Sample records for calcium-binding proteins

  1. Fractal aspects of calcium binding protein structures

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    Isvoran, Adriana [West University of Timisoara, Department of Chemistry, Pestalozzi 16, 300115 Timisoara (Romania)], E-mail:; Pitulice, Laura [West University of Timisoara, Department of Chemistry, Pestalozzi 16, 300115 Timisoara (Romania); Craescu, Constantin T. [INSERM U759/Institute Curie-Recherche, Centre Universitaire Paris-Sud, Batiment 112, 91405 Orsay (France); Chiriac, Adrian [West University of Timisoara, Department of Chemistry, Pestalozzi 16, 300115 Timisoara (Romania)


    The structures of EF-hand calcium binding proteins may be classified into two distinct groups: extended and compact structures. In this paper we studied 20 different structures of calcium binding proteins using the fractal analysis. Nine structures show extended shapes, one is semi-compact and the other 10 have compact shapes. Our study reveals different fractal characteristics for protein backbones belonging to different structural classes and these observations may be correlated to the physicochemical forces governing the protein folding.

  2. Apolipoprotein B is a calcium binding protein

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    Dashti, N.; Lee, D.M.; Mok, T.


    Human hepatocarcinoma Hep G2 cells were grown in culture medium containing (/sup 45/Ca/sup 2 +/). The secreted lipoproteins of d < 1.063 g/ml and d 1.063-1.21 g/ml were isolated from the culture media and analyzed by 3.3% and 7% SDS-polyacrylamide gel electrophoresis. Radioactivity profiles of (/sup 45/Ca) from the gels showed that the peak of radioactivity corresponded to the apolipoprotein B band. The molar ratio of the incorporated (/sup 45/Ca/sup 2 +/) and apolipoprotein B was close to unity. No radioactivity was found associated with any other secreted apolipoproteins. To confirm these findings, apolipoprotein B-containing lipoproteins were precipitated with anti-apolipoprotein B and high density lipoproteins were precipitated with anti-apolipoprotein A-I. Only the former precipitate was radioactive. These results suggest that apolipoprotein B is a calcium binding protein.

  3. Calcium-binding proteins from human platelets

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    Gogstad, G.O.; Krutnes, M.B.; Solum, N.O.


    Calcium-binding platelet proteins were examined by crossed immunoelectrophoresis of solubilized platelets against antibodies to whole platelets followed by incubation of the immunoplates with /sup 45/Ca/sup 2 +/ and autoradiography. When the immunoplates had been pretreated with EDTA at pH 9.0 in order to remove divalent cations, three immunoprecipitates were markedly labelled with /sup 45/Ca/sup 2 +/. These corresponded to the glycoprotein IIb-IIIa complex, glycoprotein Ia and a presently unidentified antigen termed G18. These antigens were membrane-bound and surface-oriented. When an excess of EDTA was introduced in the incubation media the results revealed that the glycoprotein IIb-IIIa complex and antigen G18, but not glycoprotein Ia, contained sites with a stronger affinity for calcium than has EDTA at pH 7.4. Immunoprecipitates of the separate glycoproteins IIb and IIIa both bound calcium in the same manner as the glycoprotein IIb-IIIa complex. As another approach, platelet-rich plasma was incubated with /sup 45/Ca/sup 2 +/ prior to crossed immunoelectrophoresis of the solubilized platelets. A single immunoprecipitate was weakly labelled. This did not correspond to any of the immunoprecipitates which were visible after staining with Coomassie blue. The labelling of this antigen was markedly increased when the platelet-rich plasma had been preincubated with EDTA and in this case a weak labelling of the glycoprotein IIB-IIIa precipitate also became apparent. No increased incorporation of calcium occured in any of these immunoprecipitates when the platelets were aggregated with ADP in the presence of /sup 45/Ca/sup 2 +/.

  4. ALG-2, a multifunctional calcium binding protein?

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    Tarabykina, Svetlana; Mollerup, Jens; Winding Gojkovic, P.


    ALG-2 was originally discovered as a pro-apoptotic protein in a genetic screen. Due to its ability to bind calcium with high affinity it was postulated to provide a link between the known effect of calcium in programmed cell death and the molecular death execution machinery. This review article...

  5. Calcium-binding properties of a calcium-dependent protein kinase from Plasmodium falciparum and the significance of individual calcium-binding sites for kinase activation. (United States)

    Zhao, Y; Pokutta, S; Maurer, P; Lindt, M; Franklin, R M; Kappes, B


    Calcium-dependent protein kinase from Plasmodium falciparum (PfCPK) is a multidomain protein composed of an N-terminal kinase domain connected via a linker region to a C-terminal CaM-like calcium-binding domain. The kinase can be activated by Ca2+ alone and associates with 45Ca2+. Here we describe the calcium-binding properties of the kinase and the significance of the individual calcium-binding sites with respect to enzymatic activation, as well as the Ca(2+)-induced conformational change as detected by circular dichroism. As predicted from the cDNA sequence, the kinase has four EF-hand calcium-binding sites in the C-terminal domain. To understand the roles of the individual calcium-binding sites, two series of mutations were generated at the individual EF-hand motifs. The highly conserved glutamic acid residue at position 12 in each calcium-binding loop was mutated to either lysine or glutamine, and therefore a total of eight mutants were generated. Either of these mutations (to lysine or glutamine) is sufficient to eliminate calcium binding at the mutated site. Sites I and II appear to be crucial for both Ca(2+)-induced conformational change and enzymatic activation. Whereas mutations at site II almost completely abolish kinase activity, mutations at site I are also deleterious and dramatically reduce the sensitivity of the Ca(2+)-induced conformational change and the Ca(2+)-dependent activation. Mutations at sites III and IV have minor effects.

  6. Neutrophils and the calcium-binding protein MRP-14 mediate carrageenan-induced antinociception in mice

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    Rosana L. Pagano


    Full Text Available Background: We have previously shown that the calcium-binding protein MRP-14 secreted by neutrophils mediates the antinociceptive response in an acute inflammatory model induced by the intraperitoneal injection of glycogen in mice.

  7. Distribution of calcium-binding proteins in the chick visual system

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    C.P. Pfeiffer


    Full Text Available The calcium-binding proteins calbindin (CB, calretinin (CR, and parvalbumin (PV have been extensively studied over the last decade since they appear to be important as buffers of intracellular calcium. In the present study we investigated the distribution of these proteins in the chick visual system by means of conventional immunocytochemistry. The results indicated that CB, CR, and PV are widely distributed in retinorecipient areas of the chick brain. In some regions, all three calcium-binding proteins were present at different intensities and often in different neurons such as in the dorsolateral thalamic complex. In other areas, such as the nucleus geniculatus lateralis ventralis, only CB and CR were detected, whereas PV was absent. These results show that these three calcium-binding proteins are differentially distributed in the visual system of the chick, with varying degrees of co-localization

  8. Exploring NMR ensembles of calcium binding proteins: Perspectives to design inhibitors of protein-protein interactions

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    Craescu Constantin T


    Full Text Available Abstract Background Disrupting protein-protein interactions by small organic molecules is nowadays a promising strategy employed to block protein targets involved in different pathologies. However, structural changes occurring at the binding interfaces make difficult drug discovery processes using structure-based drug design/virtual screening approaches. Here we focused on two homologous calcium binding proteins, calmodulin and human centrin 2, involved in different cellular functions via protein-protein interactions, and known to undergo important conformational changes upon ligand binding. Results In order to find suitable protein conformations of calmodulin and centrin for further structure-based drug design/virtual screening, we performed in silico structural/energetic analysis and molecular docking of terphenyl (a mimicking alpha-helical molecule known to inhibit protein-protein interactions of calmodulin into X-ray and NMR ensembles of calmodulin and centrin. We employed several scoring methods in order to find the best protein conformations. Our results show that docking on NMR structures of calmodulin and centrin can be very helpful to take into account conformational changes occurring at protein-protein interfaces. Conclusions NMR structures of protein-protein complexes nowadays available could efficiently be exploited for further structure-based drug design/virtual screening processes employed to design small molecule inhibitors of protein-protein interactions.

  9. Cloning, purification, crystallization and preliminary crystallographic study of calcium-binding protein 5 from Entamoeba histolytica. (United States)

    Kumar, Sanjeev; Zaidi, Rana; Gourinath, Samudrala


    Entamoeba histolytica is the causative agent of human amoebiasis. Phagocytosis is the major route of food intake by this parasite and is responsible for its virulence. Calcium and calcium-binding proteins play major roles in its phagocytosis. Calcium-binding protein 5 from E. histolytica (EhCaBP5) is a cytoplasmic protein; its expression is very sensitive to serum starvation and it seems to be involved in binding to myosin I. In this study, EhCaBP5 was cloned, expressed in Escherichia coli and purified using affinity and size-exclusion chromatography. The purified protein crystallized in space group C222 and the crystals diffracted to 2 Å resolution. The Matthews coefficient indicated the presence of one molecule in the asymmetric unit, with a VM of 2.35 Å3 Da(-1) and a solvent content of 47.7%.

  10. Isolation, expression and immunological characterization of a calcium-binding protein from Parietaria pollen. (United States)

    Bonura, A; Gulino, L; Trapani, A; Di Felice, G; Tinghino, R; Amoroso, S; Geraci, D; Valenta, R; Westritschnig, K; Scala, E; Mari, A; Colombo, P


    The diagnosis and therapy of allergic disorders are usually performed with crude extracts which are a heterogeneous mixture of proteins with different allergenic potency. The knowledge of the allergenic composition is a key step for diagnostic and therapeutic options. Parietaria judaica pollen represents one of the main sources of allergens in the Mediterranean area and its major allergens have already been identified (Par j 1 and Par j 2). In addition, inhibition studies performed using a calcium-binding protein (CBP) from grass pollen (Phl p 7) showed the presence of a homologue of this cross-reactive allergen in the Parietaria extract. Screening of a cDNA library allowed us to isolate a 480bp cDNA containing the information for an 87 AA long protein with high level of homology to calcium-binding proteins from other allergenic sources. It was expressed as a recombinant allergen in Escherichia coli and purified by affinity chromatography. Its expression allowed us to study the prevalence of this allergen in a population of allergic patients in southern Europe. Immunoblotting and inhibition studies showed that this allergen shares a pattern of IgE epitopes in common with other 2-EF-hand calcium-binding proteins from botanically non-related species. The immunological properties of the Pj CBP were investigated by CD63 activation assay and CFDA-SE staining. In conclusion, DNA recombinant technology allowed the isolation, expression and immunological characterization of a cross-reactive calcium-binding protein allergen from Parietaria judaica pollen.

  11. Identification of calcium-binding proteins associated with the human sperm plasma membrane

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    Westbrook Anne


    Full Text Available Abstract Background The precise composition of the human sperm plasma membrane, the molecular interactions that define domain specific functions, and the regulation of membrane associated proteins during the capacitation process, still remain to be fully understood. Here, we investigated the repertoire of calcium-regulated proteins associated with the human sperm plasma membrane. Methods Surface specific radioiodination was combined with two-dimensional gel electrophoresis, a 45Ca-overlay assay, computer assisted image analysis and mass spectrometry to identify calcium-binding proteins exposed on the human sperm surface. Results Nine acidic 45Ca-binding sperm proteins were excised from stained preparative 2D gels and identified by mass spectrometry. Five of the calcium binding proteins; HSPA2 (HSP70-1, HSPA5 (Bip, HYOU1 (ORP150, serum amyloid P-component (SAP and protein kinase C substrate 80K-H (80K-H were found to be accessible to Iodo-Bead catalyzed 125I-labelling on the surface of intact human sperm. Agglutination and immunofluorescence analysis confirmed that SAP is situated on the plasma membrane of intact, motile sperm as well as permeabilized cells. Western blot analysis showed increased phosphorylation of human sperm 80K-H protein following in vitro capacitation. This is the first demonstration of the 80K-H protein in a mammalian sperm. Conclusion The presence of SAP on the surface of mature sperm implies that SAP has a physiological role in reproduction, which is thought to be in the removal of spermatozoa from the female genital tract via phagocytosis. Since 80K-H is a Ca2+-sensor recently implicated in the regulation of both inositol 1,4,5-trisphosphate receptor and transient receptor potential (TRP cation channel activities, its detection in sperm represents the first direct signaling link between PKC and store-operated calcium channels identified in human sperm.

  12. Calcium-binding proteins in the laterodorsal thalamic nucleus during development of the guinea pig. (United States)

    Zakowski, Witold; Bogus-Nowakowska, Krystyna; Wasilewska, Barbara; Hermanowicz, Beata; Robak, Anna


    The laterodorsal thalamic nucleus (LD) is often treated as a part of the anterior thalamic nuclei (ATN) because of its location and similar connectivity. Our previous studies have shown that distribution of three calcium-binding proteins, i.e. calbindin D28k (CB), calretinin (CR) and parvalbumin (PV), changes within the ATN during development of the guinea pig. The aim of this study is to examine the immunoreactivity pattern of these proteins in the LD in the guinea pig ontogeny. Brains from animals ranging from 40th embryonic day to 80th postnatal day were used in the study. Two methods were applied: a single-labelling immunoenzymatic method and double-labelling immunofluorescence. No changes of the distribution pattern of the substances were observed throughout the examined developmental stages. CB and CR were the most abundantly expressed proteins in perikarya of the LD. Numerous CB- and CR-immunoreactive cell bodies were found throughout the whole extent of the nucleus. In most of these cell bodies both proteins colocalized vastly. The highest immunoreactivity of the perikarya containing CB and CR was observed in the mediodorsal part of the LD and in its rostral portion. In regard to PV, single cell bodies were observed mostly in the dorsal part of the nucleus. PV did not colocalize with the other proteins. In summary, all the studied calcium-binding proteins were already present in the LD at prenatal developmental stages and the pattern of distribution remained virtually constant until adulthood. Thus, the LD differs considerably from the ATN in an aspect of neurochemical cell differentiation. Copyright © 2014 Elsevier B.V. All rights reserved.

  13. 1H, 13C and 15N NMR assignments of a calcium-binding protein from Entamoeba histolytica. (United States)

    Verma, Deepshikha; Bhattacharya, Alok; Chary, Kandala V R


    We report almost complete sequence specific (1)H, (13)C and (15)N NMR assignments of a 150-residue long calmodulin-like calcium-binding protein from Entamoeba histolytica (EhCaBP6), as a prelude to its structural and functional characterization.

  14. Distribution and Morphology of Calcium-Binding Proteins Immunoreactive Neurons following Chronic Tungsten Multielectrode Implants.

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    Marco Aurelio M Freire

    Full Text Available The development of therapeutic approaches to improve the life quality of people suffering from different types of body paralysis is a current major medical challenge. Brain-machine interface (BMI can potentially help reestablishing lost sensory and motor functions, allowing patients to use their own brain activity to restore sensorimotor control of paralyzed body parts. Chronic implants of multielectrodes, employed to record neural activity directly from the brain parenchyma, constitute the fundamental component of a BMI. However, before this technique may be effectively available to human clinical trials, it is essential to characterize its long-term impact on the nervous tissue in animal models. In the present study we evaluated how chronic implanted tungsten microelectrode arrays impact the distribution and morphology of interneurons reactive to calcium-binding proteins calbindin (CB, calretinin (CR and parvalbumin (PV across the rat's motor cortex. Our results revealed that chronic microelectrode arrays were well tolerated by the nervous tissue, with recordings remaining viable for up to 6 months after implantation. Furthermore, neither the morphology nor the distribution of inhibitory neurons were broadly impacted. Moreover, restricted microglial activation was observed on the implanted sites. On the whole, our results confirm and expand the notion that tungsten multielectrodes can be deemed as a feasible candidate to future human BMI studies.

  15. Auditory hindbrain atrophy and anomalous calcium binding protein expression after neonatal exposure to monosodium glutamate. (United States)

    Foran, Lindsey; Blackburn, Kaitlyn; Kulesza, Randy J


    Glutamate is the most abundant excitatory neurotransmitter in the central nervous system, and is stored and released by both neurons and astrocytes. Despite the important role of glutamate as a neurotransmitter, elevated extracellular glutamate can result in excitotoxicity and apoptosis. Monosodium glutamate (MSG) is a naturally occurring sodium salt of glutamic acid that is used as a flavor enhancer in many processed foods. Previous studies have shown that MSG administration during the early postnatal period results in neurodegenerative changes in several forebrain regions, characterized by neuronal loss and neuroendocrine abnormalities. Systemic delivery of MSG during the neonatal period and induction of glutamate neurotoxicity in the cochlea have both been shown to result in fewer neurons in the spiral ganglion. We hypothesized that an MSG-induced loss of neurons in the spiral ganglion would have a significant impact on the number of neurons in the cochlear nuclei and superior olivary complex (SOC). Indeed, we found that exposure to MSG from postnatal days 4 through 10 resulted in significantly fewer neurons in the cochlear nuclei and SOC and significant dysmorphology in surviving neurons. Moreover, we found that neonatal MSG exposure resulted in a significant decrease in the expression of both calretinin and calbindin. These results suggest that neonatal exposure to MSG interferes with early development of the auditory brainstem and impacts expression of calcium binding proteins, both of which may lead to diminished auditory function. Copyright © 2017 IBRO. Published by Elsevier Ltd. All rights reserved.

  16. A Specific Peptide with Calcium-Binding Capacity from Defatted Schizochytrium sp. Protein Hydrolysates and the Molecular Properties

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    Xixi Cai


    Full Text Available Marine microorganisms have been proposed as a new kind of protein source. Efforts are needed in order to transform the protein-rich biological wastes left after lipid extraction into value-added bio-products. Thus, the utilization of protein recovered from defatted Schizochytrium sp. by-products presents an opportunity. A specific peptide Tyr-Leu (YL with calcium-binding capacity was purified from defatted Schizochytrium sp. protein hydrolysates through gel filtration chromatography and RP-HPLC. The calcium-binding activity of YL reached 126.34 ± 3.40 μg/mg. The calcium-binding mechanism was investigated through ultraviolet, fluorescence and infrared spectroscopy. The results showed that calcium ions could form dative bonds with carboxyl oxygen atoms and amino nitrogen atoms as well as the nitrogen and oxygen atoms of amide bonds. YL-Ca exhibited excellent thermal stability and solubility, which was beneficial for its absorption and transport in the basic intestinal tract of the human body. Moreover, the cellular uptake of calcium in Caco-2 cells showed that YL-Ca could enhance calcium uptake efficiency and protect calcium ions against precipitation caused by dietary inhibitors such as tannic acid, oxalate, phytate and metal ions. The findings indicate that the by-product of Schizochytrium sp. is a promising source for making peptide-calcium bio-products as algae-based functional supplements for human beings.

  17. A Specific Peptide with Calcium-Binding Capacity from Defatted Schizochytrium sp. Protein Hydrolysates and the Molecular Properties. (United States)

    Cai, Xixi; Yang, Qian; Lin, Jiaping; Fu, Nanyan; Wang, Shaoyun


    Marine microorganisms have been proposed as a new kind of protein source. Efforts are needed in order to transform the protein-rich biological wastes left after lipid extraction into value-added bio-products. Thus, the utilization of protein recovered from defatted Schizochytrium sp. by-products presents an opportunity. A specific peptide Tyr-Leu (YL) with calcium-binding capacity was purified from defatted Schizochytrium sp. protein hydrolysates through gel filtration chromatography and RP-HPLC. The calcium-binding activity of YL reached 126.34 ± 3.40 μg/mg. The calcium-binding mechanism was investigated through ultraviolet, fluorescence and infrared spectroscopy. The results showed that calcium ions could form dative bonds with carboxyl oxygen atoms and amino nitrogen atoms as well as the nitrogen and oxygen atoms of amide bonds. YL-Ca exhibited excellent thermal stability and solubility, which was beneficial for its absorption and transport in the basic intestinal tract of the human body. Moreover, the cellular uptake of calcium in Caco-2 cells showed that YL-Ca could enhance calcium uptake efficiency and protect calcium ions against precipitation caused by dietary inhibitors such as tannic acid, oxalate, phytate and metal ions. The findings indicate that the by-product of Schizochytrium sp. is a promising source for making peptide-calcium bio-products as algae-based functional supplements for human beings.

  18. Scaling properties of the radius of gyration and surface area for EF-hand calcium binding proteins

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    Pitulice, L. [West University of Timisoara, Department of Chemistry, Pestalozzi 16, 300115 Timisoara (Romania); Isvoran, A. [West University of Timisoara, Department of Chemistry, Pestalozzi 16, 300115 Timisoara (Romania)], E-mail:; Craescu, C.T. [INSERM U759/Institute Curie-Recherche, Centre Universitaire Paris-Sud, Batiment 112, 91405 Orsay (France); Chiriac, A. [West University of Timisoara, Department of Chemistry, Pestalozzi 16, 300115 Timisoara (Romania)


    In this paper, we analyze the scaling properties of both the radius of gyration and the surface area for EF-hand calcium binding proteins. These properties are different for two conformational subfamilies: proteins with extended and compact structures, respectively. The radius of gyration is a measure of the shape of protein, whereas its surface fractal dimension is a measure of its interatomic packing. Different scaling properties for the radius of gyration underline that these two subfamilies present different shapes whilst different scaling properties for the surface area reveal different strengths of their intermolecular forces. All these data suggest different mechanisms responsible for the global folding of proteins belonging to these two subfamilies.

  19. The calcium-binding protein of Entamoeba histolytica as a fusion partner for expression of peptides in Escherichia coli. (United States)

    Reddi, Honey; Bhattacharya, Alok; Kumar, Vijay


    We describe the construction of an Escherichia coli expression vector, CBP that allows the C-terminal fusion of heterologous proteins to the calcium-binding protein (CaBP) of the parasitic protozoan Entamoeba histolytica. The intrinsic nature of this protein to remain soluble on heat treatment has been exploited in its use as a novel fusion partner. The presence of a histidine tag and an enterokinase recognition site, aid in the affinity purification and proteolytic cleavage of the fusion protein. The efficacy of the vector was tested using the preS1 region of the envelope protein of the hepatitis B virus. The CaBP-preS1 fusion protein partitioned in the soluble fraction on heat treatment and this facilitated its rapid purification.

  20. APF/CBP, the small, amphipathic, anionic protein(s) in bile and gallstones, consists of lipid-binding and calcium-binding forms

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    Lafont, H.; Domingo, N.; Groen, A.; Kaler, E. W.; Lee, S. P.; Koehler, R.; Ostrow, J. D.; Veis, A.


    Two very similar small anionic, amphipathic proteins, a phospholipid-binding apoprotein (anionic polypeptide fraction [APF]) and a calcium-binding polypeptide (CBP), are found abundantly in bile and all types of gallstones. The often disparate properties among various preparations of APF/CBP could

  1. Portraying the Effect of Calcium-Binding Proteins on Cytosolic Calcium Concentration Distribution Fractionally in Nerve Cells. (United States)

    Jha, Brajesh Kumar; Joshi, Hardik; Dave, Devanshi D


    Nerve cells like neurons and astrocytes in central nervous system (CNS) take part in the signaling process which means the transformation of the information from one cell to another via signals. The signaling process is affected by various external parameters like buffers calcium-binding proteins, voltage-gated calcium channel. In the present paper, the role of buffers in the cytoplasmic calcium concentration distribution is shown. The elicitation in calcium concentration is due to the presence of lower amount calcium-binding proteins which can be shown graphically. The mathematical model is designed by keeping in mind the physiological condition taking place in CNS of mammalian brain. The thing to be noted here is that the more elicitation in the calcium concentration distribution results in the cell death which finally give neurodegenerative disease to the mammalian brain. The present paper gives a glimpse of Parkinson's diseases in particular. Computational results are performed in Wolfram Mathematica 9.0 and simulated on core(TM) i5-3210M CPU @ 2.50 GHz processing speed and 4 GB memory. It is found that the different types of buffer like ethylene glycol-bis([Formula: see text]-aminoethyl ether)-N,N,N',N'-tetraacetic acid, 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid and calmodulin have noteworthy effect at different fractions of time.

  2. Novel Peptide with Specific Calcium-Binding Capacity from Schizochytrium sp. Protein Hydrolysates and Calcium Bioavailability in Caco-2 Cells

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    Xixi Cai


    Full Text Available Peptide-calcium can probably be a suitable supplement to improve calcium absorption in the human body. In this study, a specific peptide Phe-Tyr (FY with calcium-binding capacity was purified from Schizochytrium sp. protein hydrolysates through gel filtration chromatography and reversed phase HPLC. The calcium-binding capacity of FY reached 128.77 ± 2.57 μg/mg. Results of ultraviolet spectroscopy, fluorescence spectroscopy, and infrared spectroscopy showed that carboxyl groups, amino groups, and amido groups were the major chelating sites. FY-Ca exhibited excellent thermal stability and solubility, which were beneficial to be absorbed and transported in the basic intestinal tract of the human body. Moreover, the calcium bioavailability in Caco-2 cells showed that FY-Ca could enhance calcium uptake efficiency by more than three times when compared with CaCl2, and protect calcium ions against dietary inhibitors, such as tannic acid, oxalate, phytate, and Zn2+. Our findings further the progress of algae-based peptide-calcium, suggesting that FY-Ca has the potential to be developed as functionally nutraceutical additives.

