INIS)

    Sakurai, A.; Takeda, K.; Ain, K.; Ceccarelli, P.; Nakai, A.; Seino, S.; Bell, G.I.; Refetoff, S.; DeGroot, L.J.

    1989-01-01

    The syndrome of generalized resistance to thyroid hormone is characterized by elevated circulating levels of thyroid hormone in the presence of an overall eumetabolic state and failure to respond normally to triiodothyronine. The authors have evaluated a family with inherited generalized resistance to thyroid hormone for abnormalities in the thyroid hormone nuclear receptors. A single guanine → cytosine replacement in the codon for amino acid 340 resulted in a glycine → arginine substitution in the hormone-binding domain of one of two alleles of the patient's thyroid hormone nuclear receptor β gene. In vitro translation products of this mutant human thyroid hormone nuclear receptor β gene did not bind triiodothyronine. Thus, generalized resistance to thyroid hormone can result from expression of an abnormal thyroid hormone nuclear receptor molecule

  7. GlycoDomainViewer

    DEFF Research Database (Denmark)

    Joshi, Hiren J; Jørgensen, Anja; Schjoldager, Katrine T

    2018-01-01

    features, which enhances visibility and accessibility of the wealth of glycoproteomic data being generated. The GlycoDomainViewer enables visual exploration of glycoproteomic data, incorporating information from recent N- and O-glycoproteome studies on human and animal cell lines and some organs and body...

  8. Role of microRNA-199a-5p and discoidin domain receptor 1 in human hepatocellular carcinoma invasion

    Directory of Open Access Journals (Sweden)

    Shen Qingli

    2010-08-01

    Full Text Available Abstract Background Micro-ribonucleic acid (miRNA-199a-5p has been reported to be decreased in hepatocellular carcinoma (HCC compared to normal tissue. Discoidin domain receptor-1 (DDR1 tyrosine kinase, involved in cell invasion-related signaling pathway, was predicted to be a potential target of miR-199a-5p by the use of miRNA target prediction algorithms. The aim of this study was to investigate the role of miR-199a-5p and DDR1 in HCC invasion. Methods Mature miR-199a-5p and DDR1 expression were evaluated in tumor and adjacent non-tumor liver tissues from 23 patients with HCC undergoing liver resection and five hepatoma cell lines by the use of real-time quantitative RT-PCR (qRT-PCR analysis. The effect of aberrant miR-199a-5p expression on cell invasion was assessed in vitro using HepG2 and SNU-182 hepatoma cell lines. Luciferase reporter assay was employed to validate DDR1 as a putative miR-199a-5p target gene. Regulation of DDR1 expression by miR-199a-5p was assessed by the use qRT-PCR and western blotting analysis. Results A significant down-regulation of miR-199a-5p was observed in 65.2% of HCC tissues and in four of five cell lines. In contrast, DDR1 expression was significantly increased in 52.2% of HCC samples and in two of five cell lines. Increased DDR1 expression in HCC was associated with advanced tumor stage. DDR1 was shown to be a direct target of miR-199a-5p by luciferase reporter assay. Transfection of miR-199a-5p inhibited invasion of HepG2 but not SNU-182 hepatoma cells. Conclusions Decreased expression of miR-199a-5p contributes to increased cell invasion by functional deregulation of DDR1 activity in HCC. However, the effect of miR-199a-5p on DDR1 varies among individuals and hepatoma cell lines. These findings may have significant translational relevance for development of new targeted therapies as well as prognostic prediction for patients with HCC.

  9. Identification of a novel antimicrobial peptide from human hepatitis B virus core protein arginine-rich domain (ARD.

    Directory of Open Access Journals (Sweden)