  3. Abnormal neuronal expression of the calcium-binding proteins, parvalbumin and calbindin D-28k, in aged dogs. (United States)

    Sisó, S; Tort, S; Aparici, C; Pérez, L; Vidal, E; Pumarola, M


    Disturbances of the gamma-aminobutyric acid (GABA) neurotransmitter system have been implicated in chronic degenerative neurological disease. Cognitive dysfunction and neuron loss are features in older dogs. GABAergic neurons also show immunoreactivity for specific calcium-binding proteins. Immunohistochemistry was used to study the neuronal expression of calbindin D-28k and parvalbumin in different areas of the brain in 13 dogs, aged between 2 and 13.5 years. Calbindin expression was found only in the cerebellum. There were significant differences in the quantity and distribution of neurons expressing these proteins between geriatric and adult brains. Parvalbumin- and calbindin-expressing neurons are relatively sensitive to degeneration in the cerebellum of older dogs. Parvalbumin labelling was associated with dystrophic structures that are commonly associated with ageing. Copyright 2002 Elsevier Science Ltd.

  4. The calcium binding protein ALG-2 binds and stabilizes Scotin, a p53-inducible gene product localized at the endoplasmic reticulum membrane

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    Draeby, Ingrid; Woods, Yvonne L; la Cour, Jonas Marstrand


    ALG-2 (apoptosis linked gene 2 product) is a calcium binding protein for which no clear cellular function has been established. In this study we identified Scotin as a novel ALG-2 target protein containing 6 PXY and 4 PYP repeats, earlier identified in the ALG-2 binding regions of AIP1/ALIX and TSG...

  5. The calcium-binding protein complex S100A8/A9 has a crucial role in controlling macrophage-mediated renal repair following ischemia/reperfusion

    NARCIS (Netherlands)

    Dessing, M.C.; Tammaro, A.; Pulskens, W.P.C.; Teske, G.J.; Butter, L.M.; Claessen, N.; Eijk, M. van; Poll, T. van der; Vogl, T.; Roth, J.; Florquin, S.; Leemans, J.C.


    Upon ischemia/reperfusion (I/R)-induced injury, several damage-associated molecular patterns are expressed including the calcium-binding protein S100A8/A9 complex. S100A8/A9 can be recognized by Toll-like receptor-4 and its activation is known to deleteriously contribute to renal I/R-induced injury.

  6. Comparative anatomical distribution of neuronal calcium-binding protein (NECAB) 1 and -2 in rodent and human spinal cord. (United States)

    Zhang, Ming-Dong; Barde, Swapnali; Szodorai, Edit; Josephson, Anna; Mitsios, Nicholas; Watanabe, Masahiko; Attems, Johannes; Lubec, Gert; Kovács, Gábor G; Uhlén, Mathias; Mulder, Jan; Harkany, Tibor; Hökfelt, Tomas


    Neuronal calcium-binding protein 1 and -2 (NECAB1/2) localize to multiple excitatory neuron populations in the mouse spinal cord. Here, we analyzed rat and human spinal cord, combining in situ hybridization and immunohistochemistry, complementing newly collated data on mouse spinal cord for direct comparisons. Necab1/2 mRNA transcripts showed complementary distribution in rodent's spinal cord. Multiple-labeling fluorescence histochemistry with neuronal phenotypic markers localized NECAB1 to a dense fiber plexus in the dorsal horn, to neurons mainly in superficial layers and to commissural interneurons in both rodent species. NECAB1-positive (+) motor neurons were only found in mice. NECAB1 distribution in the human spinal cord was similar with the addition of NECAB1-like immunoreactivity surrounding myelinated axons. NECAB2 was mainly present in excitatory synaptic boutons in the dorsal horn of all three species, and often in calbindin-D28k(+) neuronal somata. Rodent ependymal cells expressed calbindin-D28k. In humans, they instead were NECAB2(+) and/or calretinin(+). Our results reveal that the association of NECAB2 to excitatory neuronal circuits in the spinal cord is evolutionarily conserved across the mammalian species investigated so far. In contrast, NECAB1 expression is more heterogeneous. Thus, our study suggests that the phenotypic segregation of NECAB1 and -2 to respective excitatory and inhibitory spinal systems can underpin functional modalities in determining the fidelity of synaptic neurotransmission and neuronal responsiveness, and might bear translational relevance to humans.

  7. Subdivisions of the auditory midbrain (n. mesencephalicus lateralis, pars dorsalis in zebra finches using calcium-binding protein immunocytochemistry.

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    Priscilla Logerot

    Full Text Available The midbrain nucleus mesencephalicus lateralis pars dorsalis (MLd is thought to be the avian homologue of the central nucleus of the mammalian inferior colliculus. As such, it is a major relay in the ascending auditory pathway of all birds and in songbirds mediates the auditory feedback necessary for the learning and maintenance of song. To clarify the organization of MLd, we applied three calcium binding protein antibodies to tissue sections from the brains of adult male and female zebra finches. The staining patterns resulting from the application of parvalbumin, calbindin and calretinin antibodies differed from each other and in different parts of the nucleus. Parvalbumin-like immunoreactivity was distributed throughout the whole nucleus, as defined by the totality of the terminations of brainstem auditory afferents; in other words parvalbumin-like immunoreactivity defines the boundaries of MLd. Staining patterns of parvalbumin, calbindin and calretinin defined two regions of MLd: inner (MLd.I and outer (MLd.O. MLd.O largely surrounds MLd.I and is distinct from the surrounding intercollicular nucleus. Unlike the case in some non-songbirds, however, the two MLd regions do not correspond to the terminal zones of the projections of the brainstem auditory nuclei angularis and laminaris, which have been found to overlap substantially throughout the nucleus in zebra finches.

  8. Neuroprotective Effect of Ginseng against Alteration of Calcium Binding Proteins Immunoreactivity in the Mice Hippocampus after Radiofrequency Exposure

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    Dhiraj Maskey


    Full Text Available Calcium binding proteins (CaBPs such as calbindin D28-k, parvalbumin, and calretinin are able to bind Ca2+ with high affinity. Changes in Ca2+ concentrations via CaBPs can disturb Ca2+ homeostasis. Brain damage can be induced by the prolonged electromagnetic field (EMF exposure with loss of interacellular Ca2+ balance. The present study investigated the radioprotective effect of ginseng in regard to CaBPs immunoreactivity (IR in the hippocampus through immunohistochemistry after one-month exposure at 1.6 SAR value by comparing sham control with exposed and ginseng-treated exposed groups separately. Loss of dendritic arborization was noted with the CaBPs in the Cornu Ammonis areas as well as a decrease of staining intensity of the granule cells in the dentate gyrus after exposure while no loss was observed in the ginseng-treated group. A significant difference in the relative mean density was noted between control and exposed groups but was nonsignificant in the ginseng-treated group. Decrease in CaBP IR with changes in the neuronal staining as observed in the exposed group would affect the hippocampal trisynaptic circuit by alteration of the Ca2+ concentration which could be prevented by ginseng. Hence, ginseng could contribute as a radioprotective agent against EMF exposure, contributing to the maintenance of Ca2+ homeostasis by preventing impairment of intracellular Ca2+ levels in the hippocampus.

  9. The calcium-binding protein S100P in normal and malignant human tissues

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    Pastorek Jaromir


    Full Text Available Abstract Background S100P is a Ca2+ binding protein overexpressed in a variety of cancers, and thus, has been considered a potential tumor biomarker. Very little has been studied about its normal expression and functions. Methods We examined S100P expression in normal human tissues by quantitative reverse transcription polymerase chain reaction and immunohistochemistry. S100P protein expression was also studied in a series of tumors, consisting of 74 ovarian, 11 pancreatic, 56 gastric, 57 colorectal, 89 breast and 193 prostate carcinomas using a novel anti-S100P monoclonal antibody. Results Among the normal tissues, the highest S100P mRNA levels were observed in the placenta and esophagus. Moderate signals were also detected in the stomach, duodenum, large intestine, prostate and leukocytes. At the protein level, the highest reactions for S100P were seen in the placenta and stomach. Immunostaining of tumor specimens showed that S100P protein is expressed in all the tumor categories included in the study, being most prevalent in gastric tumors. Conclusion Based on our observations, S100P is widely expressed in both normal and malignant tissues. The high expression in some tumors suggests that it may represent a potential target molecule for future diagnostic and therapeutic applications.

  10. Ectopic expression of the calcium-binding protein parvalbumin in mouse liver endothelial cells

    DEFF Research Database (Denmark)

    Castillo, M B; Berchtold, M W; Rülicke, T


    To elucidate the physiological role of the Ca2+ binding protein parvalbumin, we have generated transgenic mice carrying the full-length complementary DNA (cDNA) of rat parvalbumin under the control of the heavy-metal inducible metallothionein IIA promoter. Immunohistochemical and biochemical...... vasoconstriction via calcium signalling, were investigated in the mouse liver perfused in situ. Vasoconstriction, thought to be mediated by the Ito cell, was not affected in the transgenic animals, whereas microvascular exchange, probed with the multiple indicator dilution technique, was markedly decreased...

  11. Structure and self-assembly of the calcium binding matrix protein of human metapneumovirus. (United States)

    Leyrat, Cedric; Renner, Max; Harlos, Karl; Huiskonen, Juha T; Grimes, Jonathan M


    The matrix protein (M) of paramyxoviruses plays a key role in determining virion morphology by directing viral assembly and budding. Here, we report the crystal structure of the human metapneumovirus M at 2.8 Å resolution in its native dimeric state. The structure reveals the presence of a high-affinity Ca²⁺ binding site. Molecular dynamics simulations (MDS) predict a secondary lower-affinity site that correlates well with data from fluorescence-based thermal shift assays. By combining small-angle X-ray scattering with MDS and ensemble analysis, we captured the structure and dynamics of M in solution. Our analysis reveals a large positively charged patch on the protein surface that is involved in membrane interaction. Structural analysis of DOPC-induced polymerization of M into helical filaments using electron microscopy leads to a model of M self-assembly. The conservation of the Ca²⁺ binding sites suggests a role for calcium in the replication and morphogenesis of pneumoviruses. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

  12. Relating form and function of EF-hand calcium binding proteins. (United States)

    Chazin, Walter J


    The EF hand, a helix-loop-helix structure, is one of the most common motifs found in animal genomes, and EF-hand Ca(2+)-binding proteins (EFCaBPs) are widely distributed throughout the cell. However, researchers remain confounded by a lack of understanding of how peptide sequences code for specific functions and by uncertainty about the molecular mechanisms that enable EFCaBPs to distinguish among many diverse cellular targets. Such knowledge could define the roles of EFCaBPs in health and disease and ultimately enable control or even design of Ca(2+)-dependent functions in medicine and biotechnology. In this Account, we describe our structural and biochemical research designed to understand the sequence-to-function relationship in EFCaBPs. The first structural goal was to define conformational changes induced by binding Ca(2+), and our group and others established that solution NMR spectroscopy is well suited for this task. We pinpointed residues critical to the differences in Ca(2+) response of calbindin D(9k) and calmodulin (CaM), homologous EFCaBPs from different functional classes, by using direct structure determination with site-directed mutagenesis and protein engineering. Structure combined with biochemistry provided the foundation for identifying the fundamental mechanism of cooperativity in the binding of Ca(2+) ions: this cooperativity provides EFCaBPs with the ability to detect the relatively small changes in concentration that constitute Ca(2+) signals. Using calbindin D(9k) as a model system, studies of the structure and fast time scale dynamics of each of the four ion binding states in a typical EF-hand domain provided direct evidence that site-site communication lowers the free energy cost of reorganization for binding the second ion. Our work has also extended models of how EFCaBPs interact with their cellular targets. We determined the unique dimeric architecture of S100 proteins, a specialized subfamily of EFCaBPs found exclusively in

  13. Identification of calconectin, a calcium-binding protein specifically expressed by the mantle of Pinctada margaritifera. (United States)

    Duplat, D; Puisségur, M; Bédouet, L; Rousseau, M; Boulzaguet, H; Milet, C; Sellos, D; Van Wormhoudt, A; Lopez, E


    Nacre or mother-of-pearl in the shell of Pinctada margaritifera is composed of 95-99% calcium carbonate and 1-5% organic matrix. In this study, we developed an original technique to characterize the genes differentially expressed in nacre-forming cells (NFC) by combining suppression subtractive hybridization (SSH), to establish a cDNA subtractive library, with rapid amplification of cDNA ends (RACE)-PCR. Seventy-two specific cDNA sequences have been obtained so far. These include a protein containing two EF-hand Ca2+-binding domains which was completely sequenced after amplification by RACE-PCR. Its specific expression as well as the specificity of the SSH method was confirmed by semi-quantitative RT-PCR on NFC and mantle cells.

  14. Expression of the calcium-binding proteins MRP8 and MRP14 in monocytes is regulated by a calcium-induced suppressor mechanism.


    ROTH, J.; Goebeler, M; Wrocklage, V; van den Bos, C; Sorg, C.


    MRP8 and MRP14 are two calcium-binding proteins of the S-100 family the expression of which is restricted to distinct stages of monocytic differentiation. Heteromeric MRP8/MRP14 complexes have been shown to represent their biologically active forms. However, it is not as yet clear whether biochemical modification of complexes, or regulation on the transcriptional level, are responsible for the control of MRP8/MRP14 expression. Employing Western-blot analysis and metabolic labelling we have de...

  15. Molecular cloning of the apoptosis-related calcium-binding protein AsALG-2 in Avena sativa. (United States)

    Hoat, Trinh Xuan; Nakayashiki, Hitoshi; Yang, Qian; Tosa, Yukio; Mayama, Shigeyuki


    Victorin, the host-selective toxin produced by the fungus Cochliobolus victoriae, induces programmed cell death (PCD) in victorin-sensitive oat lines with characteristic features of animal apoptosis, such as mitochondrial permeability transition, chromatin condensation, nuclear DNA laddering and rRNA/mRNA degradation. In this study, we characterized a calcium-binding protein, namely AsALG-2, which might have a role in the victorin-induced PCD. AsALG-2 is homologous to the Apoptosis-Linked Gene ALG-2 identified in mammalian cells. Northern blot analysis revealed that the accumulation of AsALG-2 transcripts increased during victorin-induced PCD, but not during necrotic cell death. Salicylic acid, chitosan and chitin strongly activated the expression of general defence response genes, such as PR-10; however, neither induced cell death nor the accumulation of AsALG-2 mRNA. Pharmacological studies indicated that victorin-induced DNA laddering and AsALG-2 expression were regulated through similar pathways. The calcium channel blocker, nifedipine, moderately inhibited the accumulation of AsALG-2 mRNA during cell death. Trifluoperazine (calmodulin antagonist) and K252a (serine-threonine kinase inhibitor) reduced the victorin-induced phytoalexin accumulation, but did not prevent the victorin-induced DNA laddering or accumulation of AsALG-2 mRNA. Taken together, our investigations suggest that there is a calcium-mediated signalling pathway in animal and plant PCD in common. © 2012 THE AUTHORS. MOLECULAR PLANT PATHOLOGY © 2012 BSPP AND BLACKWELL PUBLISHING LTD.

  16. A novel biomarker associated with distress in humans: calcium-binding protein, spermatid-specific 1 (CABS1). (United States)

    Ritz, Thomas; Rosenfield, David; St Laurent, Chris D; Trueba, Ana F; Werchan, Chelsey A; Vogel, Pia D; Auchus, Richard J; Reyes-Serratos, Eduardo; Befus, A Dean


    Calcium-binding protein spermatid-specific 1 (CABS1) is expressed in the human submandibular gland and has an anti-inflammatory motif similar to that in submandibular rat 1 in rats. Here, we investigate CABS1 in human saliva and its association with psychological and physiological distress and inflammation in humans. Volunteers participated across three studies: 1) weekly baseline measures; 2) a psychosocial speech and mental arithmetic stressor under evaluative threat; and 3) during academic exam stress. Salivary samples were analyzed for CABS1 and cortisol. Additional measures included questionnaires of perceived stress and negative affect; exhaled nitric oxide; respiration and cardiac activity; lung function; and salivary and nasal inflammatory markers. We identified a CABS1 immunoreactive band at 27 kDa in all participants and additional molecular mass forms in some participants. One week temporal stability of the 27-kDa band was satisfactory (test-retest reliability estimate = 0.62-0.86). Acute stress increased intensity of 18, 27, and 55 kDa bands; 27-kDa increases were associated with more negative affect and lower heart rate, sympathetic activity, respiration rate, and minute ventilation. In both acute and academic stress, changes in 27 kDa were positively associated with salivary cortisol. The 27-kDa band was also positively associated with VEGF and salivary leukotriene B4 levels. Participants with low molecular weight CABS1 bands showed reduced habitual stress and negative affect in response to acute stress. CABS1 is readily detected in human saliva and is associated with psychological and physiological indicators of stress. The role of CABS1 in inflammatory processes, stress, and stress resilience requires careful study. Copyright © 2017 the American Physiological Society.

  17. Deficiency in Calcium-Binding Protein S100A4 Impairs the Adjuvant Action of Cholera Toxin

    Directory of Open Access Journals (Sweden)

    Jia-Bin Sun


    Full Text Available The calcium-binding protein S100A4 has been described to promote pathological inflammation in experimental autoimmune and inflammatory disorders and in allergy and to contribute to antigen presentation and antibody response after parenteral immunization with an alum-adjuvanted antigen. In this study, we extend these findings by demonstrating that mice lacking S100A4 have a defective humoral and cellular immune response to mucosal (sublingual immunization with a model protein antigen [ovalbumin (OVA] given together with the strong mucosal adjuvant cholera toxin (CT, and that this impairment is due to defective adjuvant-stimulated antigen presentation by antigen-presenting cells. In comparison to wild-type (WT mice, mice genetically lacking S100A4 had reduced humoral and cellular immune responses after immunization with OVA plus CT, including a complete lack of detectable germinal center reaction. Further, when stimulated in vitro with OVA plus CT, S100A4−/− dendritic cells (DCs showed impaired responses in several CT-stimulated immune regulatory molecules including the co-stimulatory molecule CD86, inflammasome-associated caspase-1 and IL-1β. Coculture of OVA-specific OT-II T cells with S100A4−/− DCs that had been pulse incubated with OVA plus CT resulted in impaired OT-II T cell proliferation and reduced production of Th1, Th2, and Th17 cytokines compared to similar cocultures with WT DCs. In accordance with these findings, transfection of WT DCs with S100A4-targeting small interfering RNA (siRNA but not mock-siRNA resulted in significant reductions in the expression of caspase-1 and IL-1β as well as CD86 in response to CT. Importantly, also engraftment of WT DCs into S100A4−/− mice effectively restored the immune response to immunization in the recipients. In conclusion, our results demonstrate that deficiency in S100A4 has a strong impact on the development of both humoral and cellular immunity after mucosal immunization using CT

  18. Comparative distribution of relaxin-3 inputs and calcium-binding protein-positive neurons in rat amygdala

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    Fabio N Santos


    Full Text Available The neural circuits involved in mediating complex behaviors are being rapidly elucidated using various newly developed and powerful anatomical and molecular techniques, providing insights into the neural basis for anxiety disorders, depression, addiction, and dysfunctional social behaviors. Many of these behaviors and associated physiological processes involve the activation of the amygdala in conjunction with cortical and hippocampal circuits. Ascending subcortical projections provide modulatory inputs to the extended amygdala and its related nodes (or ‘hubs’ within these key circuits. One such input arises from the nucleus incertus (NI in the tegmentum, which sends amino acid- and peptide-containing projections throughout the forebrain. Notably, a distinct population of GABAergic NI neurons expresses the highly-conserved neuropeptide, relaxin-3, and relaxin-3 signaling has been implicated in the modulation of reward/motivation and anxiety- and depressive-like behaviors in rodents via actions within the extended amygdala. Thus, a detailed description of the relaxin-3 innervation of the extended amygdala would provide an anatomical framework for an improved understanding of NI and relaxin-3 modulation of these and other specific amygdala-related functions. Therefore, in this study, we examined the distribution of NI projections and relaxin-3-positive elements (axons/fibers/terminals within the amygdala, relative to the distribution of neurons expressing the calcium-binding proteins, parvalbumin, calretinin and/or calbindin. Anterograde tracer injections into the NI revealed a topographic distribution of NI efferents within the amygdala that was near identical to the distribution of relaxin-3-immunoreactive fibers. Highest densities of anterogradely-labeled elements and relaxin-3-immunoreactive fibers were observed in the medial nucleus of the amygdala, medial divisions of the bed nucleus of the stria terminalis (BST and in the endopiriform

  19. Crystallization and preliminary crystallographic analysis of calcium-binding protein-2 from Entamoeba histolytica and its complexes with strontium and the IQ1 motif of myosin V

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    Gourinath, S., E-mail:; Padhan, Narendra; Alam, Neelima; Bhattacharya, Alok [School of Life Sciences, Jawaharlal Nehru University, New Delhi 110067 (India)


    Calcium-binding protein-2 (EhCaBP2) crystals were grown using MPD as a precipitant. EhCaBP2 also crystallized in complex with strontium (replacing calcium) at similar conditions. Preliminary data for EhCaBP2 crystals in complex with an IQ motif are also reported. Calcium plays a pivotal role in the pathogenesis of amoebiasis, a major disease caused by Entamoeba histolytica. Two domains with four canonical EF-hand-containing calcium-binding proteins (CaBPs) have been identified from E. histolytica. Even though they have very high sequence similarity, these bind to different target proteins in a Ca{sup 2+}-dependent manner, leading to different functional pathways. Calcium-binding protein-2 (EhCaBP2) crystals were grown using MPD as a precipitant. The crystals belong to space group P2{sub 1}, with unit-cell parameters a = 111.74, b = 68.83, c = 113.25 Å, β = 116.7°. EhCaBP2 also crystallized in complex with strontium (replacing calcium) at similar conditions. The crystals belong to space group P2{sub 1}, with unit-cell parameters a = 69.18, b = 112.03, c = 93.42 Å, β = 92.8°. Preliminary data for EhCaBP2 crystals in complex with an IQ motif are also reported. This complex was crystallized with MPD and ethanol as precipitating agents. These crystals belong to space group P2{sub 1}, with unit-cell parameters a = 60.5, b = 69.86, c = 86.5 Å, β = 97.9°.

  20. Identification of a novel calcium binding motif based on the detection of sequence insertions in the animal peroxidase domain of bacterial proteins.

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    Saray Santamaría-Hernando

    Full Text Available Proteins of the animal heme peroxidase (ANP superfamily differ greatly in size since they have either one or two catalytic domains that match profile PS50292. The orf PP_2561 of Pseudomonas putida KT2440 that we have called PepA encodes a two-domain ANP. The alignment of these domains with those of PepA homologues revealed a variable number of insertions with the consensus G-x-D-G-x-x-[GN]-[TN]-x-D-D. This motif has also been detected in the structure of pseudopilin (pdb 3G20, where it was found to be involved in Ca(2+ coordination although a sequence analysis did not reveal the presence of any known calcium binding motifs in this protein. Isothermal titration calorimetry revealed that a peptide containing this consensus motif bound specifically calcium ions with affinities ranging between 33-79 µM depending on the pH. Microcalorimetric titrations of the purified N-terminal ANP-like domain of PepA revealed Ca(2+ binding with a K(D of 12 µM and stoichiometry of 1.25 calcium ions per protein monomer. This domain exhibited peroxidase activity after its reconstitution with heme. These data led to the definition of a novel calcium binding motif that we have termed PERCAL and which was abundantly present in animal peroxidase-like domains of bacterial proteins. Bacterial heme peroxidases thus possess two different types of calcium binding motifs, namely PERCAL and the related hemolysin type calcium binding motif, with the latter being located outside the catalytic domains and in their C-terminal end. A phylogenetic tree of ANP-like catalytic domains of bacterial proteins with PERCAL motifs, including single domain peroxidases, was divided into two major clusters, representing domains with and without PERCAL motif containing insertions. We have verified that the recently reported classification of bacterial heme peroxidases in two families (cd09819 and cd09821 is unrelated to these insertions. Sequences matching PERCAL were detected in all kingdoms of

  1. The calcium-binding protein ALG-2 regulates protein secretion and trafficking via interactions with MISSL and MAP1B proteins. (United States)

    Takahara, Terunao; Inoue, Kuniko; Arai, Yumika; Kuwata, Keiko; Shibata, Hideki; Maki, Masatoshi


    Mobilization of intracellular calcium is essential for a wide range of cellular processes, including signal transduction, apoptosis, and vesicular trafficking. Several lines of evidence have suggested that apoptosis-linked gene 2 (ALG-2, also known as PDCD6), a calcium-binding protein, acts as a calcium sensor linking calcium levels with efficient vesicular trafficking, especially at the endoplasmic reticulum (ER)-to-Golgi transport step. However, how ALG-2 regulates these processes remains largely unclear. Here, we report that MAPK1-interacting and spindle-stabilizing (MISS)-like (MISSL), a previously uncharacterized protein, interacts with ALG-2 in a calcium-dependent manner. Live-cell imaging revealed that upon a rise in intracellular calcium levels, GFP-tagged MISSL (GFP-MISSL) dynamically relocalizes in a punctate pattern and colocalizes with ALG-2. MISSL knockdown caused disorganization of the components of the ER exit site, the ER-Golgi intermediate compartment, and Golgi. Importantly, knockdown of either MISSL or ALG-2 attenuated the secretion of secreted alkaline phosphatase (SEAP), a model secreted cargo protein, with similar reductions in secretion by single- and double-protein knockdowns, suggesting that MISSL and ALG-2 act in the same pathway to regulate the secretion process. Furthermore, ALG-2 or MISSL knockdown delayed ER-to-Golgi transport of procollagen type I. We also found that ALG-2 and MISSL interact with microtubule-associated protein 1B (MAP1B) and that MAP1B knockdown reverts the reduced secretion of SEAP caused by MISSL or ALG-2 depletion. These results suggest that a change in the intracellular calcium level plays a role in regulation of the secretory pathway via interaction of ALG-2 with MISSL and MAP1B. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  2. The serum concentration of the calcium binding protein S100B is positively associated with cognitive performance in older adults

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    Virginie eLam


    Full Text Available S100B is a calcium-binding peptide produced predominantly by astroglial cells in the central nervous system. S100B paradoxically has neurotrophic and apoptotic effects, dependent on extracellular concentration. This study investigated the relationship between serum S100B levels and neuropsychological performance across a range of cognitive domains in healthy older aged adults. A cohort of 219 participants between the ages of 43 and 84 years (141 female were recruited. Subjects provided a fasting blood sample for S100B measurement (Mean = 0.24 ng/mL, SD = 0.14 and completed a battery of neuropsychological tests. S100B concentrations (both with and without the covariates of age and sex were positively associated with the following measures of cognitive performance: digit symbol coding, Stroop test and measures of verbal ability. The results from this study show that serum S100B is positively associated with better cognitive performance in healthy older adults.

  3. Acute Sleep Deprivation Increases Serum Levels of Neuron-Specific Enolase (NSE) and S100 Calcium Binding Protein B (S-100B) in Healthy Young Men (United States)

    Benedict, Christian; Cedernaes, Jonathan; Giedraitis, Vilmantas; Nilsson, Emil K.; Hogenkamp, Pleunie S.; Vågesjö, Evelina; Massena, Sara; Pettersson, Ulrika; Christoffersson, Gustaf; Phillipson, Mia; Broman, Jan-Erik; Lannfelt, Lars; Zetterberg, Henrik; Schiöth, Helgi B.


    Study Objectives: To investigate whether total sleep deprivation (TSD) affects circulating concentrations of neuron-specific enolase (NSE) and S100 calcium binding protein B (S-100B) in humans. These factors are usually found in the cytoplasm of neurons and glia cells. Increasing concentrations of these factors in blood may be therefore indicative for either neuronal damage, impaired blood brain barrier function, or both. In addition, amyloid β (Aβ) peptides 1-42 and 1-40 were measured in plasma to calculate their ratio. A reduced plasma ratio of Aβ peptides 1-42 to 1-40 is considered an indirect measure of increased deposition of Aβ 1-42 peptide in the brain. Design: Subjects participated in two conditions (including either 8-h of nocturnal sleep [22:30-06:30] or TSD). Fasting blood samples were drawn before and after sleep interventions (19:30 and 07:30, respectively). Setting: Sleep laboratory. Participants: 15 healthy young men. Results: TSD increased morning serum levels of NSE (P = 0.002) and S-100B (P = 0.02) by approximately 20%, compared with values obtained after a night of sleep. In contrast, the ratio of Aβ peptides 1-42 to 1-40 did not differ between the sleep interventions. Conclusions: Future studies in which both serum and cerebrospinal fluid are sampled after sleep loss should elucidate whether the increase in serum neuron-specific enolase and S100 calcium binding protein B is primarily caused by neuronal damage, impaired blood brain barrier function, or is just a consequence of increased gene expression in non-neuronal cells, such as leukocytes. Citation: Benedict C; Cedernaes J; Giedraitis V; Nilsson EK; Hogenkamp PS; Vågesjö E; Massena S; Pettersson U; Christoffersson G; Phillipson M; Broman JE; Lannfelt L; Zetterberg H; Schiöth HB. Acute Sleep Deprivation Increases Serum Levels of Neuron-Specific Enolase (NSE) and S100 Calcium Binding Protein B (S-100B) in Healthy Young Men. SLEEP 2014;37(1):195-198. PMID:24470708

  4. Gene expression and protein localisation of calcyclin, a calcium-binding protein of the S-100 family in fresh neuroblastomas. (United States)

    Tonini, G P; Fabretti, G; Kuznicki, J; Massimo, L; Scaruffi, P; Brisigotti, M; Mazzocco, K


    Calcyclin gene, a Ca(2+)-binding protein with homology to S-100, has been found to be expressed at different levels in leukaemic cells and in other tumour cells. We recently reported the expression of the gene in human neuroblastoma (NB) cell lines, and suggested a possible role of calcyclin in cell differentiation. To extend our findings, we investigated the expression of the gene in NB cells induced to differentiate by retinoic acid (RA), using the reverse transcriptase-polymerase chain reaction (RT-PCR) technique. Time-course experiments employing LA-N-5 cells showed that calcyclin mRNA appeared 2 h after RA treatment, long before the cells were blocked in the G1 cell-cycle phase and before the neurite-like structures outgrew from the cell bodies. This suggests the involvement of the gene in the early phase of cell differentiation. Furthermore, we investigated mRNA expression in a series of fresh neuroblastomas. NB tumours showed a heterogeneous pattern of calcyclin expression, although calcyclin seemed to be expressed more frequently in cases with a favourable Shimada histology. We also studied the expression of the protein in formalin fixed and paraffin embedded tissues, by using a specific anticalcyclin antibody. The protein was detected in stromal cells which characterise a more mature histological type, and in nerve sheaths, whereas neuroblasts were negative. The tissue that expressed calcyclin protein showed a Schwann-like differentiation and, unlike S-100 protein, calcyclin was expressed in the perineurium.

  5. Flexibility of EF-hand motifs: structural and thermodynamic studies of Calcium Binding Protein-1 from Entamoeba histolytica with Pb2+, Ba2+, and Sr2+

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    Kumar Shivesh


    Full Text Available Abstract Background EF-hand proteins can be activated by the binding of various heavy metals other than calcium, and such complexes can disturb the calcium-signaling pathway and cause toxicity and disease causing state. So far, no comprehensive study has been done to understand different heavy metals binding to calcium signaling proteins. Results In this work, the flexibility of the EF-hand motifs are examined by crystallographic and thermodynamic studies of binding of Pb2+, Ba2+ and Sr2+ to Calcium Binding Protein-1 from Entamoeba histolytica (EhCaBP1. The structures of the EhCaBP1- heavy metal complexes are found to be overall similar, nevertheless specific differences in metal coordination, and small differences in the coordination distances between the metal and the ligands in the metal binding loop. The largest such distances occur for the Ba2+- EhCaBP1 complex, where two bariums are bound with partial occupancy at the EF2 motif. Thermodynamic studies confirm that EhCaBP1 has five binding sites for Ba2+ compared to four binding sites for the other metals. These structures and thermodynamic studies reveal that the EF-hand motifs can accommodate several heavy atoms with similar binding affinities. The binding of Ca2+ to the 1st, 2nd and 4th sites and the binding of Ba2+ to the 1st, 2nd, 4th and 5th sites are both enthalpically and entropically driven, whereas the binding of Sr2+ to the 1st, 2nd and 4th sites are simply enthalpy driven, interestingly in agreement with ITC data, Sr2+ do not coordinate with water in this structure. For all the metals, binding to the 3rd site is only entropy driven. Conclusion Energetically, Ca2+ is preferred in three sites, while in one site Ba2+ has better binding energy. The Sr2+-coordination in the EF hand motifs is similar to that of the native Ca2+ bound structure, except for the lack of water coordination. Sr2+ coordination seems to be a pre-formed in nature since all seven coordinating atoms are from the

  6. Changes of calcium binding proteins, c-Fos and COX in hippocampal formation and cerebellum of Niemann-Pick, type C mouse. (United States)

    Byun, Kyunghee; Kim, Daesik; Bayarsaikhan, Enkhjaigal; Oh, Jeehyun; Kim, Jisun; Kwak, Grace; Jeong, Goo-Bo; Jo, Seung-Mook; Lee, Bonghee


    Niemann-Pick disease, type C (NPC) is an intractable disease that is accompanied by ataxia, dystonia, neurodegeneration, and dementia due to an NPC gene defect. Disruption of calcium homeostasis in neurons is important in patients with NPC. Thus, we used immunohistochemistry to assess the expression levels of calcium binding proteins (calbindin D28K, parvalbumin, and calretinin), c-Fos and cyclooxygenase-1,2 (COX-1,2) in the hippocampal formation and cerebellum of 4 and 8 week old NPC+/+, NPC+/-, and NPC-/- mice. General expression of these proteins decreased in the hippocampus and cerebellum of NPC-/- compared to that in both young and adult NPC+/+ or NPC+/- mice. Parvalbumin, COX-1,2 or c-Fos-immunoreactive neurons were widely detected in the CA1, CA3, and DG of the hippocampus, but the immunoreactivities were decreased sharply in all areas of hippocampus of NPC-/- compared to NPC+/+ and NPC+/- mice. Taken together, reduction of these proteins may be one of the strong phenotypes related to the neuronal degeneration in NPC-/- mice. Copyright © 2013 Elsevier B.V. All rights reserved.

  7. [Mass spectrometry identification and immune cross-reactivity of a minor shrimp allergen-sarcoplasmic calcium binding protein from Litopenaeus vannamei]. (United States)

    Wang, Cai-xia; Huang, Jian-fang; Xiang, Jun-jian; Sun, Yi-fan; Lv, Si; Guo, Jie


    To identify sarcoplasmic calcium-binding protein (SCP) as a minor shrimp allergen by mass spectrometry, and to analyze the immune cross-reactivity among crustacean SCPs. The M(r); 21 000 allergen from Litopenaeus vannamei was identified by MALDI-TOF/TOF-MS. BLAST and ClustalW were used to compare amino acid sequence identity of the allergen among crustaceans. The puritifed M(r); 21 000 allergen was injected subcutaneously in mice to produce the specific polyclonal antibodies to analyze immune cross-reactivity of the allergen with proteins from 8 other species of crustaceans by Western blotting. The M(r); 21 000 shrimp allergen was identified as SCP. Sequence comparison revealed that SCP had 81%-100% amino acid identity among crustaceans. Western blotting showed that the proteins with M(r); about 21 000, corresponding to SCP from Metapenaeus ensis, Penaeus monodon, Oratosquilla oratoria, Macrobrachium rosenbergii, Procambarus clarkii, Portunus pelagicus, Charybdis feriatus, Eriocheir sinensis were recognized by polyclonal antibodies against SCP of Litopenaeus vannamei. SCP is a minor shrimp allergen, and SCPs have a high sequence homology and strong immune cross-reactivity among crustaceans, which can be used as detective, diagnostic and safe immunotherapeutic agents for subjects with shrimp allergy.

  8. Structural Insights into Membrane Targeting by the Flagellar Calcium-binding Protein (FCaBP) a Myristoylated and Palmitoylated Calcium Sensor in Trypanosoma cruzi

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    J Wingard; J Ladner; M Vanarotti; A Fisher; H Robinson; K Buchanan; D Engman; J Ames


    The flagellar calcium-binding protein (FCaBP) of the protozoan Trypanosoma cruzi is targeted to the flagellar membrane where it regulates flagellar function and assembly. As a first step toward understanding the Ca{sup 2+}-induced conformational changes important for membrane-targeting, we report here the x-ray crystal structure of FCaBP in the Ca{sup 2+}-free state determined at 2.2{angstrom} resolution. The first 17 residues from the N terminus appear unstructured and solvent-exposed. Residues implicated in membrane targeting (Lys-19, Lys-22, and Lys-25) are flanked by an exposed N-terminal helix (residues 26-37), forming a patch of positive charge on the protein surface that may interact electrostatically with flagellar membrane targets. The four EF-hands in FCaBP each adopt a 'closed conformation' similar to that seen in Ca{sup 2+}-free calmodulin. The overall fold of FCaBP is closest to that of grancalcin and other members of the penta EF-hand superfamily. Unlike the dimeric penta EF-hand proteins, FCaBP lacks a fifth EF-hand and is monomeric. The unstructured N-terminal region of FCaBP suggests that its covalently attached myristoyl group at the N terminus may be solvent-exposed, in contrast to the highly sequestered myristoyl group seen in recoverin and GCAP1. NMR analysis demonstrates that the myristoyl group attached to FCaBP is indeed solvent-exposed in both the Ca{sup 2+}-free and Ca{sup 2+}-bound states, and myristoylation has no effect on protein structure and folding stability. We propose that exposed acyl groups at the N terminus may anchor FCaBP to the flagellar membrane and that Ca{sup 2+}-induced conformational changes may control its binding to membrane-bound protein targets..

  9. The lack of relationships between vitamin D3 metabolites and calcium-binding protein in the eggshell gland of laying birds. (United States)

    Bar, A; Rosenberg, J; Hurwitz, S


    The relationship of the metabolism of vitamin D3 and calcium-binding protein (CaBP) to calcium transport by the eggshell gland (ESG) was assessed in chickens. Plasma or ESG 1,25 dihydroxyvitamin D3 (1,25(OH)2D3) and ESG CaBP were no different between periods of ESG inactivity and of shell calcification. A severe dietary calcium deficiency resulted in increased kidney 25-hydroxycholecalciferol-1-hydroxylase activity (542%), plasma and ESG 1,25(OH)2D3 concentrations (193 and 274%, respectively), but in decreased ESG CaBP (34%), associated with the production of poorly calcified eggs. Significant correlations were found between 25 hydroxycholecalciferol-1-hydroxylase, plasma 1,25(OH)2D3 and ESG 1,25(OH)2D3, but not between ESG 1,25(OH)2D3 and CaBP. Hens with a low shell density had a significantly lower (55%) ESG CaBP than those with high shell density, without any significant change in ESG 1,25(OH)2D3. Significant correlations were found between ESG CaBP and shell calcium. Total receptors for 1,25(OH)2D3 were lower in ESG than in the intestine. The results suggest that CaBP level and calcium transport in the ESG are not regulated by 1,25(OH)2D3.

  10. Comparative Analysis of Calcium-Binding Myeloid-Related Protein-8/14 in Saliva and Serum of Patients With Periodontitis and Healthy Individuals. (United States)

    Haririan, Hady; Andrukhov, Oleh; Pablik, Eleonore; Neuhofer, Michaela; Moritz, Andreas; Rausch-Fan, Xiaohui


    This study aims to investigate calcium-binding myeloid-related protein (MRP)-8/14 in the saliva and serum of individuals with periodontitis and periodontally healthy individuals for the assessment of its role in the pathogenesis and clinical diagnosis of periodontitis. This cross-sectional study includes 56 patients with periodontitis and 44 periodontally healthy individuals. Saliva and serum were collected for the detection of MRP-8/14 and calcium levels. Periodontopathic bacteria were determined by polymerase chain reaction in saliva. Correlations between salivary and serum MRP-8/14 levels and clinical parameters, bacteria, and calcium were analyzed with Pearson correlation in a multiple regression model. MRP-8/14 levels were documented with receiver operating characteristic (ROC) curves. Compared with healthy individuals, MRP-8/14 levels were significantly higher in both the saliva and serum of patients with periodontitis, but calcium was increased only in saliva. A high diagnostic potential of salivary MRP-8/14 was detected for periodontitis (ROC = 0.86). Salivary MRP-8/14 levels correlated significantly with the presence of the periodontopathogen Treponema denticola, as well as with the clinical parameters of periodontitis. MRP-8/14 in saliva might be a potential diagnostic parameter for periodontal disease.

  11. Prenatal acoustic stimulation influences neuronal size and the expression of calcium-binding proteins (calbindin D-28K and parvalbumin) in chick hippocampus. (United States)

    Chaudhury, Sraboni; Nag, Tapas Chandra; Wadhwa, Shashi


    Prenatal auditory enrichment by species-specific sounds and sitar music enhances the expression of immediate early genes, synaptic proteins and calcium binding proteins (CaBPs) as well as modifies the structural components of the brainstem auditory nuclei and auditory imprinting area in chicks. There is also facilitation of postnatal auditory preference of the chicks to maternal calls following both types of sound stimulation indicating prenatal perceptual learning. To examine whether the sound enrichment protocol also affects the areas related to learning and memory, we assessed morphological changes in the hippocampus at post-hatch day 1 of control and prenatally sound-stimulated chicks. Additionally, the proportions of neurons containing calbindin D-28K and parvalbumin immunoreactivity as well as their protein levels were determined. Fertilized eggs of domestic chick were incubated under normal conditions of temperature, humidity, forced draft of air as well as light and dark (12:12h) photoperiods. They were exposed to patterned sounds of species-specific and sitar music at 65 dB for 15 min per hour over a day/night cycle from day 10 of incubation till hatching. The hippocampal volume, neuronal nuclear size and total number of neurons showed a significant increase in the music-stimulated group as compared to the species-specific sound-stimulated and control groups. However, in both the auditory-stimulated groups the protein levels of calbindin and parvalbumin as well as the percentage of the immunopositive neurons were increased. The enhanced proportion of CaBPs in the sound-enriched groups suggests greater Ca(2+) influx, which may influence long-term potentiation and short-term memory.

  12. A calcium-binding protein, rice annexin OsANN1, enhances heat stress tolerance by modulating the production of H2O2. (United States)

    Qiao, Bei; Zhang, Qian; Liu, Dongliang; Wang, Haiqi; Yin, Jingya; Wang, Rui; He, Mengli; Cui, Meng; Shang, Zhonglin; Wang, Dekai; Zhu, Zhengge


    OsANN1 is a member of the annexin protein family in rice. The function of this protein and the mechanisms of its involvement in stress responses and stress tolerance are largely unknown. Here it is reported that OsANN1 confers abiotic stress tolerance by modulating antioxidant accumulation under abiotic stress. OsANN1-knockdown [RNA interference (RNAi)] plants were more sensitive to heat and drought stresses, whereas OsANN1-overexpression (OE) lines showed improved growth with higher expression of OsANN1 under abiotic stress. Overexpression of OsANN1 promoted SOD (superoxide dismutase) and CAT (catalase) activities, which regulate H2O2 content and redox homeostasis, suggesting the existence of a feedback mechanism between OsANN1 and H2O2 production under abiotic stress. Higher expression of OsANN1 can provide overall cellular protection against abiotic stress-induced damage, and a significant accumulation of OsANN1-green fluorescent protein (GFP) signals was found in the cytosol after heat shock treatment. OsANN1 also has calcium-binding and ATPase activities in vitro, indicating that OsANN1 has multiple functions in rice growth. Furthermore, yeast two-hybrid and bimolecular fluorescence complementation (BiFC) assays demonstrated that OsANN1 interacts with OsCDPK24. This cross-talk may provide additional layers of regulation in the abiotic stress response. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email:

  13. Chitinase III in pomegranate seeds (Punica granatum Linn.): a high-capacity calcium-binding protein in amyloplasts. (United States)

    Yang, Haixia; Zhang, Tuo; Masuda, Taro; Lv, Chenyan; Sun, Lei; Qu, Guiqin; Zhao, Guanghua


    Chitinases are a class of ubiquitous proteins that are widely distributed in plants. Defense is the major natural role for chitinases, primarily against fungal pathogens. Little is known regarding their non-defensive roles in seeds. In this study, a new class III chitinase from pomegranate seeds (pomegranate seed chitinase, PSC) was isolated and purified to homogeneity. The native state of PSC is a monomer with a molecular weight of approximately 30 kDa. This chitinase naturally binds calcium ions with high capacity and low affinity, suggesting that PSC is a calcium storage protein. Consistent with this idea, its amino acid sequence (inferred from cDNA) is rich in acidic amino acid residues, especially Asp, similar to reported calcium storage proteins. The presence of calcium considerably improves the stability of the protein but has little effect on its enzymatic activity. Transmission electron microscopy analyses indicate that, similar to phytoferritin, this enzyme is widely distributed in the stroma of amyloplasts of the embryonic cells, suggesting that amyloplasts in seeds could serve as an alternative plastid for calcium storage. Indeed, the transmission electron microscopy results showed that, within the embryonic cells, calcium ions are mainly distributed in the stroma of the amyloplasts, consistent with a role for PSC in calcium storage. Thus, the plant appears to have evolved a new plastid for calcium storage in seeds. During seed germination, the content of this enzyme decreases with time, suggesting that it is involved in the germination process. © 2011 The Authors. The Plant Journal © 2011 Blackwell Publishing Ltd.

  14. Expression of calcium-binding proteins and selected neuropeptides in the human, chimpanzee, and crab-eating macaque claustrum. (United States)

    Pirone, Andrea; Castagna, Maura; Granato, Alberto; Peruffo, Antonella; Quilici, Francesca; Cavicchioli, Laura; Piano, Ilaria; Lenzi, Carla; Cozzi, Bruno


    The claustrum is present in all mammalian species examined so far and its morphology, chemoarchitecture, physiology, phylogenesis and ontogenesis are still a matter of debate. Several morphologically distinct types of immunostained cells were described in different mammalian species. To date, a comparative study on the neurochemical organization of the human and non-human primates claustrum has not been fully described yet, partially due to technical reasons linked to the postmortem sampling interval. The present study analyze the localization and morphology of neurons expressing parvalbumin (PV), calretinin (CR), NPY, and somatostatin (SOM) in the claustrum of man (# 5), chimpanzee (# 1) and crab-eating monkey (# 3). Immunoreactivity for the used markers was observed in neuronal cell bodies and processes distributed throughout the anterior-posterior extent of human, chimpanzee and macaque claustrum. Both CR- and PV-immunoreactive (ir) neurons were mostly localized in the central and ventral region of the claustrum of the three species while SOM- and NPY-ir neurons seemed to be equally distributed throughout the ventral-dorsal extent. In the chimpanzee claustrum SOM-ir elements were not observed. No co-localization of PV with CR was found, thus suggesting the existence of two non-overlapping populations of PV and CR-ir interneurons. The expression of most proteins (CR, PV, NPY), was similar in all species. The only exception was the absence of SOM-ir elements in the claustrum of the chimpanzee, likely due to species specific variability. Our data suggest a possible common structural organization shared with the adjacent insular region, a further element that emphasizes a possible common ontogeny of the claustrum and the neocortex.

  15. Expression of calcium-binding proteins and selected neuropeptides in the human, chimpanzee, and crab-eating macaque claustrum

    Directory of Open Access Journals (Sweden)

    Andrea ePirone


    Full Text Available The claustrum is present in all mammalian species examined so far and its morphology, chemoarchitecture, physiology, phylogenesis and ontogenesis are still a matter of debate. Several morphologically distinct types of immunostained cells were described in different mammalian species. To date, a comparative study on the neurochemical organization of the human and non-human primates claustrum has not been fully described yet, partially due to technical reasons linked to the postmortem sampling interval. The present study analyzes the localization and morphology of neurons expressing parvalbumin (PV, calretinin (CR, NPY, and somatostatin (SOM in the claustrum of man (# 5, chimpanzee (# 1 and crab-eating monkey (#3. Immunoreactivity for the used markers was observed in neuronal cell bodies and processes distributed throughout the anterior-posterior extent of human, chimpanzee and macaque claustrum. Both CR- and PV-immunoreactive (ir neurons were mostly localized in the central and ventral region of the claustrum of the three species while SOM- and NPY-ir neurons seemed to be equally distributed throughout the ventral-dorsal extent. In the chimpanzee claustrum SOM-ir elements were not observed. No co-localization of PV with CR was found, thus suggesting the existence of two non-overlapping populations of PV and CR-ir interneurons. The expression of most proteins (CR, PV, NPY, was similar in all species. The only exception was the absence of SOM-ir elements in the claustrum of the chimpanzee, likely due to species specific variability. Our data suggest a possible common structural organization shared with the adjacent insular region, a further element that emphasizes a possible common ontogeny of the claustrum and the neocortex.

  16. Short-term exposure to L-type calcium channel blocker, verapamil, alters the expression pattern of calcium-binding proteins in the brain of goldfish, Carassius auratus. (United States)

    Palande, Nikhil V; Bhoyar, Rahul C; Biswas, Saikat P; Jadhao, Arun G


    The influx of calcium ions (Ca(2+)) is responsible for various physiological events including neurotransmitter release and synaptic modulation. The L-type voltage dependent calcium channels (L-type VDCCs) transport Ca(2+) across the membrane. Calcium-binding proteins (CaBPs) bind free cytosolic Ca(2+) and prevent excitotoxicity caused by sudden increase in cytoplasmic Ca(2+). The present study was aimed to understand the regulation of expression of neuronal CaBPs, namely, calretinin (CR) and parvalbumin (PV) following blockade of L-type VDCCs in the CNS of Carassius auratus. Verapamil (VRP), a potent L-type VDCC blocker, selectively blocks Ca(2+) entry at the plasma membrane level. VRP present in the aquatic environment at a very low residual concentration has shown ecotoxicological effects on aquatic animals. Following acute exposure for 96h, median lethal concentration (LC50) for VRP was found to be 1.22mg/L for goldfish. At various doses of VRP, the behavioral alterations were observed in the form of respiratory difficulty and loss of body balance confirming the cardiovascular toxicity caused by VRP at higher doses. In addition to affecting the cardiovascular system, VRP also showed effects on the nervous system in the form of altered expression of PV. When compared with controls, the pattern of CR expression did not show any variations, while PV expression showed significant alterations in few neuronal populations such as the pretectal nucleus, inferior lobes, and the rostral corpus cerebellum. Our result suggests possible regulatory effect of calcium channel blockers on the expression of PV. Copyright © 2015 Elsevier Inc. All rights reserved.