    Heng-Li Chen

    Full Text Available The rise of multidrug-resistant (MDR pathogens causes an increasing challenge to public health. Antimicrobial peptides are considered a possible solution to this problem. HBV core protein (HBc contains an arginine-rich domain (ARD at its C-terminus, which consists of 16 arginine residues separated into four clusters (ARD I to IV. In this study, we demonstrated that the peptide containing the full-length ARD I-IV (HBc147-183 has a broad-spectrum antimicrobial activity at micro-molar concentrations, including some MDR and colistin (polymyxin E-resistant Acinetobacter baumannii. Furthermore, confocal fluorescence microscopy and SYTOX Green uptake assay indicated that this peptide killed Gram-negative and Gram-positive bacteria by membrane permeabilization or DNA binding. In addition, peptide ARD II-IV (HBc153-176 and ARD I-III (HBc147-167 were found to be necessary and sufficient for the activity against P. aeruginosa and K. peumoniae. The antimicrobial activity of HBc ARD peptides can be attenuated by the addition of LPS. HBc ARD peptide was shown to be capable of direct binding to the Lipid A of lipopolysaccharide (LPS in several in vitro binding assays. Peptide ARD I-IV (HBc147-183 had no detectable cytotoxicity in various tissue culture systems and a mouse animal model. In the mouse model by intraperitoneal (i.p. inoculation with Staphylococcus aureus, timely treatment by i.p. injection with ARD peptide resulted in 100-fold reduction of bacteria load in blood, liver and spleen, as well as 100% protection of inoculated animals from death. If peptide was injected when bacterial load in the blood reached its peak, the protection rate dropped to 40%. Similar results were observed in K. peumoniae using an IVIS imaging system. The finding of anti-microbial HBc ARD is discussed in the context of commensal gut microbiota, development of intrahepatic anti-viral immunity and establishment of chronic infection with HBV. Our current results suggested that

  10. Structure of the lutein-binding domain of human StARD3 at 1.74 Å resolution and model of a complex with lutein

    Energy Technology Data Exchange (ETDEWEB)

    Horvath, Martin P., E-mail: martin.horvath@utah.edu; George, Evan W.; Tran, Quang T.; Baumgardner, Kody; Zharov, Gabe; Lee, Sarah [University of Utah, 257 S 1400 E, Salt Lake City, UT 84112 (United States); Sharifzadeh, Hassan; Shihab, Saeed; Mattinson, Ty; Li, Binxing; Bernstein, Paul S., E-mail: martin.horvath@utah.edu [Moran Eye Center, University of Utah School of Medicine, Salt Lake City, UT 84132 (United States)

    2016-07-27

    The structure of a START-domain protein known to bind lutein in the human retina is reported to an improved resolution limit. Rigid-body docking demonstrates that at least a portion of lutein must protrude from the large tunnel-like cavity characteristic of this helix-grip protein and suggests a mechanism for lutein binding specificity. A crystal structure of the lutein-binding domain of human StARD3 (StAR-related lipid-transfer protein 3; also known as MLN64) has been refined to 1.74 Å resolution. A previous structure of the same protein determined to 2.2 Å resolution highlighted homology with StARD1 and shared cholesterol-binding character. StARD3 has since been recognized as a carotenoid-binding protein in the primate retina, where its biochemical function of binding lutein with specificity appears to be well suited to recruit this photoprotective molecule. The current and previous structures correspond closely to each other (r.m.s.d. of 0.25 Å), especially in terms of the helix-grip fold constructed around a solvent-filled cavity. Regions of interest were defined with alternate conformations in the current higher-resolution structure, including Arg351 found within the cavity and Ω1, a loop of four residues found just outside the cavity entrance. Models of the complex with lutein generated by rigid-body docking indicate that one of the ionone rings must protrude outside the cavity, and this insight has implications for molecular interactions with transport proteins and enzymes that act on lutein. Interestingly, models with the ∊-ionone ring characteristic of lutein pointing towards the bottom of the cavity were associated with fewer steric clashes, suggesting that steric complementarity and ligand asymmetry may play a role in discriminating lutein from the other ocular carotenoids zeaxanthin and meso-zeaxanthin, which only have β-ionone rings.

  11. Molecular mimicry of human tRNALys anti-codon domain by HIV-1 RNA genome facilitates tRNA primer annealing.

    Science.gov (United States)

    Jones, Christopher P; Saadatmand, Jenan; Kleiman, Lawrence; Musier-Forsyth, Karin