  17. Intracellular amyloid-β accumulation in calcium-binding protein-deficient neurons leads to amyloid-β plaque formation in animal model of Alzheimer's disease. (United States)

    Moon, Minho; Hong, Hyun-Seok; Nam, Dong Woo; Baik, Sung Hoon; Song, Hyundong; Kook, Sun-Young; Kim, Yong Soo; Lee, Jeewoo; Mook-Jung, Inhee


    One of the major hallmarks of Alzheimer's disease (AD) is the extracellular deposition of amyloid-β (Aβ) as senile plaques in specific brain regions. Clearly, an understanding of the cellular processes underlying Aβ deposition is a crucial issue in the field of AD research. Recent studies have found that accumulation of intraneuronal Aβ (iAβ) is associated with synaptic deficits, neuronal death, and cognitive dysfunction in AD patients. In this study, we found that Aβ deposits had several shapes and sizes, and that iAβ occurred before the formation of extracellular amyloid plaques in the subiculum of 5XFAD mice, an animal model of AD. We also observed pyroglutamate-modified Aβ (N3pE-Aβ), which has been suggested to be a seeding molecule for senile plaques, inside the Aβ plaques only after iAβ accumulation, which argues against its seeding role. In addition, we found that iAβ accumulates in calcium-binding protein (CBP)-free neurons, induces neuronal death, and then develops into senile plaques in 2-4-month-old 5XFAD mice. These findings suggest that N3pE-Aβ-independent accumulation of Aβ in CBP-free neurons might be an early process that triggers neuronal damage and senile plaque formation in AD patients. Our results provide new insights into several long-standing gaps in AD research, namely how Aβ plaques are formed, what happens to iAβ and how Aβ causes selective neuronal loss in AD patients.

  18. S100 calcium binding protein B as a biomarker of delirium duration in the intensive care unit – an exploratory analysis

    Directory of Open Access Journals (Sweden)

    Khan BA


    Full Text Available Babar A Khan,1–3 Mark O Farber,1 Noll Campbell,2–5 Anthony Perkins,2,3 Nagendra K Prasad,6 Siu L Hui,1–3 Douglas K Miller,1–3 Enrique Calvo-Ayala,1 John D Buckley,1 Ruxandra Ionescu,1 Anantha Shekhar,1 E Wesley Ely,7,8 Malaz A Boustani1–3 1Indiana University School of Medicine, 2Indiana University Center for Aging Research, 3Regenstrief Institute, Inc., 4Wishard Health Services, Indianapolis, 5Department of Pharmacy Practice, Purdue University College of Pharmacy, West Lafayette, 6Indiana University Melvin and Bren Simon Cancer Center, Indianapolis, IN, 7Vanderbilt University School of Medicine, 8VA Tennessee Valley Geriatric Research Education Clinical Center (GRECC, Nashville, TN, USA Background: Currently, there are no valid and reliable biomarkers to identify delirious patients predisposed to longer delirium duration. We investigated the hypothesis that elevated S100 calcium binding protein B (S100β levels will be associated with longer delirium duration in critically ill patients. Methods: A prospective observational cohort study was performed in the medical, surgical, and progressive intensive care units (ICUs of a tertiary care, university affiliated, and urban hospital. Sixty-three delirious patients were selected for the analysis, with two samples of S100β collected on days 1 and 8 of enrollment. The main outcome measure was delirium duration. Using the cutoff of <0.1 ng/mL and $0.1 ng/mL as normal and abnormal levels of S100β, respectively, on day 1 and day 8, four exposure groups were created: Group A, normal S100β levels on day 1 and day 8; Group B, normal S100β level on day 1 and abnormal S100β level on day 8; Group C, abnormal S100β level on day 1 and normal on day 8; and Group D, abnormal S100β levels on both day 1 and day 8. Results: Patients with abnormal levels of S100β showed a trend towards higher delirium duration (P=0.076; Group B (standard deviation (7.0 [3.2] days, Group C (5.5 [6.3] days, and Group D

  19. Calcium binding to beta-2-microglobulin at physiological pH drives the occurrence of conformational changes which cause the protein to precipitate into amorphous forms that subsequently transform into amyloid aggregates.

    Directory of Open Access Journals (Sweden)

    Sukhdeep Kumar

    Full Text Available Using spectroscopic, calorimetric and microscopic methods, we demonstrate that calcium binds to beta-2-microglobulin (β2m under physiological conditions of pH and ionic strength, in biological buffers, causing a conformational change associated with the binding of up to four calcium atoms per β2m molecule, with a marked transformation of some random coil structure into beta sheet structure, and culminating in the aggregation of the protein at physiological (serum concentrations of calcium and β2m. We draw attention to the fact that the sequence of β2m contains several potential calcium-binding motifs of the DXD and DXDXD (or DXEXD varieties. We establish (a that the microscopic aggregation seen at physiological concentrations of β2m and calcium turns into actual turbidity and visible precipitation at higher concentrations of protein and β2m, (b that this initial aggregation/precipitation leads to the formation of amorphous aggregates, (c that the formation of the amorphous aggregates can be partially reversed through the addition of the divalent ion chelating agent, EDTA, and (d that upon incubation for a few weeks, the amorphous aggregates appear to support the formation of amyloid aggregates that bind to the dye, thioflavin T (ThT, resulting in increase in the dye's fluorescence. We speculate that β2m exists in the form of microscopic aggregates in vivo and that these don't progress to form larger amyloid aggregates because protein concentrations remain low under normal conditions of kidney function and β2m degradation. However, when kidney function is compromised and especially when dialysis is performed, β2m concentrations probably transiently rise to yield large aggregates that deposit in bone joints and transform into amyloids during dialysis related amyloidosis.

  20. Blood Levels of S-100 Calcium-Binding Protein B, High-Sensitivity C-Reactive Protein, and Interleukin-6 for Changes in Depressive Symptom Severity after Coronary Artery Bypass Grafting: Prospective Cohort Nested within a Randomized, Controlled Trial (United States)

    Pearlman, Daniel M.; Brown, Jeremiah R.; MacKenzie, Todd A.; Hernandez, Felix; Najjar, Souhel


    Background Cross-sectional and retrospective studies have associated major depressive disorder with glial activation and injury as well as blood–brain barrier disruption, but these associations have not been assessed prospectively. Here, we aimed to determine the relationship between changes in depressive symptom severity and in blood levels of S-100 calcium-binding protein B (S-100B), high-sensitivity C-reactive protein, and interleukin-6 following an inflammatory challenge. Methods Fifty unselected participants were recruited from a randomized, controlled trial comparing coronary artery bypass grafting procedures performed with versus without cardiopulmonary bypass for the risk of neurocognitive decline. Depressive symptom severity was measured at baseline, discharge, and six-month follow-up using the Beck Depression Inventory II (BDI-II). The primary outcome of the present biomarker study was acute change in depressive symptom severity, defined as the intra-subject difference between baseline and discharge BDI-II scores. Blood biomarker levels were determined at baseline and 2 days postoperative. Results Changes in S-100B levels correlated positively with acute changes in depressive symptom severity (Spearman ρ, 0.62; P = 0.0004) and accounted for about one-fourth of their observed variance (R2, 0.23; P = 0.0105). This association remained statistically significant after adjusting for baseline S-100B levels, age, weight, body-mass index, or β-blocker use, but not baseline BDI-II scores (P = 0.064). There was no statistically significant association between the primary outcome and baseline S-100B levels, baseline high-sensitivity C-reactive protein or interleukin-6 levels, or changes in high-sensitivity C-reactive protein or interleukin-6 levels. Among most participants, levels of all three biomarkers were normal at baseline and markedly elevated at 2 days postoperative. Conclusions Acute changes in depressive symptom severity were specifically

  1. Structure of thrombospondin type 3 repeats in bacterial outer membrane protein A reveals its intra-repeat disulfide bond-dependent calcium-binding capability

    Energy Technology Data Exchange (ETDEWEB)

    Dai, Shuyan; Sun, Cancan; Tan, Kemin; Ye, Sheng; Zhang, Rongguang


    Eukaryotic thrombospondin type 3 repeat (TT3R) is an efficient calcium ion (Ca2+) binding motif only found in mammalian thrombospondin family. TT3R has also been found in prokaryotic cellulase Cel5G, which was thought to forfeit the Ca2+-binding capability due to the formation of intra-repeat disulfide bonds, instead of the inter-repeat ones possessed by eukaryotic TT3Rs. In this study, we have identified an enormous number of prokaryotic TT3R-containing proteins belonging to several different protein families, including outer membrane protein A (OmpA), an important structural protein connecting the outer membrane and the periplasmic peptidoglycan layer in gram-negative bacteria. Here, we report the crystal structure of the periplasmic region of OmpA from Capnocytophaga gingivalis, which contains a linker region comprising five consecutive TT3Rs. The structure of OmpA-TT3R exhibits a well-ordered architecture organized around two tightly-coordinated Ca2+ and confirms the presence of abnormal intra-repeat disulfide bonds. Further mutagenesis studies showed that the Ca2+-binding capability of OmpA-TT3R is indeed dependent on the proper formation of intra-repeat disulfide bonds, which help to fix a conserved glycine residue at its proper position for Ca2+ coordination. Additionally, despite lacking inter repeat disulfide bonds, the interfaces between adjacent OmpA-TT3Rs are enhanced by both hydrophobic and conserved aromatic-proline interactions.

  2. Calcium Binding Ability of Recombinant Buffalo Regucalcin: A Study Using Circular Dichroism Spectroscopy. (United States)

    Harikrishna, P; Thomas, Jobin; Shende, A M; Bhure, S K


    Regucalcin is a calcium regulating multifunctional protein reported to have many important functions like calcium homeostasis, anti-oxidative, anti-apoptotic and anti-cancerous functions. Although it is demonstrated as a calcium regulating protein, the calcium binding ability of regucalcin is still a controversy. The main reason for the controversy is that it lacks a typical EF hand motif which is common to most of the calcium binding proteins. Even though many studies reported regucalcin as a calcium binding protein, there are some studies reporting regucalcin as non-calcium binding also. In the present study, we investigated the calcium binding ability of recombinant buffalo regucalcin by assessing the secondary structural changes of the protein using circular dichroism spectroscopy after adding Ca(2+) to the protein solution. Two types of calcium binding studies were done, one with different concentration of calcium chloride (0.5 mM CaCl2, 1 mM CaCl2, 2 mM CaCl2) and other at different time interval (no incubation and 10 min incubation) after addition of calcium chloride. Significant structural changes were observed in both studies which prove the calcium binding ability of recombinant regucalcin. A constant increase in the α-helix (1.1% with 0.5 mM CaCl2, 1.4% with 1 mM CaCl2, 3.5% with 2 mM CaCl2) and a decrease in β-sheets (78.5% with 0.5 mM CaCl2, 77.4% with 1 mM CaCl2, 75.7% with 2 mM CaCl2) were observed with the increase in calcium chloride concentration. There was a rapid increase in α-helix and decrease in β-sheets immediately after addition of calcium chloride, which subsides after 10 min incubation.

  3. Expressão da proteína ligadora de cálcio S100 A7 (psoriasina no carcinoma laríngeo Expression of calcium binding protein S100 A7 (psoriasin in laryngeal carcinoma

    Directory of Open Access Journals (Sweden)

    Rogério Costa Tiveron


    Full Text Available Muitos estudos relatam o aumento da expressão de S100 A7 (psoriasina em lesões neoplásicas. Destacam-se trabalhos em carcinoma da mama, espinocelular da bexiga, pele e cavidade oral. Não foi demonstrada expressão da S100 A7 em câncer de laringe. OBJETIVO: Identificar a expressão da proteína ligadora de cálcio S100 A7 e sua correlação com carcinomas espinocelular da laringe. MATERIAL E MÉTODOS: Amostras de tecido neoplásico de 63 pacientes foram submetidos à imunohis toquímica com o anticorpo S110 A7. Os resultados foram classificados e comparados. RESULTADOS: O grupo bem diferenciado teve a maior pontuação de falha no tratamento. O grupo moderadamente diferenciado apresentou escores mais elevados do que o grupo pouco diferenciado. Pontuações mais altas predominaram nos estágios I e II no grupo moderadamente diferenciado, enquanto a distribuição do escore foi mais homogênea em estados avançados (III e IV. Em relação às falhas no tratamento, o grupo pontuação zero (04/03 complicações: 75% diferiu significativamente da pontuação restante (13/59: 22%. CONCLUSÕES: A S100 A7 foi expressa em 93,7% dos casos de câncer de laringe, com maior positividade nos tumores mais diferenciados e taxa significativamente menor de falha no tratamento. A pontuação obtida não teve impacto sobre a sobrevivência.Many studies have reported increased expression of S100 A7 (psoriasin in neoplastic lesions. Among them are studies on breast carcinoma, bladder squamous cell carcinoma, skin tumors and oral cavity squamous cell carcinoma. The expression of S100 A7 has not been described for laryngeal cancer. OBJECTIVE: This study aims to identify the expression of the calcium-binding protein S100 A7 and its correlation with squamous cell carcinomas of the larynx. MATERIAL AND METHODS: Specimens from 63 patients were submitted to immunohistochemistry testing with antibody S100 A7. Results were classified and compared. RESULTS: The group with

  4. Protein (Cyanobacteria): 495464111 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  5. Protein (Cyanobacteria): 230294 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  6. Calcium Binding and Disulfide Bonds Regulate the Stability of Secretagogin towards Thermal and Urea Denaturation. (United States)

    Sanagavarapu, Kalyani; Weiffert, Tanja; Ní Mhurchú, Niamh; O'Connell, David; Linse, Sara


    Secretagogin is a calcium-sensor protein with six EF-hands. It is widely expressed in neurons and neuro-endocrine cells of a broad range of vertebrates including mammals, fishes and amphibia. The protein plays a role in secretion and interacts with several vesicle-associated proteins. In this work, we have studied the contribution of calcium binding and disulfide-bond formation to the stability of the secretagogin structure towards thermal and urea denaturation. SDS-PAGE analysis of secretagogin in reducing and non-reducing conditions identified a tendency of the protein to form dimers in a redox-dependent manner. The denaturation of apo and Calcium-loaded secretagogin was studied by circular dichroism and fluorescence spectroscopy under conditions favoring monomer or dimer or a 1:1 monomer: dimer ratio. This analysis reveals significantly higher stability towards urea denaturation of Calcium-loaded secretagogin compared to the apo protein. The secondary and tertiary structure of the Calcium-loaded form is not completely denatured in the presence of 10 M urea. Reduced and Calcium-loaded secretagogin is found to refold reversibly after heating to 95°C, while both oxidized and reduced apo secretagogin is irreversibly denatured at this temperature. Thus, calcium binding greatly stabilizes the structure of secretagogin towards chemical and heat denaturation.

  7. Calcium binding and homoassociation of E-cadherin domains. (United States)

    Koch, A W; Pokutta, S; Lustig, A; Engel, J


    Cadherins are single pass transmembrane glycoproteins which mediate calcium dependent cell-cell adhesion by homophilic interactions. To reveal the molecular details of calcium binding and homoassociation, we recombinantly expressed in Escherichia coli a domain pair consisting of the first two domains of E-cadherin (ECAD12) and the single domains 1, 2, and 5. ECAD12 encompasses the most N-terminal of the four putative calcium-binding pockets in the extracellular region of E-cadherin. Equilibrium dialysis experiments revealed that the single domains do not bind Ca2+, but ECAD12 was found to bind three calcium ions. ECAD12 dimerizes (Kd = 0.08 +/- 0.02 mM) in the presence of Ca2+ as we could demonstrate by analytical ultracentrifugation. Calcium binding to ECAD12 induces conformational changes which were monitored by electrophoretic mobility and by circular dichroism. By analyzing our equilibrium dialysis data with a single binding site model, we found an average Kd of 460 microM for the three bound Ca2+. Assuming a model for three binding sites, which slightly increased the quality of the fit, we obtained two identical Kds of 330 microM and a third much higher Kd of 2 mM. The entire extracellular region of E-cadherin, which was recombinantly expressed in mammalian cells, binds nine Ca2+ with a much lower average Kd of 30 microM. Therefore, we conclude that the four calcium binding pockets are not identical. Since binding to ECAD12 occurs at Ca2+ concentrations close to those in the extracellular space, we suggest that the N-terminal domain pair might be involved in calcium regulation of E-cadherin mediated cell-cell adhesion.

  8. Kinetics of calcium binding to dental biofilm bacteria. (United States)

    Leitão, Tarcísio Jorge; Cury, Jaime Aparecido; Tenuta, Livia Maria Andaló


    Dental biofilm bacteria can bind calcium ions and release them during a pH drop, which could decrease the driving force for dental demineralization (i.e. hydroxyapatite dissolution) occurring at reduced pHs. However, the kinetics of this binding and release is not completely understood. Here we validated a method to evaluate the kinetics of calcium binding and release to/from Streptococcus mutans, and estimated the importance of this reservoir as a source of ions. The kinetics of calcium binding was assessed by measuring the amount of bound calcium in S. mutans Ingbrit 1600 pellets treated with PIPES buffer, pH 7.0, containing 1 or 10 mM Ca; for the release kinetics, bacterial pellets previously treated with 1 mM or 10 mM Ca were exposed to the calcium-free or 1 mM Ca PIPES buffer, pH 7.0, for up to 60 min. Binding and release curves were constructed and parameters of kinetics were calculated. Also, calcium release was assessed by exposing pellets previously treated with calcium to a pH 5.0 buffer for 10 min. Calcium binding to bacteria was concentration-dependent and rapid, with maximum binding reached at 5 min. On the other hand, calcium release was slower, and according to the calculations, would never be complete in the groups pretreated with 10 mM Ca. Decreasing pH from 7.0 to 5.0 caused a release of calcium able to increase the surrounding fluid calcium concentration in 2 mM. The results suggest that dental biofilm bacteria may act as a calcium reservoir, rapidly binding ions from surrounding fluids, releasing them slowly at neutral pH and promptly during a pH drop.

  9. Calcium-binding capacity of centrin2 is required for linear POC5 assembly but not for nucleotide excision repair.

    Directory of Open Access Journals (Sweden)

    Tiago J Dantas

    Full Text Available Centrosomes, the principal microtubule-organising centres in animal cells, contain centrins, small, conserved calcium-binding proteins unique to eukaryotes. Centrin2 binds to xeroderma pigmentosum group C protein (XPC, stabilising it, and its presence slightly increases nucleotide excision repair (NER activity in vitro. In previous work, we deleted all three centrin isoforms present in chicken DT40 cells and observed delayed repair of UV-induced DNA lesions, but no centrosome abnormalities. Here, we explore how centrin2 controls NER. In the centrin null cells, we expressed centrin2 mutants that cannot bind calcium or that lack sites for phosphorylation by regulatory kinases. Expression of any of these mutants restored the UV sensitivity of centrin null cells to normal as effectively as expression of wild-type centrin. However, calcium-binding-deficient and T118A mutants showed greatly compromised localisation to centrosomes. XPC recruitment to laser-induced UV-like lesions was only slightly slower in centrin-deficient cells than in controls, and levels of XPC and its partner HRAD23B were unaffected by centrin deficiency. Interestingly, we found that overexpression of the centrin interactor POC5 leads to the assembly of linear, centrin-dependent structures that recruit other centrosomal proteins such as PCM-1 and NEDD1. Together, these observations suggest that assembly of centrins into complex structures requires calcium binding capacity, but that such assembly is not required for centrin activity in NER.

  10. Conformational changes of the recombinant extracellular domain of E-cadherin upon calcium binding. (United States)

    Pokutta, S; Herrenknecht, K; Kemler, R; Engel, J


    The cell-adhesion protein E-cadherin/uvomorulin exhibits a calcium-dependent homoassociation. The effect of Ca2+ on the extracellular fragment of E-cadherin was studied using the recombinant protein expressed in the baculovirus expression system. The recombinant and native fragment of E-cadherin were found to be similar by many biochemical criteria [Herrenknecht, K. & Kemler, R. (1993) J. Cell Sci. 17, 147-154]. A large and reversible conformational transition was observed upon Ca2+ depletion. A change from a rod-like structure, 22 nm in length, to a more globular assembly of the five subdomains became evident by electron-microscopical analysis. In the presence of Ca2+, the circular dichroic spectra indicated predominantly beta-structure but a more negative ellipticity was observed in the absence of Ca2+. The intrinsic tryptophan fluorescence decreased by 12% upon Ca2+ depletion. Both effects were used for calcium titrations which indicated calcium binding to several sites with average K(d) values of 45-150 microM. Cleavage of the protein fragment by trypsin occurred only at low Ca2+ concentrations and from the calcium-dependence of cleavage rates, a K(d) value of 24 microM was derived. The major site of cleavage was identified by partial sequencing to be located between the two putative calcium-binding sites in the third subdomain from the N-terminus. In agreement with earlier results with the native fragment, the recombinant protein did not associate in the presence or absence of Ca2+. We suggest the calcium-dependent homoassociation therefore depends on additional effects connected with the cell surface association of E-cadherin.

  11. Involvement of SNP marker located on the Calcium binding protein ...

    African Journals Online (AJOL)

    Olive trees importance is mainly due to the economic and health benefits, especially in the Mediterranean basin. Unfortunately, to enhance productivity and quality of olive oil, the study of both molecular and phenotypic characterizations of olive cultivars is crucial. We consider the analysis of 14 Tunisian olive cultivars of ...

  12. Fragment molecular orbital method for studying lanthanide interactions with proteins

    Energy Technology Data Exchange (ETDEWEB)

    Tsushima, Satoru [Helmholtz-Zentrum Dresden-Rossendorf e.V., Dresden (Germany). Biophysics; Komeiji, Y. [National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba (Japan); Mochizuki, Y. [Rikkyo Univ., Tokyo (Japan)


    The binding affinity of the calcium-binding protein calmodulin towards Eu{sup 3+} was studied as a model for lanthanide protein interactions in the large family of ''EF-hand'' calcium-binding proteins.

  13. The early asthmatic response is associated with glycolysis, calcium binding and mitochondria activity as revealed by proteomic analysis in rats

    Directory of Open Access Journals (Sweden)

    Xu Yu-Dong


    Full Text Available Abstract Background The inhalation of allergens by allergic asthmatics results in the early asthmatic response (EAR, which is characterized by acute airway obstruction beginning within a few minutes. The EAR is the earliest indicator of the pathological progression of allergic asthma. Because the molecular mechanism underlying the EAR is not fully defined, this study will contribute to a better understanding of asthma. Methods In order to gain insight into the molecular basis of the EAR, we examined changes in protein expression patterns in the lung tissue of asthmatic rats during the EAR using 2-DE/MS-based proteomic techniques. Bioinformatic analysis of the proteomic data was then performed using PPI Spider and KEGG Spider to investigate the underlying molecular mechanism. Results In total, 44 differentially expressed protein spots were detected in the 2-DE gels. Of these 44 protein spots, 42 corresponded to 36 unique proteins successfully identified using mass spectrometry. During subsequent bioinformatic analysis, the gene ontology classification, the protein-protein interaction networking and the biological pathway exploration demonstrated that the identified proteins were mainly involved in glycolysis, calcium binding and mitochondrial activity. Using western blot and semi-quantitative RT-PCR, we confirmed the changes in expression of five selected proteins, which further supports our proteomic and bioinformatic analyses. Conclusions Our results reveal that the allergen-induced EAR in asthmatic rats is associated with glycolysis, calcium binding and mitochondrial activity, which could establish a functional network in which calcium binding may play a central role in promoting the progression of asthma.

  14. Calcium binding to low molecular weight compounds and health promoting products

    DEFF Research Database (Denmark)

    Vavrusova, Martina

    absorption. Therefore, calcium as an essential nutrient should not be underestimated in our diet. Milk and dairy products are good sources of bioavailable calcium due to specific protein binding. Other sources of calcium, apart from a balanced and healthy diet, are calcium supplements and calcium fortified......Calcium precipitation in the almost neutral environment of the intestines is a process related to weight loss management and plays an important role in the prevention of colon cancer development. This process also affects calcium bioavailability which is decreased due to decreased calcium...... food. Therefore, an understanding of the basic chemistry of calcium binding to low molecular weight compounds can contribute to a general knowledge about calcium bioavailability and also to product improvement. Calcium precipitation with palmitate was described by a first-order reaction for conditions...

  15. Protein (Cyanobacteria): 428310068 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  16. Protein (Cyanobacteria): 493576985 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  17. Protein (Cyanobacteria): 427729770 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  18. Calcium binding to low molecular weight compounds and health promoting products

    DEFF Research Database (Denmark)

    Vavrusova, Martina

    to acidic amino acids and dipeptides was also investigated. It was found that aspartate binds calcium stronger than glutamate. This was confirmed by electrochemical determination of the association constants for aqueous solutions at ionic strength 0.20 and 1.0. The mixed dipeptides Asp/Glu showed additive...... affinity of calcium binding as determined by the association constants and also compared with the values predicted from the individual amino acids. In contrast, the AspAsp dipeptide showed an affinity less than additive, while the GluGlu dipeptide showed affinity of calcium binding to be more than additive....... Additionally, the affinity of calcium to serine was small but was increased for the SerGlu dipeptide and the values became comparable to phosphorylated serine. An increase in ionic strength lead to decreased calcium binding of the studied free amino acids, dipeptides and phosphorylated serine as expected...

  19. Two structural motifs within canonical EF-hand calcium-binding domains identify five different classes of calcium buffers and sensors.