    2013-02-01

    The primer for initiating reverse transcription in human immunodeficiency virus type 1 (HIV-1) is tRNA(Lys3). Host cell tRNA(Lys) is selectively packaged into HIV-1 through a specific interaction between the major tRNA(Lys)-binding protein, human lysyl-tRNA synthetase (hLysRS), and the viral proteins Gag and GagPol. Annealing of the tRNA primer onto the complementary primer-binding site (PBS) in viral RNA is mediated by the nucleocapsid domain of Gag. The mechanism by which tRNA(Lys3) is targeted to the PBS and released from hLysRS prior to annealing is unknown. Here, we show that hLysRS specifically binds to a tRNA anti-codon-like element (TLE) in the HIV-1 genome, which mimics the anti-codon loop of tRNA(Lys) and is located proximal to the PBS. Mutation of the U-rich sequence within the TLE attenuates binding of hLysRS in vitro and reduces the amount of annealed tRNA(Lys3) in virions. Thus, LysRS binds specifically to the TLE, which is part of a larger LysRS binding domain in the viral RNA that includes elements of the Psi packaging signal. Our results suggest that HIV-1 uses molecular mimicry of the anti-codon of tRNA(Lys) to increase the efficiency of tRNA(Lys3) annealing to viral RNA.

  12. Distribution of the blood flow supplied by the vertebral artery in humans as assessed by emission CT

    International Nuclear Information System (INIS)

    Taki, W.; Handa, H.; Higa, T.; Tanada, S.; Fukuyama, H.; Fujita, T.; Yonekawa, Y.; Kameyama, M.; Torizuka, K.

    1984-01-01

    Though unilateral blood distribution of the vertebral artery has not been studied in humans, it is significant when considering the ischemic events especially embolic occlusion in vertebrobasilar system. To clarify the blood distribution, Krypton-81m was continuously infused into the vertebral arteries of 10 patients. Radioactivity was recorded by a rotating gamma camera. The perfusion images, reconstructed by computer in the transaxial plane, were classified according to whether the distribution was equal, or predominantly unilateral. Overlapping of blood supply from each vertebral artery was studied. In addition, the perfusion from one of the vertebral arteries revealed four distribution patterns with respect to laterality. A) ipsilateral; B) contralateral; C) bilateral; D) mosaic. Images obtained following perfusion of the vertebral artery show less uniformity compared to that of the internal carotid artery, suggesting a complicated vascular supply of the vertebral basilar territory. This complex distribution can be classified by the use of continuous infusion of Krypton-81m

  13. Non-destructive, preclinical evaluation of root canal anatomy of human teeth with flat-panel detector volume CT (FD-VCT)

    International Nuclear Information System (INIS)

    Heidrich, G.; Hassepass, F.; Dullin, C.; Grabbe, E.; Attin, T.; Hannig, C.

    2005-01-01

    Purpose: Successful endodontic diagnostics and therapy call for adequate depiction of the root canal anatomy with multimodal diagnostic imaging. The aim of the present study is to evaluate visualization of the endodont with flat-panel detector volume CT (FD-VCT). Materials and methods: 13 human teeth were examined with the prototype of a FD-VCT. After data acquisition and generation of volume data sets in volume rendering technology (VRT), the findings obtained were compared to conventional X-rays and cross-section preparations of the teeth. Results: The anatomical structures of the endodont such as root canals, side canals and communications between different root canals as well as dentricles could be detected precisely with FD-VCT. The length of curved root canals was also determined accurately. The spatial resolution of the system is around 140 μm. Only around 73% of the main root canals detected with FD-VCT and 87% of the roots could be visualized with conventional dental X-rays. None of the side canals, shown with FD-VCT, was detectable on conventional X-rays. In all cases the enamel and dentin of the teeth could be well delineated. No differences in image quality could be discerned between stored and freshly extracted teeth, or between primary and adult teeth. (orig.)

  14. A comparative study of N-glycolylneuraminic acid (Neu5Gc and cytotoxic T cell (CT carbohydrate expression in normal and dystrophin-deficient dog and human skeletal muscle.

    Directory of Open Access Journals (Sweden)

    Paul T Martin

    Full Text Available The expression of N-glycolylneuraminic acid (Neu5Gc and the cytotoxic T cell (CT carbohydrate can impact the severity of muscular dystrophy arising from the loss of dystrophin in mdx mice. Here, we describe the expression of these two glycans in skeletal muscles of dogs and humans with or without dystrophin-deficiency. Neu5Gc expression was highly reduced (>95% in muscle from normal golden retriever crosses (GR, n = 3 and from golden retriever with muscular dystrophy (GRMD, n = 5 dogs at multiple ages (3, 6 and 13 months when compared to mouse muscle, however, overall sialic acid expression in GR and GRMD muscles remained high at all ages. Neu5Gc was expressed on only a minority of GRMD satellite cells, CD8⁺ T lymphocytes and macrophages. Human muscle from normal (no evident disease, n = 3, Becker (BMD, n = 3 and Duchenne (DMD, n = 3 muscular dystrophy individuals had absent to very low Neu5Gc staining, but some punctate intracellular muscle staining was present in BMD and DMD muscles. The CT carbohydrate was localized to the neuromuscular junction in GR muscle, while GRMD muscles had increased expression on a subset of myofibers and macrophages. In humans, the CT carbohydrate was ectopically expressed on the sarcolemmal membrane of some BMD muscles, but not normal human or DMD muscles. These data are consistent with the notion that altered Neu5Gc and CT carbohydrate expression may modify disease severity resulting from dystrophin deficiency in dogs and humans.

  15. Domain crossing

    DEFF Research Database (Denmark)

    Schraefel, M. C.; Rouncefield, Mark; Kellogg, Wendy

    2012-01-01

    In CSCW, how much do we need to know about another domain/culture before we observe, intersect and intervene with designs. What optimally would that other culture need to know about us? Is this a “how long is a piece of string” question, or an inquiry where we can consider a variety of contexts a...

  16. Electron absorbed fractions of energy and S-values in an adult human skeleton based on {mu}CT images of trabecular bone

    Energy Technology Data Exchange (ETDEWEB)

    Kramer, R; Cassola, V F; Khoury, H J; De O Lira, C A B [Department of Nuclear Energy, Federal University of Pernambuco, Avenida Professor Luiz Freire, 1000, CEP 50740-540, Recife (Brazil); Richardson, R B [Radiation Protection Research and Instrumentation Branch, Atomic Energy of Canada Limited, Chalk River Laboratories, Chalk River, ON, K0J 1J0 (Canada); Vieira, J W [Federal Institute of Education, Science and Technology of Pernambuco, Recife (Brazil); Brown, K Robson, E-mail: rkramer@uol.com.br [Imaging Laboratory, Department of Archaeology and Anthropology, University of Bristol, Bristol (United Kingdom)

    2011-03-21

    When the human body is exposed to ionizing radiation, among the soft tissues at risk are the active marrow (AM) and the bone endosteum (BE) located in tiny, irregular cavities of trabecular bone. Determination of absorbed fractions (AFs) of energy or absorbed dose in the AM and the BE represent one of the major challenges of dosimetry. Recently, at the Department of Nuclear Energy at the Federal University of Pernambuco, a skeletal dosimetry method based on {mu}CT images of trabecular bone introduced into the spongiosa voxels of human phantoms has been developed and applied mainly to external exposure to photons. This study uses the same method to calculate AFs of energy and S-values (absorbed dose per unit activity) for electron-emitting radionuclides known to concentrate in skeletal tissues. The modelling of the skeletal tissue regions follows ICRP110, which defines the BE as a 50 {mu}m thick sub-region of marrow next to the bone surfaces. The paper presents mono-energetic AFs for the AM and the BE for eight different skeletal regions for electron source energies between 1 keV and 10 MeV. The S-values are given for the beta emitters {sup 14}C, {sup 59}Fe, {sup 131}I, {sup 89}Sr, {sup 32}P and {sup 90}Y. Comparisons with results from other investigations showed good agreement provided that differences between methodologies and trabecular bone volume fractions were properly taken into account. Additionally, a comparison was made between specific AFs of energy in the BE calculated for the actual 50 {mu}m endosteum and the previously recommended 10 {mu}m endosteum. The increase in endosteum thickness leads to a decrease of the endosteum absorbed dose by up to 3.7 fold when bone is the source region, while absorbed dose increases by {approx}20% when the beta emitters are in marrow.

  17. X-treme CT analysis of cancellous bone at the rotator cuff insertion in human individuals with osteoporosis: superficial versus deep quality.