    Directory of Open Access Journals (Sweden)

    Konstantin Denessiouk

    Full Text Available Proteins with EF-hand calcium-binding motifs are essential for many cellular processes, but are also associated with cancer, autism, cardiac arrhythmias, and Alzheimer's, skeletal muscle and neuronal diseases. Functionally, all EF-hand proteins are divided into two groups: (1 calcium sensors, which function to translate the signal to various responses; and (2 calcium buffers, which control the level of free Ca2+ ions in the cytoplasm. The borderline between the two groups is not clear, and many proteins cannot be described as definitive buffers or sensors. Here, we describe two highly-conserved structural motifs found in all known different families of the EF-hand proteins. The two motifs provide a supporting scaffold for the DxDxDG calcium binding loop and contribute to the hydrophobic core of the EF hand domain. The motifs allow more precise identification of calcium buffers and calcium sensors. Based on the characteristics of the two motifs, we could classify individual EF-hand domains into five groups: (1 Open static; (2 Closed static; (3 Local dynamic; (4 Dynamic; and (5 Local static EF-hand domains.

  20. Protein (Cyanobacteria): 428204399 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  1. Protein (Cyanobacteria): 493575929 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  2. Protein (Cyanobacteria): 493554048 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  3. Protein (Cyanobacteria): 218438228 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  4. An explicitly solvated full atomistic model of the cardiac thin filament and application on the calcium binding affinity effects from familial hypertrophic cardiomyopathy linked mutations (United States)

    Williams, Michael; Schwartz, Steven


    The previous version of our cardiac thin filament (CTF) model consisted of the troponin complex (cTn), two coiled-coil dimers of tropomyosin (Tm), and 29 actin units. We now present the newest revision of the model to include explicit solvation. The model was developed to continue our study of genetic mutations in the CTF proteins which are linked to familial hypertrophic cardiomyopathies. Binding of calcium to the cTnC subunit causes subtle conformational changes to propagate through the cTnC to the cTnI subunit which then detaches from actin. Conformational changes propagate through to the cTnT subunit, which allows Tm to move into the open position along actin, leading to muscle contraction. Calcium disassociation allows for the reverse to occur, which results in muscle relaxation. The inclusion of explicit TIP3 water solvation allows for the model to get better individual local solvent to protein interactions; which are important when observing the N-lobe calcium binding pocket of the cTnC. We are able to compare in silica and in vitro experimental results to better understand the physiological effects from mutants, such as the R92L/W and F110V/I of the cTnT, on the calcium binding affinity compared to the wild type.

  5. Structure and calcium binding activity of LipL32, the major surface antigen of pathogenic Leptospira sp

    Energy Technology Data Exchange (ETDEWEB)

    Hauk, Pricila; Roman-Ramos, Henrique; Ho, Paulo Lee [Instituto Butantan, Sao Paulo, SP (Brazil). Centro de Biotecnologia; Guzzo, Cristiane R.; Farah, Chuck S. [Universidade de Sao Paulo (USP), SP (Brazil). Inst. de Quimica. Dept. de Bioquimica


    Leptospirosis, caused by the spirochaete Leptospira is an important emerging infectious disease. LipL32 is the major exposed outer membrane protein found exclusively in pathogenic leptospira. It is highly immunogenic and has been shown to bind to host extracellular matrix components, including collagens, fibronectin and laminin. In this work we crystallized recombinant LipL32 protein and determined its structure to 2.25 A resolution. Initial phases were determined using the multi-wavelength anomalous dispersion technique with data collected from selenomethionine-containing crystals at the MX2 beamline at the LNLS. The LipL32 monomer is made of a jelly-roll fold core from which protrude several peripheral secondary structures. Some structural features suggested that LipL32 could bind Ca{sup 2+} ions and indeed, spectroscopic data (circular (dichroism. intrinsic tryptophan fluorescence and extrinsic 1-amino-2-anaphthol-4-sulfonic acid fluorescence) confirmed the calcium binding properties of LipL32. (author)

  6. Accesion number Protein name ENOA_MOUSE Alpha-enolase ...

    Indian Academy of Sciences (India)

    Sandra Feijoo Bandin

    Mitochondrial inner membrane protein. CMC1_MOUSE. Calcium-binding mitochondrial carrier protein Aralar1. CMC2_MOUSE. Calcium-binding mitochondrial carrier protein Aralar2. Biological process. Metabolic process. Glycolysis. Lipid metabolism. Respiratory electron transport chain. Others. Calcium ion homeostasis.

  7. Structural investigations of calcium binding and its role in activity and activation of outer membrane phospholipase A from Escherichia coli

    NARCIS (Netherlands)

    Snijder, H.J.; Kingma, R.L.; Kalk, K.H.; Dekker, N.; Egmond, M.R.|info:eu-repo/dai/nl/112946593; Dijkstra, B.W.


    Outer membrane phospholipase A (OMPLA) is an integral membrane enzyme that catalyses the hydrolysis of phospholipids. Enzymatic activity is regulated by reversible dimerisation and calcium-binding. We have investigated the role of calcium by X-ray crystallography. In monomeric OMPLA, one calcium ion

  8. Anticoagulant and calcium-binding properties of high molecular weight derivatives of human fibrinogen (plasmin fragments Y)

    NARCIS (Netherlands)

    Nieuwenhuizen, W.; Voskuilen, M.; Hermans, J.


    The present study was undertaken as a step to delineate further the localization of the calcium-binding sites in fibrinogen and to assess the anticlotting properties of fibrinogen degradation products. To this purpose, fragments Y were prepared by plasmin digestion of human fibrinogen in the

  9. Recombinant γT305A fibrinogen indicates severely impaired fibrin polymerization due to the aberrant function of hole 'A' and calcium binding sites. (United States)

    Ikeda, Minami; Kobayashi, Tamaki; Arai, Shinpei; Mukai, Saki; Takezawa, Yuka; Terasawa, Fumiko; Okumura, Nobuo


    We examined a 6-month-old girl with inherited fibrinogen abnormality and no history of bleeding or thrombosis. Routine coagulation screening tests showed a markedly low level of plasma fibrinogen determined by functional measurement and also a low level by antigenic measurement (functional/antigenic ratio=0.295), suggesting hypodysfibrinogenemia. DNA sequence analysis was performed, and γT305A fibrinogen was synthesized in Chinese hamster ovary cells based on the results. We then functionally analyzed and compared with that of nearby recombinant γN308K fibrinogen. DNA sequence analysis revealed a heterozygous γT305A substitution (mature protein residue number). The γT305A fibrinogen indicated markedly impaired thrombin-catalyzed fibrin polymerization both in the presence or absence of 1mM calcium ion compared with that of γN308K fibrinogen. Protection of plasmin degradation in the presence of calcium ion or Gly-Pro-Arg-Pro peptide (analogue for so-called knob 'A') and factor XIIIa-catalyzed fibrinogen crosslinking demonstrated that the calcium binding sites, hole 'a' and D:D interaction sites were all markedly impaired, whereas γN308Kwas impaired at the latter two sites. Molecular modeling demonstrated that γT305 is localized at a shorter distance than γN308 from the high affinity calcium binding site and hole 'a'. Our findings suggest that γT305 might be important for construction of the overall structure of the γ module of fibrinogen. Substitution of γT305A leads to both dysfibrinogenemic and hypofibrinogenemic characterization, namely hypodysfibrinogenemia. We have already reported that recombinant γT305A fibrinogen was synthesized normally and secreted slightly, but was significantly reduced. Copyright © 2014 Elsevier Ltd. All rights reserved.

  10. Antisense expression of a gene encoding a calcium-binding protein ...

    Indian Academy of Sciences (India)


    and, after washing, the blots were incubated with secondary antibodies conjugated to alkaline phosphatase or horseradish peroxidase and developed by BCIP/NBT substrate or ECL. (Amersham Bioscience, NJ, USA). 2.3 Transmission electron microscopy (TEM). Immersion fixation of leaf tissues for ultrastructural studies.

  11. Calcium-binding protein, secretagogin, characterizes novel groups of interneurons in the rat striatum. (United States)

    Kosaka, Toshio; Yasuda, Seiko; Kosaka, Katsuko


    In the rat striatum numerous secretagogin (SCGN) positive neurons were scattered. They were heterogeneous in their morphological and chemical properties. We examined the colocalization of SCGN with known four interneuron markers, parvalbumin (PV), calretinin (CR), nitric oxide synthase (NOS) and choline acetyl transferase (ChAT). 60-70% of SCGN positive striatal neurons contained either PV or CR or ChAT, but none contained NOS. On the other hand the remaining 30-40% expressed none of these markers, most of which were GAD positive. The present study indicates that there are hitherto unknown groups of striatal interneurons in the rat striatum. Copyright © 2017 Elsevier Ireland Ltd and Japan Neuroscience Society. All rights reserved.

  12. Effect of calcium-binding peptide from Pacific cod (Gadus macrocephalus) bone on calcium bioavailability in rats. (United States)

    Peng, Zhe; Hou, Hu; Zhang, Kai; Li, Bafang


    Bone collagen peptide with high affinity to Ca was extracted from Pacific cod (Gadus macrocephalus) bone. FTIR spectra of calcium-binding bone collagen peptide showed that band at 3381cm(-1) shifted to 3361cm(-1), 1455cm(-1) moved to 1411cm(-1), and amide II became deeper valley, compared with that of bone collagen peptide. This peptide was sequenced by Q-TOF-MS and sequences of Gly-Pro-Glu-Gly, Gly-Glu-Lys, Gly-Pro-Leu-Gly and Gly-Leu-Pro-Gly appeared repeatedly in some peptides. From SEM, after chelated with calcium, the loose and porous structure turned into granular structure. From the animal experiment, Ca apparent absorption rate, Ca retention rate and femur Ca content of calcium-binding bone collagen peptide group were significantly higher than those of model and CaCO3 groups (P<0.05), while serum ALP was significantly lower than model group (P<0.05) and similar to control group. The results suggested that calcium-binding bone collagen peptide could improve bioavailability of Ca and thus prevented Ca deficiency. Copyright © 2016 Elsevier Ltd. All rights reserved.

  13. Ionized calcium-binding adaptor molecule 1 positive macrophages and HO-1 up-regulation in intestinal muscularis resident macrophages

    DEFF Research Database (Denmark)

    Mikkelsen, Hanne B; Huizinga, Jan D; Larsen, Jytte O


    the reaction of resident macrophages of the musculature to a pro-inflammatory stimulator, lipopolysaccharide (LPS). Mice were injected with LPS or saline and sacrificed after 6 hours. Whole mounts were stained with antibodies toward CD169, ionized calcium-binding adaptor molecule 1 (iba1) (microglial...... macrophages in serosa and at AP, suggesting a M2 phenotype. LPS-treatment results in an up-regulation of HO-1(pos) /CD169(neg) cells in serosa and at AP. This article is protected by copyright. All rights reserved....

  14. Proteomic identification of calcium-binding chaperone calreticulin as a potential mediator for the neuroprotective and neuritogenic activities of fruit-derived glycoside amygdalin. (United States)

    Cheng, Yuanyuan; Yang, Chuanbin; Zhao, Jia; Tse, Hung Fat; Rong, Jianhui


    Amygdalin is a fruit-derived glycoside with the potential for treating neurodegenerative diseases. This study was designed to identify the neuroprotective and neuritogenic activities of amygdalin. We initially demonstrated that amygdalin enhanced nerve growth factor (NGF)-induced neuritogenesis and attenuated 6-hydroxydopamine (6-OHDA)-induced neurotoxicity in rat dopaminergic PC12 cells. To define protein targets for amygdalin, we selected a total of 11 mostly regulated protein spots from two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels for protein identification by matrix-assisted laser desorption/ionization-time-of-flight-mass spectrometry. We verified the effect of amygdalin on six representative proteins (i.e., calreticulin, Hsp90β, Grp94, 14-3-3η, 14-3-3ζ/δ and Rab GDI-α) for biological relevance to neuronal survival and differentiation. Calcium-binding chaperone calreticulin is of special interest for its activities to promote folding, oligomeric assembly and quality control of proteins that modulate cell survival and differentiation. We transiently knocked down calreticulin expression by specific siRNA and studied its effect on the neuroprotective and neuritogenic activities of amygdalin. We found that amygdalin failed to enhance NGF-induced neuritogenesis in calreticulin-siRNA transfected cells. On the other hand, amygdalin rescued 6-OHDA-induced loss of calreticulin expression. We also found that amygdalin increased the intracellular calcium concentration possibly via inducing calreticulin. Collectively, our results demonstrated the role of calreticulin in mediating the neuroprotective and neuritogenic activities of amygdalin. Copyright © 2015 Elsevier Inc. All rights reserved.

  15. Structural Changes in the Lectin Domain of CD23, the Low-Affinity IgE Receptor, upon Calcium Binding

    Energy Technology Data Exchange (ETDEWEB)

    Wurzburg, Beth A.; Tarchevskaya, Svetlana S.; Jardetzky, Theodore S. (NWU)


    CD23, the low-affinity receptor for IgE (Fc{var_epsilon}RII), regulates IgE synthesis and also mediates IgE-dependent antigen transport and processing. CD23 is a unique Fc receptor belonging to the C-type lectin-like domain superfamily and binds IgE in an unusual, non-lectin-like manner, requiring calcium but not carbohydrate. We have solved the high-resolution crystal structures of the human CD23 lectin domain in the presence and absence of Ca{sup 2+}. The crystal structures differ significantly from a previously determined NMR structure and show that calcium binding occurs at the principal binding site, but not at an auxiliary site that appears to be absent in human CD23. Conformational differences between the apo and Ca{sup 2+} bound structures suggest how IgE-Fc binding can be both calcium-dependent and carbohydrate-independent.

  16. The PEF family proteins sorcin and grancalcin interact in vivo and in vitro

    DEFF Research Database (Denmark)

    Hansen, Christian; Tarabykina, Svetlana; la Cour, Jonas Marstrand


    The penta-EF hand (PEF) family of calcium binding proteins includes grancalcin, peflin, sorcin, calpain large and small subunits as well as ALG-2. Systematic testing of the heterodimerization abilities of the PEF proteins using the yeast two-hybrid and glutathione S-transferase pull-down assays...... be a way to regulate and fine tune processes mediated by calcium binding proteins of the penta-EF hand type....

  17. Isolation and characterization of calcium binding glycoproteins of cardiac sarcolemmal vesicles

    Energy Technology Data Exchange (ETDEWEB)

    Michalak, M.; Fliegel, L.; Wlasichuk, K. (Univ. of Alberta, Edmonton (Canada))


    Two major Ca2(+)-binding glycoproteins Mr 120,000 and 100,000 were isolated from 3-((3-cholamidopropyl)dimethylammonio)-1-propanesulfonic acid -solubilized bovine heart sarcolemma membrane. Peroxidase-conjugated concanavalin A and wheat germ agglutinin lectins bind strongly to the isolated 120- and 100-kDa glycoproteins. Treatment with endoglycosidase F resulted in conversion of the 120-kDa glycoprotein to a form migrating at about 97 kDa. Treatment of the 100-kDa band with endoglycosidase F produced form of about 80 kDa. Endoglycosidase H digestion removes only 5% of the mass of both glycoproteins. the carbohydrate structure of both glycoproteins, is therefore, predicted to be at least 75% complex structure and 25% high mannose or hybrid structure. The 120- and 100-kDa glycoproteins are the major Ca2(+)-binding proteins in the sarcolemma membranes. Intact and endoglycosidase-treated glycoproteins bind 45Ca2+ as analyzed by a 45Ca2+ overlay technique. Using polyclonal antibodies, the 120- and 100-kDa glycoproteins were identified in muscle plasma membranes (ventricles, atria, and uterus smooth muscle). They were, however, not present in non-muscle tissues such as pancreas, liver, and kidney. The 120- and 100-kDa glycoproteins appear to be homologous molecules as judged by their similar V8 protease peptide maps, cross-reactivity with polyclonal antibody, and other physicochemical properties.

  18. Expression of calcium-binding proteins and nNOS in the human vestibular and precerebellar brainstem. (United States)

    Baizer, Joan S; Broussard, Dianne M


    Information about the position and movement of the head in space is coded by vestibular receptors and relayed to four nuclei that comprise the vestibular nuclear complex (VNC). Many additional brainstem nuclei are involved in the processing of vestibular information, receiving signals either directly from the eighth nerve or indirectly via projections from the VNC. In cats, squirrel monkeys, and macaque monkeys, we found neurochemically defined subdivisions within the medial vestibular nucleus (MVe) and within the functionally related nucleus prepositus hypoglossi (PrH). In humans, different studies disagree about the borders, sizes, and possible subdivisions of the vestibular brainstem. In an attempt to clarify this organization, we have begun an analysis of the neurochemical characteristics of the human using brains from the Witelson Normal Brain Collection and standard techniques for antigen retrieval and immunohistochemistry. Using antibodies to calbindin, calretinin, parvalbumin, and nitric oxide synthase, we find neurochemically defined subdivisions within the MVe similar to the subdivisions described in cats and monkeys. The neurochemical organization of PrH is different. We also find unique neurochemical profiles for several structures that suggest reclassification of nuclei. These data suggest both quantitative and qualitative differences among cats, monkeys, and humans in the organization of the vestibular brainstem. These results have important implications for the analysis of changes in that organization subsequent to aging, disease, or loss of input.

  19. The calcium-binding protein parvalbumin modulates the firing 1 properties of the reticular thalamic nucleus bursting neurons. (United States)

    Albéri, Lavinia; Lintas, Alessandra; Kretz, Robert; Schwaller, Beat; Villa, Alessandro E P


    The reticular thalamic nucleus (RTN) of the mouse is characterized by an overwhelming majority of GABAergic neurons receiving afferences from both the thalamus and the cerebral cortex and sending projections mainly on thalamocortical neurons. The RTN neurons express high levels of the "slow Ca(2+) buffer" parvalbumin (PV) and are characterized by low-threshold Ca(2+) currents, I(T). We performed extracellular recordings in ketamine/xylazine anesthetized mice in the rostromedial portion of the RTN. In the RTN of wild-type and PV knockout (PVKO) mice we distinguished four types of neurons characterized on the basis of their firing pattern: irregular firing (type I), medium bursting (type II), long bursting (type III), and tonically firing (type IV). Compared with wild-type mice, we observed in the PVKOs the medium bursting (type II) more frequently than the long bursting type and longer interspike intervals within the burst without affecting the number of spikes. This suggests that PV may affect the firing properties of RTN neurons via a mechanism associated with the kinetics of burst discharges. Ca(v)3.2 channels, which mediate the I(T) currents, were more localized to the somatic plasma membrane of RTN neurons in PVKO mice, whereas Ca(v)3.3 expression was similar in both genotypes. The immunoelectron microscopy analysis showed that Ca(v)3.2 channels were localized at active axosomatic synapses, thus suggesting that the differential localization of Ca(v)3.2 in the PVKOs may affect bursting dynamics. Cross-correlation analysis of simultaneously recorded neurons from the same electrode tip showed that about one-third of the cell pairs tended to fire synchronously in both genotypes, independent of PV expression. In summary, PV deficiency does not affect the functional connectivity between RTN neurons but affects the distribution of Ca(v)3.2 channels and the dynamics of burst discharges of RTN cells, which in turn regulate the activity in the thalamocortical circuit.

  20. Calcium ion-protein interactions in prothrombin activation

    Energy Technology Data Exchange (ETDEWEB)

    Brenckle, G.M.; Carlisle, T.L.; Jackson, C.M.


    1. The protein concentration dependence observed in the calcium binding to fragment 1 indicates that calcium-mediated dimerization is responsible for the cooperative calcium binding behavior usually observed. ''Unusual'' fragment 1, which exhibits negative cooperativity (the type of binding behavior expected for ions interacting with a charged protein) at high concentration, also exhibit altered self-association behavior. 2. The calcium-induced spectral perturbations that are observed by fluorescence and ultraviolet difference spectroscopy are influenced by calcium-mediated dimerization. Similar spectral perturbations may also be induced by other divalent, trivalent, and monovalent ions, as well as changes in pH. Because this is a multi-site system, only limited interpretation of the spectral data is possible without calcium binding data. 3. Although strong side chain CD signals make estimation of fragment 1 secondary structure ambiguous, the CD data do indicate small changes in structure during calcium binding. Similar changes are observed upon addition of monovalent ions at high concentration or after lowering the pH. No coupling between changes in conformation and the cooperative calcium binding behavior has yet been observed to exist.

  1. Expression of the high capacity calcium-binding domain of calreticulin increases bioavailable calcium stores in plants (United States)

    Wyatt, Sarah E.; Tsou, Pei-Lan; Robertson, Dominique; Brown, C. S. (Principal Investigator)


    Modulation of cytosolic calcium levels in both plants and animals is achieved by a system of Ca2+-transport and storage pathways that include Ca2+ buffering proteins in the lumen of intracellular compartments. To date, most research has focused on the role of transporters in regulating cytosolic calcium. We used a reverse genetics approach to modulate calcium stores in the lumen of the endoplasmic reticulum. Our goals were two-fold: to use the low affinity, high capacity Ca2+ binding characteristics of the C-domain of calreticulin to selectively increase Ca2+ storage in the endoplasmic reticulum, and to determine if those alterations affected plant physiological responses to stress. The C-domain of calreticulin is a highly acidic region that binds 20-50 moles of Ca2+ per mole of protein and has been shown to be the major site of Ca2+ storage within the endoplasmic reticulum of plant cells. A 377-bp fragment encoding the C-domain and ER retention signal from the maize calreticulin gene was fused to a gene for the green fluorescent protein and expressed in Arabidopsis under the control of a heat shock promoter. Following induction on normal medium, the C-domain transformants showed delayed loss of chlorophyll after transfer to calcium depleted medium when compared to seedlings transformed with green fluorescent protein alone. Total calcium measurements showed a 9-35% increase for induced C-domain transformants compared to controls. The data suggest that ectopic expression of the calreticulin C-domain increases Ca2+ stores, and that this Ca2+ reserve can be used by the plant in times of stress.

  2. Enhanced expression of a calcium-dependent protein kinase from ...

    Indian Academy of Sciences (India)

    Among the downstream targets of calcium in plants, calcium-dependent protein kinases (CDPKs) form an interesting class of kinases which are activated by calcium binding. They have been implicated in a diverse array of responses to hormonal and environmental stimuli. In order to dissect the role of CDPKs in the moss ...

  3. Localization of Nitric Oxide Synthase-containing Neurons in the Bat Visual Cortex and Co-localization with Calcium-binding Proteins. (United States)

    Gu, Ya-Nan; Kim, Hang-Gu; Jeon, Chang-Jin


    Microchiroptera (microbats) is a suborder of bats thought to have degenerated vision. However, many recent studies have shown that they have visual ability. In this study, we labeled neuronal nitric oxide synthase (nNOS)-the synthesizing enzyme of the gaseous non-synaptic neurotransmitter nitric oxide-and co-localized it with calbindin D28K (CB), calretinin (CR), and parvalbumin (PV) in the visual cortex of the greater horseshoe bat (Rhinolophus ferrumequinum, a species of microbats). nNOS-immunoreactive (IR) neurons were found in all layers of the visual cortex. Intensely labeled neurons were most common in layer IV, and weakly labeled neurons were most common in layer VI. Majority of the nNOS-IR neurons were round- or oval-type neurons; no pyramidal-type neurons were found. None of these neurons co-localized with CB, CR, or PV. However, the synthesis of nitric oxide in the bat visual cortex by nNOS does not depend on CB, CR, or PV.

  4. Visual and motor connectivity and the distribution of calcium-binding proteins in macaque frontal eye field: implications for saccade target selection

    Directory of Open Access Journals (Sweden)

    Pierre Pouget


    Full Text Available The frontal eye field (FEF contributes to directing visual attention and saccadic eye movement through intrinsic processing, interactions with extrastriate visual cortical areas (e.g. V4, and projections to subcortical structures (e.g. superior colliculus; SC. Several models have been proposed to describe the relationship between the allocation of visual attention and the production of saccades. We obtained anatomical information that might provide useful constraints on these models by evaluating two characteristics of FEF. First, we investigated the laminar distribution of efferent connections from FEF to visual areas V4 + TEO and to SC. Second, we examined the laminar distribution of different populations of GABAergic neurons in FEF. We found that the neurons in FEF that project to V4 + TEO are located predominantly in the supragranular layers, colocalized with the highest density of calbindin- and calretinin-immunoreactive inhibitory interneurons. In contrast, the cell bodies of neurons that project to SC are found only in layer 5 of FEF, colocalized primarily with parvalbumin inhibitory interneurons. None of the neurons in layer 5 that project to V4 + TEO also project to SC. These results provide useful constraints for cognitive models of visual attention and saccade production by indicating that different populations of neurons project to extrastriate visual cortical areas and to SC. This finding also suggests that FEF neurons projecting to visual cortex and superior colliculus are embedded in different patterns of intracortical circuitry.

  5. Polypepide synthesis in response to 20-hydroxyecdysone, with particular reference to calcium-binding proteins, in the epidermis of a larval insect

    Energy Technology Data Exchange (ETDEWEB)

    Ouellette, Y.


    Epidermal cells respond to 20-hydroxyecdysone (20HE) by altering their morphology, rate of proliferation and level of cell-to-cell communication. To study changes in polypeptide synthesis induced by 20HE, epidermal polypeptides from the midinstar larvae of the mealworm Tenebrio molitor were separated by one- and two-dimensional polyacrylamide gel electrophoresis. {sup 35}S-methionine-labelled polypeptides were extracted from cells incubated either in the absence or presence of 2{mu}g/ml 20HE for 14-16 h in vitro. Cells incubated with the hormone incorporated 28% more label into polypeptides. Over 250 polypeptides were detected in total cell lysates by this technique. Polypeptides that showed altered rates of synthesis in response to 20HE treatment were identified and characterized.

  6. Epidermal growth factor receptor ligands as new extracellular targets for the metastasis-promoting S100A4 protein

    DEFF Research Database (Denmark)

    Klingelhöfer, Jörg; Møller, Henrik D.; Sumer, Eren U


    The function of S100A4, a member of the calcium-binding S100 protein family, has been associated with tumor invasion and metastasis. Although an essential pro-metastatic role of extracellular S100A4 in tumor progression has been demonstrated, the identification of the precise underlying mechanisms...

  7. Protein kinase C interaction with calcium: a phospholipid-dependent process.

    LENUS (Irish Health Repository)

    Bazzi, M D


    The calcium-binding properties of calcium- and phospholipid-dependent protein kinase C (PKC) were investigated by equilibrium dialysis in the presence and the absence of phospholipids. Calcium binding to PKC displayed striking and unexpected behavior; the free proteins bound virtually no calcium at intracellular calcium concentrations and bound limited calcium (about 1 mol\\/mol of PKC) at 200 microM calcium. However, in the presence of membranes containing acidic phospholipids, PKC bound at least eight calcium ions per protein. The presence of 1 microM phorbol dibutyrate (PDBu) in the dialysis buffer had little effect on these calcium-binding properties. Analysis of PKC-calcium binding by gel filtration under equilibrium conditions gave similar results; only membrane-associated PKC bound significant amounts of calcium. Consequently, PKC is a member of what may be a large group of proteins that bind calcium in a phospholipid-dependent manner. The calcium concentrations needed to induce PKC-membrane binding were similar to those needed for calcium binding (about 40 microM calcium at the midpoint). However, the calcium concentration required for PKC-membrane binding was strongly influenced by the phosphatidylserine composition of the membranes. Membranes with higher percentages of phosphatidylserine required lower concentrations of calcium. These properties suggested that the calcium sites may be generated at the interface between PKC and the membrane. Calcium may function as a bridge between PKC and phospholipids. These studies also suggested that calcium-dependent PKC-membrane binding and PKC function could be regulated by a number of factors in addition to calcium levels and diacylglycerol content of the membrane.

  8. Chemical alteration by tooth bleaching of human salivary proteins that infiltrated subsurface enamel lesions: experimental study with bovine lesion model systems

    NARCIS (Netherlands)

    Iizuka, J.; Mukai, Y.; Taniguchi, M.; ten Cate, J.M.; Mikuni-Takagaki, Y.; Teranaka, T.


    Salivary macromolecules infiltrate white and brown spot enamel lesions and adsorb onto hydroxyapatite. Calcium-binding salivary proteins such as statherin hinder remineralization of these lesions. We assessed whether bleaching agents can remove salivary components that have infiltrated and bound to

  9. RapA2 Is a Calcium-binding Lectin Composed of Two Highly Conserved Cadherin-like Domains That Specifically Recognize Rhizobium leguminosarum Acidic Exopolysaccharides* (United States)

    Abdian, Patricia L.; Caramelo, Julio J.; Ausmees, Nora; Zorreguieta, Angeles


    In silico analyses have revealed a conserved protein domain (CHDL) widely present in bacteria that has significant structural similarity to eukaryotic cadherins. A CHDL domain was shown to be present in RapA, a protein that is involved in autoaggregation of Rhizobium cells, biofilm formation, and adhesion to plant roots as shown by us and others. Structural similarity to cadherins suggested calcium-dependent oligomerization of CHDL domains as a mechanistic basis for RapA action. Here we show by circular dichroism spectroscopy, light scattering, isothermal titration calorimetry, and other methods that RapA2 from Rhizobium leguminosarum indeed exhibits a cadherin-like β-sheet conformation and that its proper folding and stability are dependent on the binding of one calcium ion per protein molecule. By further in silico analysis we also reveal that RapA2 consists of two CHDL domains and expand the range of CHDL-containing proteins in bacteria and archaea. However, light scattering assays at various concentrations of added calcium revealed that RapA2 formed neither homo-oligomers nor hetero-oligomers with RapB (a distinct CHDL protein), indicating that RapA2 does not mediate cellular interactions through a cadherin-like mechanism. Instead, we demonstrate that RapA2 interacts specifically with the acidic exopolysaccharides (EPSs) produced by R. leguminosarum in a calcium-dependent manner, sustaining a role of these proteins in the development of the biofilm matrix made of EPS. Because EPS binding by RapA2 can only be attributed to its two CHDL domains, we propose that RapA2 is a calcium-dependent lectin and that CHDL domains in various bacterial and archaeal proteins confer carbohydrate binding activity to these proteins. PMID:23235153

  10. The SARS Coronavirus 3a protein binds calcium in its cytoplasmic domain. (United States)

    Minakshi, Rinki; Padhan, Kartika; Rehman, Safikur; Hassan, Md Imtaiyaz; Ahmad, Faizan


    The Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV) is a positive stranded RNA virus with ∼30kb genome. Among all open reading frames (orfs) of this virus, the orf3a is the largest, and encodes a protein of 274 amino acids, named as 3a protein. Sequence analysis suggests that the orf3a aligned to one calcium pump present in Plasmodium falciparum and the enzyme glutamine synthetase found in Leptospira interrogans. This sequence similarity was found to be limited only to amino acid residues 209-264 which form the cytoplasmic domain of the orf3a. Furthermore, this region was predicted to be involved in the calcium binding. Owing to this hypothesis, we were driven to establish its calcium binding property in vitro. Here, we expressed and purified the cytoplasmic domain of the 3a protein, called Cyto3a, as a recombinant His-tagged protein in the E. coli. The calcium binding nature was established by performing various staining methods such as ruthenium red and stains-all. (45)Ca overlay method was also done to further support the data. Since the 3a protein forms ion channels, we were interested to see any conformational changes occurring in the Cyot3a upon calcium binding, using fluorescence spectroscopy and circular dichroism. These studies clearly indicate a significant change in the conformation of the Cyto3a protein after binding with calcium. Our results strongly suggest that the cytoplasmic domain of the 3a protein of SARS-CoV binds calcium in vitro, causing a change in protein conformation. Copyright © 2014 Elsevier B.V. All rights reserved.

  11. Significance of calcium binding, tyrosine phosphorylation, and lysine trimethylation for the essential function of calmodulin in vertebrate cells analyzed in a novel gene replacement system

    DEFF Research Database (Denmark)

    Panina, Svetlana; Stephan, Alexander; la Cour, Jonas Marstrand


    system to study the effect ofCaMgene deletion. Here, we present a novel genetic system based on the chicken DT40 cell line, in which the two functional CaM genes were deleted and one allele replaced with a CaM transgene that can be artificially regulated.Weshow that CaM is essential for survival......Calmodulin (CaM) was shown to be essential for survival of lower eukaryotes by gene deletion experiments. So far, no CaM gene deletion was reported in higher eukaryotes. In vertebrates, CaM is expressed from several genes, which encode an identical protein, making it difficult to generate a model...

  12. GRP94: An HSP90-like protein specialized for protein folding and quality control in the endoplasmic reticulum

    DEFF Research Database (Denmark)

    Marzec, Michal; Eletto, Davide; Argon, Yair


    Glucose-regulated protein 94 is the HSP90-like protein in the lumen of the endoplasmic reticulum and therefore it chaperones secreted and membrane proteins. It has essential functions in development and physiology of multicellular organisms, at least in part because of this unique clientele. GRP94...... shares many biochemical features with other HSP90 proteins, in particular its domain structure and ATPase activity, but also displays distinct activities, such as calcium binding, necessitated by the conditions in the endoplasmic reticulum. GRP94's mode of action varies from the general HSP90 theme...

  13. Structures of apicomplexan calcium-dependent protein kinases reveal mechanism of activation by calcium

    Energy Technology Data Exchange (ETDEWEB)

    Wernimont, Amy K; Artz, Jennifer D.; Jr, Patrick Finerty; Lin, Yu-Hui; Amani, Mehrnaz; Allali-Hassani, Abdellah; Senisterra, Guillermo; Vedadi, Masoud; Tempel, Wolfram; Mackenzie, Farrell; Chau, Irene; Lourido, Sebastian; Sibley, L. David; Hui, Raymond (Toronto); (WU-MED)


    Calcium-dependent protein kinases (CDPKs) have pivotal roles in the calcium-signaling pathway in plants, ciliates and apicomplexan parasites and comprise a calmodulin-dependent kinase (CaMK)-like kinase domain regulated by a calcium-binding domain in the C terminus. To understand this intramolecular mechanism of activation, we solved the structures of the autoinhibited (apo) and activated (calcium-bound) conformations of CDPKs from the apicomplexan parasites Toxoplasma gondii and Cryptosporidium parvum. In the apo form, the C-terminal CDPK activation domain (CAD) resembles a calmodulin protein with an unexpected long helix in the N terminus that inhibits the kinase domain in the same manner as CaMKII. Calcium binding triggers the reorganization of the CAD into a highly intricate fold, leading to its relocation around the base of the kinase domain to a site remote from the substrate binding site. This large conformational change constitutes a distinct mechanism in calcium signal-transduction pathways.

  14. Nonphosphorylated neurofilament protein is expressed by scattered neurons in the vestibular and precerebellar brainstem


    Baizer, Joan S.


    Vestibular information is essential for the control of posture, balance, and eye movements. The vestibular nerve projects to the four nuclei of the vestibular nuclear complex (VNC), as well as to several additional brainstem nuclei and the cerebellum. We have found that expression of the calcium-binding proteins calretinin (CR) and calbindin (CB), and the synthetic enzyme for nitric oxide synthase (nNOS) define subdivisions of the medial vestibular nucleus (MVe) and the nucleus prepositus (Pr...

  15. Point mutation in calcium-binding domain of mouse polyomavirus VP1 protein does not prevent virus-like particle formation, but changes VP1 interactions with Saccharomyces cerevisiae cell structures

    Czech Academy of Sciences Publication Activity Database

    Adamec, T.; Palková, Zdena; Velková, K.; Štokrová, Jitka; Forstová, J.


    Roč. 5, 4-5 (2005), s. 331-340 ISSN 1567-1356 R&D Projects: GA ČR GA204/03/0593 Institutional research plan: CEZ:AV0Z5052915 Keywords : polyomavirus VP1 * Saccharomyces cerevisiae * heterologous expression Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.477, year: 2005

  16. Dentin Matrix Proteins in Bone Tissue Engineering. (United States)

    Ravindran, Sriram; George, Anne


    Dentin and bone are mineralized tissue matrices comprised of collagen fibrils and reinforced with oriented crystalline hydroxyapatite. Although both tissues perform different functionalities, they are assembled and orchestrated by mesenchymal cells that synthesize both collagenous and noncollagenous proteins albeit in different proportions. The dentin matrix proteins (DMPs) have been studied in great detail in recent years due to its inherent calcium binding properties in the extracellular matrix resulting in tissue calcification. Recent studies have shown that these proteins can serve both as intracellular signaling proteins leading to induction of stem cell differentiation and also function as nucleating proteins in the extracellular matrix. These properties make the DMPs attractive candidates for bone and dentin tissue regeneration. This chapter will provide an overview of the DMPs, their functionality and their proven and possible applications with respect to bone tissue engineering.

  17. S100 Proteins As an Important Regulator of Macrophage Inflammation

    Directory of Open Access Journals (Sweden)

    Chang Xia


    Full Text Available The S100 proteins, a family of calcium-binding cytosolic proteins, have a broad range of intracellular and extracellular functions through regulating calcium balance, cell apoptosis, migration, proliferation, differentiation, energy metabolism, and inflammation. The intracellular functions of S100 proteins involve interaction with intracellular receptors, membrane protein recruitment/transportation, transcriptional regulation and integrating with enzymes or nucleic acids, and DNA repair. The S100 proteins could also be released from the cytoplasm, induced by tissue/cell damage and cellular stress. The extracellular S100 proteins, serving as a danger signal, are crucial in regulating immune homeostasis, post-traumatic injury, and inflammation. Extracellular S100 proteins are also considered biomarkers for some specific diseases. In this review, we will discuss the multi-functional roles of S100 proteins, especially their potential roles associated with cell migration, differentiation, tissue repair, and inflammation.

  18. Enrichment of low-abundance brain proteins by preparative electrophoresis. (United States)

    Fountoulakis, Michael; Juranville, Jean François


    Detection of low-copy-number gene products is essential for the development of novel drugs, however, it represents a major drawback of proteomics and simultaneously a scientific challenge. We studied the enrichment of rat brain cytosolic proteins by preparative electrophoresis using the PrepCell apparatus. The electrophoresis was performed in the presence of 0.1% lithium dodecyl sulfate. The proteins eluted from the gel were analyzed by two-dimensional gel electrophoresis and identified by matrix-assisted laser desorption ionization mass specrometry. Lithium dodecyl sulfate was easily exchanged against agents compatible with isoelectric focusing. Low-abundance proteins, which had not been found before, including neuronal-specific and calcium-binding proteins, were detected. In particular, low-molecular-mass proteins, such as hippocalcin, visinin-like proteins, and 14-3-3 proteins were strongly enriched by preparative electrophoresis.

  19. Protein (Cyanobacteria): 440682381 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  20. Molecular cloning, occurrence, and expression of a novel partially secreted protein "psoriasin" that is highly up-regulated in psoriatic skin

    DEFF Research Database (Denmark)

    Madsen, Peder; Rasmussen, H H; Leffers, H


    the vaccinia virus expression system. Analysis of the predicted sequence revealed a potential calcium-binding sequence of the EF-hand type, as well as the absence of a signal sequence at its amino terminal. Psoriasin is not related to other proteins that migrate closely in 2D gels (MRP 14, also known...... as calgranulin B, L1 and calprotectin; MRP 8, or calgranulin A and cystatin A or stefin A), and bears no significant sequence homology with any other protein of known primary structure. Increased expression of psoriasin mRNA in psoriatic keratinocytes was confirmed by Northern blotting and in situ hybridization...

  1. Arabidopsis protein kinase PKS5 inhibits the plasma membrane H+ -ATPase by preventing interaction with 14-3-3 protein

    DEFF Research Database (Denmark)

    Fuglsang, Anja Thoe; Guo, Yan; Cuin, Tracey A.


    that an Arabidopsis thaliana Ser/Thr protein kinase, PKS5, is a negative regulator of the plasma membrane proton pump (PM Hþ-ATPase). Loss-of-function pks5 mutant plants are more tolerant of high external pH due to extrusion of protons to the extracellular space. PKS5 phosphorylates the PM Hþ-ATPase AHA2 at a novel...... site, Ser-931, in the C-terminal regulatory domain. Phosphorylation at this site inhibits interaction between the PM Hþ-ATPase and an activating 14-3-3 protein in a yeast expression system. We show that PKS5 interacts with the calcium binding protein SCaBP1 and that high external pH can trigger...... an increase in the concentration of cytosolic-free calcium. These results suggest that PKS5 is part of a calcium-signaling pathway mediating PM Hþ-ATPase regulation....

  2. The vitellogenin cDNA of Cherax quadricarinatus encodes a lipoprotein with calcium binding ability, and its expression is induced following the removal of the androgenic gland in a sexually plastic system. (United States)

    Abdu, Uri; Davis, Claytus; Khalaila, Isam; Sagi, Amir


    Oocyte maturation in decapod crustaceans is a two step process. Primary vitellogenesis is followed by a variable hiatus that lasts up to the onset of secondary vitellogenesis, which is marked by the rapid accumulation of yolk proteins in the oocytes. We have cloned a complete Cherax quadricarinatus vitellogenin cDNA. The sequenced cDNA contains a 2584 aa open reading frame which shows sequence similarity to vitellogenins from other crustaceans. The mRNA encodes at least two of the previously identified vitellin components, indicating that the primary translation product is subject to post-translational modification, including proteolytic cleavage. The region close to the 3(') end of the mRNA encodes a previously characterized negatively charged protein (provisionally designated P(106)). We show here that the negative charge of P(106) could be due to its ability to bind calcium. Northern blot data show that this gene is expressed as a single 8000 nt transcript and is present in the hepatopancreas of secondary-vitellogenic females. Primary vitellogenic and other tissues examined in male and female animals were negative. In sexually plastic intersex animals, removal of the androgenic gland results in vitellogenin transcription, indicating that the gene is negatively regulated by the androgenic gland.

  3. A study on the mechanism of protein adsorption to TiO2. (United States)

    Ellingsen, J E


    The hypothesis that calcium ions adsorb to TiO2 and further to macromolecules with high affinity for Ca2+ was tested. The reaction of human serum proteins with TiO2 was examined and compared with the reaction with hydroxyapatite. The oxide covered titanium surfaces reacted with calcium when exposed to CaCl2 and calcium was identified to a depth of 17 nm into the oxide layer. Surface adsorbed serum proteins were dissolved by EDTA and analysis by the SDS-PAGE technique revealed that the surfaces of TiO2 and hydroxyapatite appeared to take up the same proteins selectively from human serum. Albumin, prealbumin and IgG were identified by immunoelectrophoresis. It is suggested that calcium binding may be one mechanism by which proteins adsorb to TiO2. It is well established that this is the case with hydroxyapatite.

  4. The S100 proteins in epidermis: Topology and function. (United States)

    Leśniak, Wiesława; Graczyk-Jarzynka, Agnieszka


    S100 proteins are small calcium binding proteins encoded by genes located in the epidermal differentiation complex (EDC). Differently to other proteins encoded by EDC genes, which are indispensable for normal epidermal differentiation, the role of S100 proteins in the epidermis remains largely unknown. Particular S100 proteins differ in their distribution in epidermal layers, skin appendages, melanocytes and Langerhans cells. Taking into account that each epidermal component consists of specialized cells with well-defined functions, such differential distribution may be indicative of the function of a given S100 protein. We used this criterion together with the survey of the current experimental data pertinent to epidermis to provide a fairly comprehensive view on the possible function of individual S100 proteins in this tissue. S100 proteins are differently expressed and, despite extensive structural homology, perform diverse functions in the epidermis. Certain S100 proteins probably ensure constant epidermal renewal and support wound healing while others act in epidermal differentiation or have a protective role. As their expression is differently affected in various skin pathologies, particular S100 proteins could be valuable diagnostic markers. S100 proteins seem to be important although not yet fully recognized epidermal constituents. Better understanding of their role in the epidermis might be helpful in designing therapies to various skin diseases. Copyright © 2015 Elsevier B.V. All rights reserved.

  5. EDTA-insoluble, calcium-binding proteoglycan in bovine bone (United States)

    Hashimoto, Y.; Lester, G. E.; Caterson, B.; Yamauchi, M.


    A calcium ion precipitable, trypsin-generated proteoglycan fragment has been isolated from the demineralized, EDTA-insoluble matrices of bone. The demineralized matrix was completely digested with trypsin, increasing concentrations of CaCl2 were added to the supernatant, and the resulting precipitates were analyzed. The amount of precipitate gradually increased with higher concentrations of calcium and was reversibly solubilized by EDTA. After molecular sieve and anion exchange chromatography, a proteoglycan-containing peak was obtained. Immunochemical analysis showed that this peak contained chondroitin 4-sulfate and possibly keratan sulfate. Amino acid analysis showed that this proteoglycan contained high amounts of aspartic acid/asparagine (Asx), serine (Ser), glutamic acid/glutamine (Glx), proline (Pro), and glycine (Gly); however, it contained little leucine (Leu) which suggests that it is not a member of the leucine-rich small proteoglycan family. In addition, significant amounts of phosphoserine (P-Ser) and hydroxyproline (Hyp) were identified in hydrolysates of this fraction. A single band (M(r) 59 kDa) was obtained on SDS-PAGE that stained with Stains-all but not with Coomassie Brilliant Blue R-250. If bone powder was trypsinized prior to demineralization, this proteoglycan-containing fraction was not liberated. Collectively, these results indicate that a proteoglycan occurs in the demineralized matrix that is precipitated with CaCl2 and is closely associated with both mineral and collagen matrices. Such a molecule might facilitate the structural network for the induction of mineralization in bone.

  6. EDTA-insoluble, calcium-binding proteoglycan in bovine bone. (United States)

    Hashimoto, Y; Lester, G E; Caterson, B; Yamauchi, M


    A calcium ion precipitable, trypsin-generated proteoglycan fragment has been isolated from the demineralized, EDTA-insoluble matrices of bone. The demineralized matrix was completely digested with trypsin, increasing concentrations of CaCl2 were added to the supernatant, and the resulting precipitates were analyzed. The amount of precipitate gradually increased with higher concentrations of calcium and was reversibly solubilized by EDTA. After molecular sieve and anion exchange chromatography, a proteoglycan-containing peak was obtained. Immunochemical analysis showed that this peak contained chondroitin 4-sulfate and possibly keratan sulfate. Amino acid analysis showed that this proteoglycan contained high amounts of aspartic acid/asparagine (Asx), serine (Ser), glutamic acid/glutamine (Glx), proline (Pro), and glycine (Gly); however, it contained little leucine (Leu) which suggests that it is not a member of the leucine-rich small proteoglycan family. In addition, significant amounts of phosphoserine (P-Ser) and hydroxyproline (Hyp) were identified in hydrolysates of this fraction. A single band (M(r) 59 kDa) was obtained on SDS-PAGE that stained with Stains-all but not with Coomassie Brilliant Blue R-250. If bone powder was trypsinized prior to demineralization, this proteoglycan-containing fraction was not liberated. Collectively, these results indicate that a proteoglycan occurs in the demineralized matrix that is precipitated with CaCl2 and is closely associated with both mineral and collagen matrices. Such a molecule might facilitate the structural network for the induction of mineralization in bone.

  7. Analytical models of calcium binding in a calcium channel

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Jinn-Liang [Department of Applied Mathematics, National Hsinchu University of Education, Hsinchu 300, Taiwan (China); Eisenberg, Bob [Department of Molecular Biophysics and Physiology, Rush University, Chicago, Illinois 60612 (United States)


    The anomalous mole fraction effect of L-type calcium channels is analyzed using a Fermi like distribution with the experimental data of Almers and McCleskey [J. Physiol. 353, 585 (1984)] and the atomic resolution model of Lipkind and Fozzard [Biochemistry 40, 6786 (2001)] of the selectivity filter of the channel. Much of the analysis is algebraic, independent of differential equations. The Fermi distribution is derived from the configuration entropy of ions and water molecules with different sizes, different valences, and interstitial voids between particles. It allows us to calculate potentials and distances (between the binding ion and the oxygen ions of the glutamate side chains) directly from the experimental data using algebraic formulas. The spatial resolution of these results is comparable with those of molecular models, but of course the accuracy is no better than that implied by the experimental data. The glutamate side chains in our model are flexible enough to accommodate different types of binding ions in different bath conditions. The binding curves of Na{sup +} and Ca{sup 2+} for [CaCl{sub 2}] ranging from 10{sup −8} to 10{sup −2} M with a fixed 32 mM background [NaCl] are shown to agree with published Monte Carlo simulations. The Poisson-Fermi differential equation—that includes both steric and correlation effects—is then used to obtain the spatial profiles of energy, concentration, and dielectric coefficient from the solvent region to the filter. The energy profiles of ions are shown to depend sensitively on the steric energy that is not taken into account in the classical rate theory. We improve the rate theory by introducing a steric energy that lumps the effects of excluded volumes of all ions and water molecules and empty spaces between particles created by Lennard-Jones type and electrostatic forces. We show that the energy landscape varies significantly with bath concentrations. The energy landscape is not constant.

  8. Calcium binding and voltage gating in Cx46 hemichannels. (United States)

    Pinto, Bernardo I; Pupo, Amaury; García, Isaac E; Mena-Ulecia, Karel; Martínez, Agustín D; Latorre, Ramón; Gonzalez, Carlos


    The opening of connexin (Cx) hemichannels in the membrane is tightly regulated by calcium (Ca 2+ ) and membrane voltage. Electrophysiological and atomic force microscopy experiments indicate that Ca 2+ stabilizes the hemichannel closed state. However, structural data show that Ca 2+ binding induces an electrostatic seal preventing ion transport without significant structural rearrangements. In agreement with the closed-state stabilization hypothesis, we found that the apparent Ca 2+ sensitivity is increased as the voltage is made more negative. Moreover, the voltage and Ca 2+ dependence of the channel kinetics indicate that the voltage sensor movement and Ca 2+ binding are allosterically coupled. An allosteric kinetic model in which the Ca 2+ decreases the energy necessary to deactivate the voltage sensor reproduces the effects of Ca 2+ and voltage in Cx46 hemichannels. In agreement with the model and suggesting a conformational change that narrows the pore, Ca 2+ inhibits the water flux through Cx hemichannels. We conclude that Ca 2+ and voltage act allosterically to stabilize the closed conformation of Cx46 hemichannels.

  9. Calcium binding to the purple membrane : A molecular dynamics study

    NARCIS (Netherlands)

    Wassenaar, Tsjerk A.; Daura, Xavier; Padros, Esteve; Mark, Alan E.


    The purple membrane (PM) is a specialized membrane patch found in halophilic archaea, containing the photoreceptor bacteriorhodopsin (bR). It is long known that calcium ions bind to the PM, but their position and role remain elusive to date. Molecular dynamics simulations in conjunction with a

  10. Protective effect of parvalbumin on excitotoxic motor neuron death

    DEFF Research Database (Denmark)

    Van den Bosch, L.; Schwaller, B.; Vleminckx, V.