    Science.gov (United States)

    Kirchhoff, Chlodwig; Kirchhoff, Sonja; Sprecher, Christoph M; Ahrens, Philipp; Imhoff, Andreas B; Hinterwimmer, Stefan; Milz, Stefan; Braunstein, Volker

    2013-03-01

    Rotator cuff (RC) repair-especially in the elderly population-is problematic since the patients suffer to a high extent from bone mineral density loss at the reattachment site. Therefore, the study was primarily driven by the question whether it is possible to reach more or qualitatively better cancellous bone and thus a more stable postoperative result if anchors with greater length are used for RC repair and/or the conventional anchors are screwed deeper into the bone. In anatomical terms, the question is raised whether cancellous bone is of better quality close to or far off the RC enthesis. Axial HRqCT scans (X-tremeCT, Scanco Medical) of 36 human cadaveric humeral heads (75 ± 11 years) were performed to determine the ratio of bone volume to total volume (BV/TV), trabecular thickness (Trab Th), number of trabecles (Trab N), trabecular separation (Trab Sp) as well as non-metric indices such as connectivity density (Conn Dens) and structure model index (SMI). Within the greater tuberosity (GT), 6 volumes of interest (VOI) (A1, B1, C1, A2, B2, C2), in the lesser tuberosity (LT) 2 VOIs (D1, D2) and one control VOI in the subchondral bone were set. The analyzed bone cylinder of each VOI was divided into a superficial and a deep portion. The parameters BV/TV, Trab N, Trab Th and Conn Dens in all volumes of the GT and LT revealed higher values in the superficial portion reaching different levels of significance (p significance for the non-metric parameter SMI in no volume of the GT/LT, although the higher values were found superficially. Our data show that cancellous bone presents with decreasing bone quality when analyzing increasingly deeper portions of the bone cylinders of the GT and LT starting at the articular surface. This information seems to be crucial for shoulder surgeons, especially when treating elderly patients. Our results clearly prove that screwing in anchors to a deeper extent will not improve stability, since the deeper bone stock is of worse

  18. Human dosimetry and preliminary tumor distribution of 18F-fluoropaclitaxel in healthy volunteers and newly diagnosed breast cancer patients using PET/CT.

    Science.gov (United States)

    Kurdziel, Karen A; Kalen, Joseph D; Hirsch, Jerry I; Wilson, John D; Bear, Harry D; Logan, Jean; McCumisky, James; Moorman-Sykes, Kathy; Adler, Stephen; Choyke, Peter L

    2011-09-01

    -fluoropaclitaxel PET/CT tumor imaging and provides radiation dosimetry measurements in humans. Although further study is needed, it is hoped that the measured intratumoral (18)F-fluoropaclitaxel distribution can serve as a surrogate for paclitaxel, and potentially other chemotherapeutic agent retention, in solid tumors.

  19. Human Dosimetry and Preliminary Tumor Distribution of 18F-Fluoropaclitaxel in Healthy Volunteers and Newly Diagnosed Breast Cancer Patients Using PET/CT

    International Nuclear Information System (INIS)

    Kurdziel, K.A.; Logan, J.; Kalen, J.D.; Hirsch, J.I.; Wilson, J.D.; Bear, H.D.; McCumisky, J.; Moorman-Sykes, K.; Adler, S.; Choyke, P.L.

    2011-01-01

    -fluoropaclitaxel PET/CT tumor imaging and provides radiation dosimetry measurements in humans. Although further study is needed, it is hoped that the measured intratumoral 18 F-fluoropaclitaxel distribution can serve as a surrogate for paclitaxel, and potentially other chemotherapeutic agent retention, in solid tumors.

  20. CT urography

    Energy Technology Data Exchange (ETDEWEB)

    Korobkin, M. [Dept. of Radiology, Univ. of Michigan, Ann Arbor, MI (United States)

    2005-11-15

    With the advent of multidetector row CT scanners, evaluation of the urothelium of the entire urinary tract with high-resolution thin sections during a single breath-hold has become a reality. Multidetector CT urography (MDCTU) is a single examination that allows evaluation of potential urinary tract calculi, renal parenchymal masses, and both benign and malignant urothelial lesions. Initial results with this new technique are encouraging. Current investigations of MDCTU focus on methods to improve opacification and distension of the upper urinary tract - the collecting systems, pelvis, and ureters. The role of abdominal compression, infusion of saline and/or furosemide, and optimal time delay of excretory phase imaging is being explored. Upper tract urothelial malignancies, including small lesions less the 5 mmin diameter, can be detected with high sensitivity. Methods to reduce radiation exposure are being explored, including split-bolus contrast injection techniques that combine nephrographic and excretory phases into a single phase. It is likely that in the near future, radiological evaluation of significant unexplained hematuria or of known or prior urothelial malignancy will consist of a single examination - MDCTU. (orig.)

  1. A single amino acid substitution within the transmembrane domain of the human immunodeficiency virus type 1 Vpu protein renders simian-human immunodeficiency virus (SHIVKU-1bMC33) susceptible to rimantadine

    International Nuclear Information System (INIS)

    Hout, David R.; Gomez, Lisa M.; Pacyniak, Erik; Miller, Jean-Marie; Hill, M. Sarah; Stephens, Edward B.

    2006-01-01

    Previous studies from our laboratory have shown that the transmembrane domain (TM) of the Vpu protein of human immunodeficiency virus type 1 (HIV-1) contributes to the pathogenesis of SHIV KU-1bMC33 in macaques and that the TM domain of Vpu could be replaced with the M2 protein viroporin from influenza A virus. Recently, we showed that the replacement of the TM domain of Vpu with that of the M2 protein of influenza A virus resulted in a virus (SHIV M2 ) that was sensitive to rimantadine [Hout, D.R., Gomez, M.L., Pacyniak, E., Gomez, L.M., Inbody, S.H., Mulcahy, E.R., Culley, N., Pinson, D.M., Powers, M.F., Wong, S.W., Stephens, E.B., 2006. Substitution of the transmembrane domain of Vpu in simian human immunodeficiency virus (SHIV KU-1bMC33 ) with that of M2 of influenza A results in a virus that is sensitive to inhibitors of the M2 ion channel and is pathogenic for pig-tailed macaques. Virology 344, 541-558]. Based on previous studies of the M2 protein which have shown that the His-X-X-X-Trp motif within the M2 is essential to the function of the M2 proton channel, we have constructed a novel SHIV in which the alanine at position 19 of the TM domain was replaced with a histidine residue resulting in the motif His-Ile-Leu-Val-Trp. The SHIV VpuA19H replicated with similar kinetics as the parental SHIV KU-1bMC33 and pulse-chase analysis revealed that the processing of viral proteins was similar to SHIV KU-1bMC33 . This SHIV VpuA19H virus was found to be more sensitive to the M2 ion channel blocker rimantadine than SHIV M2 . Electron microscopic examination of SHIV VpuA19H -infected cells treated with rimantadine revealed an accumulation of viral particles at the cell surface and within intracellular vesicles, which was similar to that previously observed to SHIV M2 -infected cells treated with rimantadine. These data indicate that the Vpu protein of HIV-1 can be converted into a rimantadine-sensitive ion channel with the alteration of one amino acid and provide

  2. SAM domain-dependent activity of PfTKL3, an essential tyrosine kinase-like kinase of the human malaria parasite Plasmodium falciparum.

    Science.gov (United States)

    Abdi, Abdirahman; Eschenlauer, Sylvain; Reininger, Luc; Doerig, Christian

    2010-10-01

    Over the last decade, several protein kinases inhibitors have reached the market for cancer chemotherapy. The kinomes of pathogens represent potentially attractive targets in infectious diseases. The functions of the majority of protein kinases of Plasmodium falciparum, the parasitic protist responsible for the most virulent form of human malaria, remain unknown. Here we present a thorough characterisation of PfTKL3 (PF13_0258), an enzyme that belongs to the tyrosine kinase-like kinase (TKL) group. We demonstrate by reverse genetics that PfTKL3 is essential for asexual parasite proliferation in human erythrocytes. PfTKL3 is expressed in both asexual and gametocytes stages, and in the latter the protein co-localises with cytoskeleton microtubules. Recombinant PfTKL3 displays in vitro autophosphorylation activity and is able to phosphorylate exogenous substrates, and both activities are dramatically dependent on the presence of an N-terminal "sterile alpha-motif" domain. This study identifies PfTKL3 as a validated drug target amenable to high-throughput screening.

  3. Long ranging swept-source optical coherence tomography-based angiography outperforms its spectral-domain counterpart in imaging human skin mi