    Amyotrophic lateral sclerosis, ALS, AMPA receptor, calcium-binding proteins, calcium buffering, excitotoxity, kainic acid, motor neuron, parvalbumin......Amyotrophic lateral sclerosis, ALS, AMPA receptor, calcium-binding proteins, calcium buffering, excitotoxity, kainic acid, motor neuron, parvalbumin...

  11. The aging human cochlear nucleus: Changes in the glial fibrillary acidic protein, intracellular calcium regulatory proteins, GABA neurotransmitter and cholinergic receptor. (United States)

    Sharma, Saroj; Nag, Tapas C; Thakar, Alok; Bhardwaj, Daya N; Roy, Tara Sankar


    The human auditory system is highly susceptible to environmental and metabolic insults which further affect the biochemical and physiological milieu of the cells that may contribute to progressive, hearing loss with aging. The cochlear nucleus (CN) is populated by morphologically diverse types of neurons with discrete physiological and neurochemical properties. Between the dorsal and the ventral cochlear nucleus (DCN and VCN), the VCN is further sub-divided into the rostral (rVCN) and caudal (cVCN) sub-divisions. Although, information is available on the age related neurochemical changes in the mammalian CN similar reports on human CN is still sparse. The morphometry and semiquantitative analysis of intensity of expression of glial fibrillary acidic protein (GFAP), calcium binding proteins (calbindin, calretinin and parvalbumin), gamma amino butyric acid (GABA) and nicotinic acetyl choline receptor (nAchR) beta 2 immunostaining were carried out in all three sub-divisions of the human CN from birth to 90 years. There was increased GFAP immunoreactivity in decades 2 and 3 in comparison to decade 1 in the CN. But no change was observed in rVCN from decade 4 onwards, whereas intense staining was also observed in decades 5 and 6 in cVCN and DCN. All three calcium binding proteins were highly expressed in early to middle ages, whereas a significant reduction was found in later decades in the VCN. GABA and nAchR beta 2 expressions were unchanged throughout in all the decades. The middle age may represent a critical period of onset and progression of aging changes in the CN and these alterations may add to the deterioration of hearing responses in the old age. Copyright © 2013 Elsevier B.V. All rights reserved.

  12. Large-scale identification of human cerebrovascular proteins: Inter-tissue and intracerebral vascular protein diversity.

    Directory of Open Access Journals (Sweden)

    Soo Jung Lee

    Full Text Available The human cerebrovascular system is responsible for regulating demand-dependent perfusion and maintaining the blood-brain barrier (BBB. In addition, defects in the human cerebrovasculature lead to stroke, intracerebral hemorrhage, vascular malformations, and vascular cognitive impairment. The objective of this study was to discover new proteins of the human cerebrovascular system using expression data from the Human Protein Atlas, a large-scale project which allows public access to immunohistochemical analysis of human tissues. We screened 20,158 proteins in the HPA and identified 346 expression patterns correlating to blood vessels in human brain. Independent experiments showed that 51/52 of these distributions could be experimentally replicated across different brain samples. Some proteins (40% demonstrated endothelial cell (EC-enriched expression, while others were expressed primarily in vascular smooth muscle cells (VSMC; 18%; 39% of these proteins were expressed in both cell types. Most brain EC markers were tissue oligospecific; that is, they were expressed in endothelia in an average of 4.8 out of 9 organs examined. Although most markers expressed in endothelial cells of the brain were present in all cerebral capillaries, a significant number (21% were expressed only in a fraction of brain capillaries within each brain sample. Among proteins found in cerebral VSMC, virtually all were also expressed in peripheral VSMC and in non-vascular smooth muscle cells (SMC. Only one was potentially brain specific: VHL (Von Hippel-Lindau tumor suppressor. HRC (histidine rich calcium binding protein and VHL were restricted to VSMC and not found in non-vascular tissues such as uterus or gut. In conclusion, we define a set of brain vascular proteins that could be relevant to understanding the unique physiology and pathophysiology of the human cerebrovasculature. This set of proteins defines inter-organ molecular differences in the vasculature and

  13. Cloning from purified high endothelial venule cells of hevin, a close relative of the antiadhesive extracellular matrix protein SPARC. (United States)

    Girard, J P; Springer, T A


    High endothelial venules (HEV) in lymphoid tissues support high levels of lymphocyte extravasion from the blood. We purified high endothelial cells from human tonsils by immunomagnetic selection with MECA-79 MAb to construct an HEV cDNA library. Differential screening of this library using cDNA probes from HEV (plus) or flat-walled vessel (minus) endothelial cells allowed us to characterize a novel human cDNA expressed to high levels in HEV. The cDNA encodes a secreted acidic calcium-binding glycoprotein of 664 aa residues, designated hevin, exhibiting 62% identity with the antiadhesive extracellular matrix protein SPARC, over a region of 232 aa spanning more than four fifths of the SPARC coding sequence. The primary structure and sequence of hevin and similar to SPARC-like proteins from rat and quail, called SC1 or QR1. Hevin could contribute to the induction or maintenance of features of the HEV endothelium that facilitate lymphocyte migration.

  14. Arabidopsis CDS blastp result: AK243353 [KOME

    Lifescience Database Archive (English)

    Full Text Available AK243353 J100061B04 At1g05990.1 68414.m00627 calcium-binding protein, putative strong similarity to calcium...-binding protein [Lotus japonicus] GI:18413495; contains INTERPRO:IPR002048 calcium-binding EF-hand domain 3e-11 ...

  15. Arabidopsis CDS blastp result: AK243353 [KOME

    Lifescience Database Archive (English)

    Full Text Available AK243353 J100061B04 At4g03290.1 68417.m00449 calcium-binding protein, putative similar to calcium...-binding protein [Lotus japonicus] GI:18413495; contains INTERPRO:IPR002048 calcium-binding EF-hand domain 3e-11 ...

  16. Arabidopsis CDS blastp result: AK242346 [KOME

    Lifescience Database Archive (English)

    Full Text Available AK242346 J080012M07 At1g05990.1 68414.m00627 calcium-binding protein, putative strong similarity to calcium...-binding protein [Lotus japonicus] GI:18413495; contains INTERPRO:IPR002048 calcium-binding EF-hand domain 1e-28 ...

  17. A Protein Involved in the Assembly of an Extracellular Calcium Storage Matrix* (United States)

    Glazer, Lilah; Shechter, Assaf; Tom, Moshe; Yudkovski, Yana; Weil, Simy; Aflalo, Eliahu David; Pamuru, Ramachandra Reddy; Khalaila, Isam; Bentov, Shmuel; Berman, Amir; Sagi, Amir


    Gastroliths, the calcium storage organs of crustaceans, consist of chitin-protein-mineral complexes in which the mineral component is stabilized amorphous calcium carbonate. To date, only three proteins, GAP 65, gastrolith matrix protein (GAMP), and orchestin, have been identified in gastroliths. Here, we report a novel protein, GAP 10, isolated from the gastrolith of the crayfish Cherax quadricarinatus and specifically expressed in its gastrolith disc. The encoding gene was cloned by partial sequencing of the protein extracted from the gastrolith matrix. Based on an assembled microarray cDNA chip, GAP 10 transcripts were found to be highly (12-fold) up-regulated in premolt gastrolith disc and significantly down-regulated in the hypodermis at the same molt stage. The deduced protein sequence of GAP 10 lacks chitin-binding domains and does not show homology to known proteins in the GenBankTM data base. It does, however, have an amino acid composition that has similarity to proteins extracted from invertebrate and ascidian-calcified extracellular matrices. The GAP 10 sequence contains a predicted signal peptide and predicted phosphorylation sites. In addition, the protein is phosphorylated and exhibits calcium-binding ability. Repeated daily injections of GAP 10 double strand RNA to premolt C. quadricarinatus resulted in a prolonged premolt stage and in the development of gastroliths with irregularly rough surfaces. These findings suggest that GAP 10 may be involved in the assembly of the gastrolith chitin-protein-mineral complex, particularly in the deposition of amorphous calcium carbonate. PMID:20150428

  18. Protein expression analysis of inflammation-related colon carcinogenesis

    Directory of Open Access Journals (Sweden)

    Yasui Yumiko


    Full Text Available Background: Chronic inflammation is a risk factor for colorectal cancer (CRC development. The aim of this study was to determine the differences in protein expression between CRC and the surrounding nontumorous colonic tissues in the mice that received azoxymethane (AOM and dextran sodium sulfate (DSS using a proteomic analysis. Materials and Methods: Male ICR mice were given a single intraperitoneal injection of AOM (10 mg/kg body weight, followed by 2% (w/v DSS in their drinking water for seven days, starting one week after the AOM injection. Colonic adenocarcinoma developed after 20 weeks and a proteomics analysis based on two-dimensional gel electrophoresis and ultraflex TOF/TOF mass spectrometry was conducted in the cancerous and nontumorous tissue specimens. Results: The proteomic analysis revealed 21 differentially expressed proteins in the cancerous tissues in comparison to the nontumorous tissues. There were five markedly increased proteins (beta-tropomyosin, tropomyosin 1 alpha isoform b, S100 calcium binding protein A9, and an unknown protein and 16 markedly decreased proteins (Car1 proteins, selenium-binding protein 1, HMG-CoA synthase, thioredoxin 1, 1 Cys peroxiredoxin protein 2, Fcgbp protein, Cytochrome c oxidase, subunit Va, ETHE1 protein, and 7 unknown proteins. Conclusions: There were 21 differentially expressed proteins in the cancerous tissues of the mice that received AOM and DSS. Their functions include metabolism, the antioxidant system, oxidative stress, mucin production, and inflammation. These findings may provide new insights into the mechanisms of inflammation-related colon carcinogenesis and the establishment of novel therapies and preventative strategies to treat carcinogenesis in the inflamed colon.

  19. Novel interactions of domain III from the envelope glycoprotein of dengue 2 virus with human plasma proteins. (United States)

    Huerta, Vivian; Ramos, Yassel; Yero, Alexis; Pupo, Dianne; Martín, Dayron; Toledo, Patricia; Fleitas, Noralvis; Gallien, Sebastien; Martín, Alejandro M; Márquez, Gabriel J; Pérez-Riverol, Yasset; Sarría, Mónica; Guirola, Osmany; González, Luis J; Domon, Bruno; Chinea, Glay


    Blood cells and plasma are important media for the four serotypes of dengue virus (DENV1-4) spreading into an infected person. Thus, interactions with human plasma proteins are expected to be decisive in the course of the viral infection. Affinity purification followed by MS analysis (AP/MS) was used to isolate and identify plasma-derived proteins capable to interact with a recombinant protein comprising the domain III of the envelope protein of DENV2 (DIIIE2). The elution of the AP potently inhibits DENV2 infection. Twenty-nine proteins were identified using a label-free approach as specifically captured by DIIIE2. Of these, a direct interaction with C reactive protein, thrombin and Inter-alpha-inhibitor complexes was confirmed by ELISA. Results provide further evidence of a significant representation of proteins from complement and coagulation cascades on DENV2 interactome in human plasma and stand out the domain III of the viral envelope protein as participant on these interactions. A functional clustering analysis highlights the presence of three structural motifs among putative DIIIE2-binding proteins: hydroxylation and EGF-like calcium-binding- and Gla domains. Early cycles of dengue virus replication take place in human blood cells. Thus, the characterization of the interactome of dengue virus proteins in human plasma can lead to the identification of pivotal interactions for the infection that can eventually constitute the target for the development of methods to control dengue virus-caused disease. In this work we identified 29 proteins from human plasma that potentially interact with the envelope protein of dengue 2 virus either directly or through co-complex formation. C reactive protein, thrombin and Inter-alpha-inhibitor complexes were validated as interactors of the domain III of the envelope protein of dengue 2. Results highlight the presence of three structural motifs among putative DIIIE2-binding proteins: hydroxylation and EGF-like calcium-binding

  20. Calcium-dependant binding proteins associated with human placental syncytiotrophoblast microvillous cytoskeleton. (United States)

    Webb, P D; Mahadevan, L C


    Isolated human placental syncytiotrophoblast microvillous plasma membrane vesicles were extracted with Triton X-100 to yield a detergent-insoluble residue. The residue contained approx. 50% of the total membrane protein and was qualitatively different from untreated trophoblast on SDS-polyacrylamide gel electrophoresis, Western blots and dot-immunobinding assay. Three major proteins, with molecular weights of 68, 36 and 34 kDa, dissociated from this non-ionic detergent-insoluble submembranous cytoskeletal fraction in the presence of calcium chelators. They were immunologically related to human lymphocyte cytoskeletal calcium-binding proteins, and the 36 kDa component reacted with antisera to the phospholipase A2 inhibitor, lipocortin II. Anti-lipocortin I sera did not recognise the 34 kDa protein, but did react with a series of trophoblast cytoskeletal proteins in the 34-37 kDa region. Incubation of epidermal growth factor with isolated trophoblast membrane vesicles stimulated the phosphorylation of a 36 kDa protein on tyrosine residues. Immunoprecipitation studies further showed there was no phosphorylation of the 34 kDa protein, but the 68 kDa protein was a major phosphorylated component of isolated syncytiotrophoblast membranes. p68 was principally phosphorylated on serine with slight tyrosine phosphorylation which showed an apparent increase after epidermal growth factor treatment. These results indicate a family of calcium-dependant binding proteins, some of which are phosphorylated, associated with the submembranous cytoskeleton of syncytiotrophoblast microvilli.

  1. Selective 'unlabeling' of amino acids in fractionally 13C labeled proteins: An approach for stereospecific NMR assignments of CH3 groups in Val and Leu residues

    Energy Technology Data Exchange (ETDEWEB)

    Atreya, H.S.; Chary, K.V.R. [Tata Institute of Fundamental Research, Department of Chemical Sciences (India)


    A novel methodology for stereospecific NMR assignments of methyl (CH{sub 3}) groups of Val and Leu residues in fractionally {sup 13}C-labeled proteins is presented. The approach is based on selective 'unlabeling' of specific amino acids in proteins while fractionally {sup 13}C-labeling the rest. A 2D [{sup 13}C-{sup 1}H] HSQC spectrum recorded on such a sample is devoid of peaks belonging to the 'unlabeled' amino acid residues. Such spectral simplification aids in unambiguous stereospecific assignment of diastereotopic CH{sub 3} groups in Val and Leu residues in large proteins. This methodology has been demonstrated on a 15 kDa calcium binding protein from Entamoeba histolytica (Eh-CaBP)

  2. Investigation into the applicability of the centrifugal microfluidics platform for the development of protein-ligand binding assays incorporating enhanced green fluorescent protein as a fluorescent reporter. (United States)

    Puckett, Libby G; Dikici, Emre; Lai, Siyi; Madou, Marc; Bachas, Leonidas G; Daunert, Sylvia


    The incorporation of a protein-ligand binding assay into a centrifugal microfluidics platform is described. The platform itself is a disc-shaped polymer substrate, upon which a series of microfluidic channels and reservoirs have been machined. Centrifugal microfluidics platforms require no internal moving parts, and fluid propulsion is achieved solely through rotation of the disc. Fluid flow is controlled by passive valves, the opening of which is dependent on the angular frequency of the rotating platform, the channel dimensions, and the physical properties of the fluid. To evaluate the effectiveness of incorporating a protein-based assay onto the centrifugal microfluidics analytical platform, a class-selective, homogeneous assay for the detection of phenothiazine antidepressants was employed. This class of drugs is known to bind to calmodulin, a calcium binding protein. Specifically, a fusion protein between calmodulin and enhanced green fluorescent protein was utilized. Calmodulin undergoes a conformational change upon binding to phenothiazines that alters the fluorescence properties of the attached fluorescent protein, which can be correlated to the concentration of the drug present. Another important aspect of this work was to study the efficacy of the platform to perform reconstitution assays. To do this, the biological reagent was dried on the platform and rehydrated to carry out the assay. The ability to prealiquot reagents on the platform should enhance its versatility and portability. The integration of protein-based assays in this platform should be useful in the design of analytical systems for high-throughput screening of pharmaceuticals and clinical diagnostics.

  3. Biomineralization of bone: a fresh view of the roles of non-collagenous proteins (United States)

    Gorski, Jeffrey Paul


    The impact of genetics has dramatically affected our understanding of the functions of non-collagenous proteins. Specifically, mutations and knockouts have defined their cellular spectrum of actions. However, the biochemical mechanisms mediated by non-collagenous proteins in biomineralization remain elusive. It is likely that this understanding will require more focused functional testing at the protein, cell, and tissue level. Although initially viewed as rather redundant and static acidic calcium binding proteins, it is now clear that non-collagenous proteins in mineralizing tissues represent diverse entities capable of forming multiple protein-protein nteractions which act in positive and negative ways to regulate the process of bone mineralization. Several new examples from the author’s laboratory are provided which illustrate this theme including an apparent activating effect of hydroxyapatite crystals on metalloproteinases. This review emphasizes the view that secreted non-collagenous proteins in mineralizing bone actively participate in the mineralization process and ultimately control where and how much mineral crystal is deposited, as well as determining the quality and biomechanical properties of the mineralized matrix produced. PMID:21622198

  4. Proteomic analysis on the alteration of protein expression in the placental villous tissue of early pregnancy loss. (United States)

    Liu, Ai-Xia; Jin, Fan; Zhang, Wu-Wen; Zhou, Tian-Hua; Zhou, Cai-Yun; Yao, Wei-Miao; Qian, Yu-Li; Huang, He-Feng


    Early pregnancy loss is the most common complication of human reproduction. Given the complexities of early development, it is likely that many mechanisms are involved. Knowledge of differences in protein expression in parallel profiling is essential to understand the comprehensive pathophysiological mechanism underlying early pregnancy loss. To identify proteins with different expression profiles related to early pregnancy loss, we applied a proteomic approach and performed two-dimensional gel electrophoresis (2-DE) on six placental villous tissues from patients with early pregnancy loss and six from normal pregnant women, followed by comparison of the silver-stained 2-DE profiles. It was found that 13 proteins were downregulated and 5 proteins were upregulated significantly (P ionization time-of-flight mass spectrometry. Anomalies of these proteins, including three principal antioxidant enzymes (copper/zinc-superoxide dismutase, peroxiredoxin 3, and thioredoxin-like 1 protein), S100 calcium binding protein, galectin-1, chorionic somatomammotropin hormone 1, transthyretin, fas inhibitory molecule, eukaryotic translation elongation factor, RNA-binding protein, ubiquitin-conjugating enzyme E2N, and proteasome beta-subunit, indicate widespread failure in cell regulations and processes such as antioxidative defense, differentiation, cell proliferation, metabolism, apoptosis, transcription, and proteolysis in early pregnancy loss. This study has identified several proteins that are associated with placentation and early development, shedding a new insight into the proteins that may be potentially involved in the pathophysiological mechanisms underlying early pregnancy loss.

  5. Alterations of Ca2+ responsive proteins within cholinergic neurons in aging and AD (United States)

    Riascos, David; Nicholas, Alexander; Samaeekia, Ravand; Yukhnanov, Rustam; Mesulam, M.-Marsel; Bigio, Eileen H.; Weintraub, Sandra; Guo, Ling; Geula, Changiz


    The molecular basis of selective neuronal vulnerability in Alzheimer ’s disease (AD) remains poorly understood. Using basal forebrain cholinergic neurons (BFCN) as a model and immunohistochemistry, we have demonstrated significant age-related loss of the calcium binding protein calbindin-D28K (CB) from BFCN, which was associated with tangle formation and degeneration in AD. Here we determined alterations in RNA and protein for CB and other Ca2+ responsive proteins Ca2+/calmodulin-dependent protein kinase I (CaMKI), growth-associated protein-43 (GAP43), calpain in the basal forebrain. We observed progressive downregulation of CB and CaMKI RNA in laser-captured BFCN in the normal-aged-AD continuum. We also detected progressive loss of CB, CaMKI-Delta, and GAP43 proteins in BF homogenates in aging and AD. Activated μ-calpain, a calcium-sensitive protease that degrades CaMKI and GAP-43, was significantly increased in the normal aged BF and was 10-times higher in AD BF. Overactivation of μ-calpain was confirmed using proteolytic fragments of its substrate spectrin. Substantial age and AD related alterations in Ca2+-sensing proteins most likely contribute to selective vulnerability of BFCN to degeneration in AD. PMID:24461366

  6. The effects of thermal treatments on protein profiles of Macrobrachium rosenbergii (giant river prawn) (United States)

    Sockalingam, Komathi; Misnan, Rosmilah; Yadzir, Zailatul Hani Mohd


    Prawn allergy is certainly the most frequent cause of allergic reactions in countries where this crustacean is a popular dish of seafood. The aim of this study was to determine the protein profiles of giant river prawn which scientifically known as Macrobrachium rosenbergii. Raw and cooked extracts (boiled, steamed and fried) of prawn samples were prepared and then resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). 27 protein bands between 6 to 207 kDa were detected in the SDS-PAGE gel of raw extracts while boiled, steamed and fried extracts revealed fewer protein bands. Steamed and boiled prawns presented higher numbers of protein bands compared to fried prawn. A prominent heat-resistant band between 32 to 38 kDa was seen in all extracts, might hypothesized to be tropomyosin. Other prominent bands between 17 to 20 kDa were also seen in all treated prawn extracts while bands of 24 to 27 kDa were seen in steamed and boiled prawn extracts. These positions are consistent with the known shellfish allergens myosin light chain, sacroplasmic calcium binding protein and troponin C respectively. Several other heat-sensitive protein bands at various molecular weights were also not detected in boiled, steamed and fried extracts of this prawn. This study showed that M. rosenbergii contains numerous heat-sensitive and heat-resistant proteins, which may play an important role in prawn allergy.

  7. A two-dimensional protein fragmentation-proteomic study of neuronal ceroid lipofuscinoses: identification and characterization of differentially expressed proteins. (United States)

    Wang, Peirong; Ju, Weina; Wu, Dan; Wang, Li; Yan, Ming; Zou, Junhua; He, Bing; Jenkins, Edmund C; Brown, W Ted; Zhong, Nanbert


    The neuronal ceroid lipofuscinoses (NCLs) are a group of neuronal degenerative diseases that primarily affect children. Previously we hypothesized that the similarity of the phenotypes among the variant subtypes of NCL suggests that the NCLs share a common metabolic functional pathway. To test our hypothesis, we have studied several candidate proteins identified using a proteomic approach. We analyzed their differential expression and cataloged their functions and involved pathways. Forty protein peaks, differentially expressed in NCLs, were selected from two-dimensional protein fragmentation (PF2D) maps and twenty-four proteins were identified by MALDI-TOF-MS or LC-ESI-MS/MS. Six proteins were verified by further Western blotting. Our results showed that annexin A1, annexin A2, and vimentin were significantly down-regulated in NCL1, NCL2, NCL3, and NCL8 cells; galectin-1 was down-regulated in NCL1, NCL3, and NCL8 but up-regulated in NCL2 cells; and isoform 5 of caldesmon was up-regulated in all NCL cell types. The histone 2B was down-regulated in NCL3. Functional analysis showed that the differentially expressed proteins identified by PF2D could be grouped into categories of intermediate filaments, cell motility, apoptosis, cytoskeleton, membrane trafficking, calcium binding, nucleosome assembly, pigment granule and cell development. Immunocytochemistry revealed nuclear translocalization of annexin A1 in CLN2-deficient fibroblasts and abnormal distribution of L-caldesmon in cultured CLN1, CLN2, CLN3 and CLN8-deficient fibroblasts. Finding differentially expressed proteins in variant NCLs, which showed disturbances of cytoskeleton, RAGE-dependent cellular pathways and decreased glycolysis provides evidence supporting our hypothesis. These findings may contribute to the discovery of molecular biomarkers and may help further elucidate the pathogenic mechanisms underlying the NCLs. Copyright © 2010 Elsevier B.V. All rights reserved.

  8. Evolution of EF-hand calcium-modulated proteins. II. Domains of several subfamilies have diverse evolutionary histories (United States)

    Nakayama, S.; Moncrief, N. D.; Kretsinger, R. H.


    In the first report in this series we described the relationships and evolution of 152 individual proteins of the EF-hand subfamilies. Here we add 66 additional proteins and define eight (CDC, TPNV, CLNB, LPS, DGK, 1F8, VIS, TCBP) new subfamilies and seven (CAL, SQUD, CDPK, EFH5, TPP, LAV, CRGP) new unique proteins, which we assume represent new subfamilies. The main focus of this study is the classification of individual EF-hand domains. Five subfamilies--calmodulin, troponin C, essential light chain, regulatory light chain, CDC31/caltractin--and three uniques--call, squidulin, and calcium-dependent protein kinase--are congruent in that all evolved from a common four-domain precursor. In contrast calpain and sarcoplasmic calcium-binding protein (SARC) each evolved from its own one-domain precursor. The remaining 19 subfamilies and uniques appear to have evolved by translocation and splicing of genes encoding the EF-hand domains that were precursors to the congruent eight and to calpain and to SARC. The rates of evolution of the EF-hand domains are slower following formation of the subfamilies and establishment of their functions. Subfamilies are not readily classified by patterns of calcium coordination, interdomain linker stability, and glycine and proline distribution. There are many homoplasies indicating that similar variants of the EF-hand evolved by independent pathways.

  9. Target size of calcium pump protein from skeletal muscle sarcoplasmic reticulum. (United States)

    Hymel, L; Maurer, A; Berenski, C; Jung, C Y; Fleischer, S


    The oligomeric size of calcium pump protein (CPP) in fast skeletal muscle sarcoplasmic reticulum membrane was determined using target theory analysis of radiation inactivation data. There was a parallel decrease of Ca2+-ATPase and calcium pumping activities with increasing radiation dose. The loss of staining intensity of the CPP band, observed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, also correlated directly with the loss of activity. The target size molecular weight of the CPP in the normal sarcoplasmic reticulum membrane ranged between 210,000 and 250,000, which is consistent with a dimeric structure. Essentially the same size is obtained for the non-phosphorylated CPP or for the phosphoenzyme form generated from either ATP (E1 state) or inorganic phosphate (E2 state). Hence, the oligomeric state of the pump does not appear to change during the catalytic cycle. Similar results were obtained with reconstituted sarcoplasmic reticulum membrane vesicles with different lipid to protein ratios. We conclude that the CPP is a dimer in both native and reconstituted sarcoplasmic reticulum membranes. The target size of the calcium-binding protein (calsequestrin) was found to be 50,000 daltons, approximating a monomer.

  10. Quantitative analysis of differentially expressed saliva proteins in human immunodeficiency virus type 1 (HIV-1) infected individuals

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Nawei; Zhang, Zhenyu [Beijing Chaoyang Hospital Affiliated Capital Medical University, Beijing (China); Feng, Shan [MOE Key Laboratory of Bioinformatics, School of Life Sciences, Tsinghua University, Beijing (China); Wang, Qingtao [Beijing Chaoyang Hospital Affiliated Capital Medical University, Beijing (China); Malamud, Daniel [NYU College of Dentistry, 345 East 24th Street, New York, NY 10010 (United States); Deng, Haiteng, E-mail: [MOE Key Laboratory of Bioinformatics, School of Life Sciences, Tsinghua University, Beijing (China)


    Highlights: ► A high-throughput method for profiling and quantification of the differentially expressed proteins in saliva samples was developed. ► Identified that DMBT1, S100A7, S100A8, S100A9 and alpha defensin were up-regulated in saliva from HIV-1 seropositive patients. ► Established analytical strategies are translatable to the clinical setting. -- Abstract: In the present study, we have established a new methodology to analyze saliva proteins from HIV-1-seropositive patients before highly active antiretroviral therapy (HAART) and seronegative controls. A total of 593 and 601 proteins were identified in the pooled saliva samples from 5 HIV-1 subjects and 5 controls, respectively. Forty-one proteins were found to be differentially expressed. Bioinformatic analysis of differentially expressed salivary proteins showed an increase of antimicrobial proteins and decrease of protease inhibitors upon HIV-1 infection. To validate some of these differentially expressed proteins, a high-throughput quantitation method was established to determine concentrations of 10 salivary proteins in 40 individual saliva samples from 20 seropositive patients before HAART and 20 seronegative subjects. This method was based on limited protein separation within the zone of the stacking gel of the 1D SDS PAGE and using isotope-coded synthetic peptides as internal standards. The results demonstrated that a combination of protein profiling and targeted quantitation is an efficient method to identify and validate differentially expressed salivary proteins. Expression levels of members of the calcium-binding S100 protein family and deleted in malignant brain tumors 1 protein (DMBT1) were up-regulated while that of Mucin 5B was down-regulated in HIV-1 seropositive saliva samples, which may provide new perspectives for monitoring HIV-infection and understanding the mechanism of HIV-1 infectivity.

  11. Solubilization and identification of hen eggshell membrane proteins during different times of chicken embryo development using the proteomic approach. (United States)

    Kaweewong, Kritsda; Garnjanagoonchorn, Wunwiboon; Jirapakkul, Wannee; Roytrakul, Sittiruk


    A fertilized chicken egg is a unit of life. During hatching, transport of nutrients, including calcium, have been reported from the egg components to the developing embryo. Calcium is mobilized from the eggshell with the involvement of Ca(2+)-binding proteins. In addition, other unknown proteins may also play some important roles during embryo developing process. Therefore identification and prediction of biological functions of eggshell membrane (ESM) proteins during chick embryo development was conducted by proteome analysis. Comparison of different lysis solutions indicated that the highest ability to extract ESM proteins could be obtained with 1 % sodium dodecyl sulfate in 5 mM Tris-HCl buffer pH 8.8 containing 0.1 % 2-mercaptoethanol. In this study fertilized Cornish chicken eggs were incubated at 37 °C in humidified incubators for up to 21 days. At selected times (days 1, 9, 15 and 21), samples were taken and the ESMs were carefully separated by hand, washed with distilled water, and air-dried at room temperature. The ESM proteins were then solubilized and analyzed by proteome analysis. Sodium dodecyl sulfate polyacrylamide gel electrophoresis combined with high performance liquid chromatography and mass spectrometry revealed 62 proteins in the ESM; only keratin is known ESM protein, 8 of which are egg white proteins and related while 53 others have not previously been reported. Some differences in the types of proteins and their molecular functions were noted in ESM at different incubation times. One protein which was present only at days 15 and 21 of egg incubation was identified as a calcium binding protein i.e. EGF like repeats and discoidin I like domain 3 (EDIL3 homologous protein).

  12. Activation-resistant homozygous protein C R229W mutation causing familial perinatal intracranial hemorrhage and delayed onset of thrombosis. (United States)

    Alsultan, Abdulrahman; Gale, Andrew J; Kurban, Kadijah; Khalifah, Mohammed; Albadr, Fahad B; Griffin, John H


    We describe a family with two first-degree cousins who presented with similar phenotypes characterized by neonatal intracranial hemorrhage and subsequent onset of thrombosis. We enrolled the two affected patients, five unaffected family members and fifty-five normal controls. Clinical, laboratory, and radiological characteristics of patients were obtained. Exome sequencing was performed for the older affected child. PROC c.811 C>T was genotyped by PCR in patients, family members, and controls. Protein C amidolytic activity and antigen were measured using the STACHROM® protein C kit and ELISAs. To define functional abnormalities caused by the patients' mutation, recombinant wildtype protein C and its mutants R229W, R229Q and R229A were studied. For the two cousins, protein C amidolytic activity was 61% and 59% and antigen was 57% and 73% (nl 70-140%), respectively. Exome sequencing revealed a homozygous variant in exon 9 of the protein C (PROC) gene c.811 C>T (R229W). The R229W mutation is located in the calcium binding loop of protein C's protease domain that mediates thrombomodulin interactions. Recombinant R229W-protein C mutant was strikingly defective in rate of activation by thrombin: thrombomodulin, suggesting an in vivo deficit in these children for generation of activated protein C. These cases emphasize that protein C and activated protein C are important in maintaining the integrity of the brain vascular endothelium in humans. Moreover, routine protein C assays utilizing snake venom protease fail to detect protein C mutants that are resistant to thrombin:thrombomodulin activation. Copyright © 2016 Elsevier Ltd. All rights reserved.

  13. Respective contribution of CML8 and CML9, two arabidopsis calmodulin-like proteins, to plant stress responses. (United States)

    Zhu, Xiaoyang; Perez, Manon; Aldon, Didier; Galaud, Jean-Philippe


    In their natural environment, plants have to continuously face constraints such as biotic and abiotic stresses. To achieve their life cycle, plants have to perceive and interpret the nature, but also the strength of environmental stimuli to activate appropriate physiological responses. Nowadays, it is well established that signaling pathways are crucial steps in the implementation of rapid and efficient plant responses such as genetic reprogramming. It is also reported that rapid raises in calcium (Ca2+) levels within plant cells participate in these early signaling steps and are essential to coordinate adaptive responses. However, to be informative, calcium increases need to be decoded and relayed by calcium-binding proteins also referred as calcium sensors to carry-out the appropriate responses. In a recent study, we showed that CML8, an Arabidopsis calcium sensor belonging to the calmodulin-like (CML) protein family, promotes plant immunity against the phytopathogenic bacteria Pseudomonas syringae pv tomato (strain DC3000). Interestingly, other CML proteins such as CML9 were also reported to contribute to plant immunity using the same pathosystem. In this addendum, we propose to discuss about the specific contribution of these 2 CMLs in stress responses.

  14. Orf virus 002 protein targets ovine protein S100A4 and inhibits NF-kappa B signaling

    Directory of Open Access Journals (Sweden)

    Daxiang Chen


    Full Text Available Orf virus (ORFV, a member of Parapoxvirus, has evolved various strategies to modulate the immune responses of host cells. The ORFV-encoded protein ORFV002, a regulator factor, has been found to inhibit the acetylation of NF-κB-p65 by blocking phosphorylation of NF-kB-p65 at Ser276 and also to disrupt the binding of NF-kB-p65 and p300. To explore the mechanism by which ORFV002 regulates NF-κB signaling, the understanding of ORFV002 potential binding partners in host cells is critical. In this study, ovine S100 calcium binding protein A4 (S100A4, prolylendopeptidase-like (PREPL and NADH dehydrogenase (ubiquinone 1 alpha subcomplex 8 (NDUFA8 were found to interact with ORFV002 based on the yeast two-hybrid (Y2H assay using a cDNA library derived from primary ovine fetal turbinate cells (OFTu. GST pull-down and bidirectional co-immunoprecipitation assay results demonstrate that ORFV002 interacts with S100A4 directly. Following the pEGFP-ORFV002 (p002GFP transfection, we found that cytoplasmic S100A4 translocates into the nucleus and co-localizes with ORFV002. Furthermore, the inhibitory effect of ORFV002 on NF-κB signaling was significantly restored by S100A4 knock-down phenotype, suggesting ovine S100A4 participating in the ORFV002-mediated NF-κB signaling. These data demonstrate that ORFV002 inhibits the NF-κB activation through its interaction with S100A4 along with its nucleus translocation.

  15. Protein Foods (United States)

    ... Text Size: A A A Listen En Español Protein Foods Foods high in protein such as fish, ... for the vegetarian proteins, whether they have carbohydrate. Protein Choices Plant-Based Proteins Plant-based protein foods ...

  16. Calcium binding kinetics of troponin C strongly modulate cooperative activation and tension kinetics in cardiac muscle. (United States)

    Kreutziger, Kareen L; Piroddi, Nicoletta; McMichael, Jonathan T; Tesi, Chiara; Poggesi, Corrado; Regnier, Michael


    Tension development and relaxation in cardiac muscle are regulated at the thin filament via Ca(2+) binding to cardiac troponin C (cTnC) and strong cross-bridge binding. However, the influence of cTnC Ca(2+)-binding properties on these processes in the organized structure of cardiac sarcomeres is not well-understood and likely differs from skeletal muscle. To study this we generated single amino acid variants of cTnC with altered Ca(2+) dissociation rates (k(off)), as measured in whole troponin (cTn) complex by stopped-flow spectroscopy (I61Q cTn>WT cTn>L48Q cTn), and exchanged them into cardiac myofibrils and demembranated trabeculae. In myofibrils at saturating Ca(2+), L48Q cTnC did not affect maximum tension (T(max)), thin filament activation (k(ACT)) and tension development (k(TR)) rates, or the rates of relaxation, but increased duration of slow phase relaxation. In contrast, I61Q cTnC reduced T(max), k(ACT) and k(TR) by 40-65% with little change in relaxation. Interestingly, k(ACT) was less than k(TR) with I61Q cTnC, and this difference increased with addition of inorganic phosphate, suggesting that reduced cTnC Ca(2+)-affinity can limit thin filament activation kinetics. Trabeculae exchanged with I61Q cTn had reduced T(max), Ca(2+) sensitivity of tension (pCa(50)), and slope (n(H)) of tension-pCa, while L48Q cTn increased pCa(50) and reduced n(H). Increased cross-bridge cycling with 2-deoxy-ATP increased pCa(50) with WT or L48Q cTn, but not I61Q cTn. We discuss the implications of these results for understanding the role of cTn Ca(2+)-binding properties on the magnitude and rate of tension development and relaxation in cardiac muscle. Copyright © 2010 Elsevier Ltd. All rights reserved.

  17. Protein profiling of paraquat-exposed rat lungs following treatment with Acai (Euterpe oleracea Mart.) berry extract. (United States)

    Kim, Yong-Sik; Jung, Hana; Zerin, Tamanna; Song, Ho-Yeon


    Paraquat (1,1'-dimethyl-4,4'-bipyridinium chloride, PQ) is a non-selective herbicide, and PQ poisoning by accidental or intentional ingestion is a cause of numerous fatalities around the world every year. Although a great deal of research has been conducted into the development of an acceptable treatment for PQ poisoning, no effective guidelines for patients have been developed thus far. Acai berry extract and juice have been highlighted in this regard, due to their observed antioxidant effects in various diseases. Furthermore, the acai berry has been used in dietary supplements, as it contains a variety of nutrients, including proteins, lipids, vitamins A, C and E and polyphenols. In this study, we conducted proteomic analysis of PQ-poisoned rat lungs to evaluate the changes in protein expression induced by PQ and to identify any protective effects of acai berry on the PQ poisoning. Our data revealed that the expression of the calcium signaling-related proteins calcium binding protein 1 (CaBP1), FK506 binding protein 4 (FKBP4), S100A6 and secreted protein acidic and rich in cysteine (Sparc, also known as osteonectin) were induced by PQ treatment and downregulated by acai berry treatment. However, the levels of protein kinase C substrate 80K-H were shown to be downregulated as the result of PQ treatment. Our results indicated that these proteins may function as biomarkers for acute poisoning by PQ exposure. Further studies may be necessary to understand their clinical relevance with regard to PQ poisoning.

  18. RNAi silenced Dd-grp94 (Dictyostelium discoideum glucose-regulated protein 94 kDa) cell lines in Dictyostelium exhibit marked reduction in growth rate and delay in development. (United States)

    Baviskar, Sandhya N; Shields, Malcolm S


    Glucose-regulated 94 kDa protein (Grp94) is a resident of the endoplasmic reticulum (ER) of multicellular eukaryotes. It is a constitutively expressed protein that is overexpressed in certain abnormal conditions of the cell such as depletion of glucose and calcium, and low oxygen and pH. The protein is also implicated in diseased conditions like cancer and Alzheimer's disease. In this study, the consequences of downregulation of Grp94 were investigated at both unicellular and multicellular stages of Dictyostelium discoideum. Previous studies have shown the expression of Dd-Grp94 (Dictyostelium discoideum glucose-regulated 94 kDa protein) in wild-type cells varies during development, and overexpression of Dd-Grp94 leads to abnormal cell shape and inhibition of development (i.e., formation of fruiting bodies). Grp94 is a known calcium binding protein and an efficient calcium buffer. Therefore, in the present study we hypothesized that downregulation of Dd-Grp94 protein would affect Dictyostelium cell structure, growth, and development. We found that Dd-grp94 RNAi recombinants exhibited reduced growth rate, cell size, and a subtle change in cell motility compared to the parental cells. The recombinants also exhibited a delay in development and small fruiting bodies. These results establish that Dd-grp94 plays a crucial role in determining normal cell structure, growth and differentiation.

  19. Identification of S-Nitrosylated (SNO) Proteins inEntamoeba histolyticaAdapted to Nitrosative Stress: Insights into the Role of SNO Actin andIn vitroVirulence. (United States)

    Trebicz-Geffen, Meirav; Shahi, Preeti; Nagaraja, Shruti; Vanunu, Shai; Manor, Shiran; Avrahami, Amit; Ankri, Serge


    We have recently reported that Entamoeba histolytica trophozoites can adapt to toxic levels of the nitric oxide (NO) donor, S-nitrosoglutathione (GSNO). Even if the consequences of this adaptation on the modulation of gene expression in NO-adapted trophozoites (NAT) have been previously explored, insight on S-nitrosylated (SNO) proteins in NAT is missing. Our study aims to fill this knowledge gap by performing a screening of SNO proteins in NAT. Employing SNO resin-assisted capture (RAC), we identified 242 putative SNO proteins with key functions in calcium binding, enzyme modulation, redox homeostasis, and actin cytoskeleton. Of the SNO proteins in NAT, proteins that are associated with actin family cytoskeleton protein are significantly enriched. Here we report that the formation of actin filaments (F-actin) is impaired in NAT. Consequently, the ability of NAT to ingest erythrocytes and their motility and their cytopathic activity are impaired. These phenotypes can be imitated by treating control parasite with cytochalasin D (CytD), a drug that binds to F-actin polymer and prevent polymerization of actin monomers. Removal of GSNO from the culture medium of NAT restored the sensitivity of the parasite to nitrosative stress (NS) and its ability to form F-actin formation and its virulence. These results establish the central role of NO in shaping the virulence of the parasite through its effect on F-actin formation and highlight the impressive ability of this parasite to adapt to NS.

  20. Olopatadine Suppresses the Migration of THP-1 Monocytes Induced by S100A12 Protein

    Directory of Open Access Journals (Sweden)


    Full Text Available Olopatadine hydrochloride (olopatadine is an antiallergic drug with histamine H 1 receptor antagonistic activity. Recently, olopatadine has been shown to bind to S100A12 which is a member of the S100 family of calcium-binding proteins, and exerts multiple proinflammatory activities including chemotaxis for monocytes and neutrophils. In this study, we examined the possibility that the interaction of olopatadine with S100A12 inhibits the proinflammatory effects of S100A12. Pretreatment of olopatadine with S100A12 reduced migration of THP-1, a monocyte cell line, induced by S100A12 alone, but did not affect recombinant human regulated upon activation, normal T cell expressed and secreted (RANTES-induced migration. Amlexanox, which also binds to S100A12, inhibited the THP-1 migration induced by S100A12. However, ketotifen, another histamine H 1 receptor antagonist, had little effect on the activity of S100A12. These results suggest that olopatadine has a new mechanism of action, that is, suppression of the function of S100A12, in addition to histamine H 1 receptor antagonistic activity.

  1. Aldosterone producing adrenal adenomas are characterized by activation of calcium/calmodulin-dependent protein kinase (CaMK) dependent pathways. (United States)

    Sackmann, S; Lichtenauer, U; Shapiro, I; Reincke, M; Beuschlein, F


    Primary aldosteronism is the most prevalent cause of secondary hypertension. However, insights in pathophysiological mechanisms resulting in autonomous aldosterone secretion are limited. Although transcriptional regulators of aldosterone synthase (CYP11B2) including calcium-binding calmodulin kinase (CaMK) dependent pathways have been defined in vitro, it remains uncertain whether these mechanisms play a role in the context of dysregulated steroidogenesis in aldosterone producing adrenadenomas. Thus, we compared expression and activation of key components of CaMK pathways in aldosterone producing adenomas (APAs) with normal adrenals glands (NAGs). As expected, aldosterone synthase expression in APAs was significantly higher in comparison to NAGs, suggesting transcriptional activation as a contributing factor of aldosterone excess. Along the same line, CaMKI was significantly upregulated in APAs on the mRNA and protein level. Furthermore, immunohistochemistry revealed nuclear localization of CaMKI in these tumors. The phosphorylation of CREB, a target protein for CaMKI was increased, which could represent a further stimulation of aldosterone synthase transcription. In summary, this study provides indirect evidence for a causative involvement of the CaM kinase signaling pathway in human aldosterone producing adenomas. © Georg Thieme Verlag KG Stuttgart · New York.

  2. Ethylene-mediated cross-talk between calcium-dependent protein kinase and MAPK signaling controls stress responses in plants. (United States)

    Ludwig, Andrea A; Saitoh, Hiromasa; Felix, Georg; Freymark, Gerald; Miersch, Otto; Wasternack, Claus; Boller, Thomas; Jones, Jonathan D G; Romeis, Tina


    Plants are constantly exposed to environmental changes and need to integrate multiple external stress cues. Calcium-dependent protein kinases (CDPKs) are implicated as major primary Ca2+ sensors in plants. CDPK activation, like activation of mitogen-activated protein kinases (MAPKs), is triggered by biotic and abiotic stresses, although distinct stimulus-specific stress responses are induced. To investigate whether CDPKs are part of an underlying mechanism to guarantee response specificity, we identified CDPK-controlled signaling pathways. A truncated form of Nicotiana tabacum CDPK2 lacking its regulatory autoinhibitor and calcium-binding domains was ectopically expressed in Nicotiana benthamiana. Infiltrated leaves responded to an abiotic stress stimulus with the activation of biotic stress reactions. These responses included synthesis of reactive oxygen species, defense gene induction, and SGT1-dependent cell death. Furthermore, N-terminal CDPK2 signaling triggered enhanced levels of the phytohormones jasmonic acid, 12-oxo-phytodienoic acid, and ethylene but not salicylic acid. These responses, commonly only observed after challenge with a strong biotic stimulus, were prevented when the CDPK's intrinsic autoinhibitory peptide was coexpressed. Remarkably, elevated CDPK signaling compromised stress-induced MAPK activation, and this inhibition required ethylene synthesis and perception. These data indicate that CDPK and MAPK pathways do not function independently and that a concerted activation of both pathways controls response specificity to biotic and abiotic stress.

  3. The Role of Disordered Ribosomal Protein Extensions in the Early Steps of Eubacterial 50 S Ribosomal Subunit Assembly

    Directory of Open Access Journals (Sweden)

    Youri Timsit


    Full Text Available Although during the past decade research has shown the functional importance of disorder in proteins, many of the structural and dynamics properties of intrinsically unstructured proteins (IUPs remain to be elucidated. This review is focused on the role of the extensions of the ribosomal proteins in the early steps of the assembly of the eubacterial 50 S subunit. The recent crystallographic structures of the ribosomal particles have revealed the picture of a complex assembly pathway that condenses the rRNA and the ribosomal proteins into active ribosomes. However, little is know about the molecular mechanisms of this process. It is thought that the long basic r-protein extensions that penetrate deeply into the subunit cores play a key role through disorder-order transitions and/or co-folding mechanisms. A current view is that such structural transitions may facilitate the proper rRNA folding. In this paper, the structures of the proteins L3, L4, L13, L20, L22 and L24 that have been experimentally found to be essential for the first steps of ribosome assembly have been compared. On the basis of their structural and dynamics properties, three categories of extensions have been identified. Each of them seems to play a distinct function. Among them, only the coil-helix transition that occurs in a phylogenetically conserved cluster of basic residues of the L20 extension appears to be strictly required for the large subunit assembly in eubacteria. The role of a helix-coil transitions in 23 S RNA folding is discussed in the light of the calcium binding protein calmodulin that shares many structural and dynamics properties with L20.

  4. Cardiac troponin C-L29Q, related to hypertrophic cardiomyopathy, hinders the transduction of the protein kinase A dependent phosphorylation signal from cardiac troponin I to C. (United States)

    Schmidtmann, Anja; Lindow, Christopher; Villard, Sylvie; Heuser, Arnd; Mügge, Andreas; Gessner, Reinhard; Granier, Claude; Jaquet, Kornelia


    We investigated structural and functional aspects of the first mutation in TNNC1, coding for the calcium-binding subunit (cTnC) of cardiac troponin, which was detected in a patient with hypertrophic cardiomyopathy [ Hoffmann B, Schmidt-Traub H, Perrot A, Osterziel KJ & Gessner R (2001) Hum Mut17, 524]. This mutation leads to a leucine-glutamine exchange at position 29 in the nonfunctional calcium-binding site of cTnC. Interestingly, the mutation is located in a putative interaction site for the nonphosphorylated N-terminal arm of cardiac troponin I (cTnI) [ Finley NL, Abbott MB, Abusamhadneh E, Gaponenko V, Dong W, Seabrook G, Howarth JW, Rana M, Solaro RJ, Cheung HC et al. (1999) EJB Lett453, 107-112]. According to peptide array experiments, the nonphosphorylated cTnI arm interacts with cTnC around L29. This interaction is almost abolished by L29Q, as observed upon protein kinase A-dependent phosphorylation of cTnI at serine 22 and serine 23 in wild-type troponin. With CD spectroscopy, minor changes are observed in the backbone of Ca2+-free and Ca2+-saturated cTnC upon the L29Q replacement. A small, but significant, reduction in calcium sensitivity was detected upon measuring the Ca2+-dependent actomyosin subfragment 1 (actoS1)-ATPase activity and the sliding velocity of thin filaments. The maximum actoS1-ATPase activity, but not the maximum sliding velocity, was significantly enhanced. In addition, we performed our investigations at different levels of protein kinase A-dependent phosphorylation of cTnI. The in vitro assays mainly showed that the Ca2+ sensitivity of the actoS1-ATPase activity, and the mean sliding velocity of thin filaments, were no longer affected by protein kinase A-dependent phosphorylation of cTnI owing to the L29Q exchange in cTnC. The findings imply a hindered transduction of the phosphorylation signal from cTnI to cTnC.

  5. EFhd2, a Protein Linked to Alzheimer's Disease and Other Neurological Disorders. (United States)

    Vega, Irving E


    EFhd2 is a conserved calcium binding protein linked to different neurological disorders and types of cancer. Although, EFhd2 is more abundant in neurons, it is also found in other cell types. The physiological function of this novel protein is still unclear, but it has been shown in vitro to play a role in calcium signaling, apoptosis, actin cytoskeleton, and regulation of synapse formation. Recently, EFhd2 was shown to promote cell motility by modulating the activity of Rac1, Cdc42, and RhoA. Although, EFhd2's role in promoting cell invasion and metastasis is of great interest in cancer biology, this review focusses on the evidence that links EFhd2 to Alzheimer's disease (AD) and other neurological disorders. Altered expression of EFhd2 has been documented in AD, Parkinson's disease, Huntington's disease, Amyotrophic Lateral Sclerosis, and schizophrenia, indicating that Efhd2 gene expression is regulated in response to neuropathological processes. However, the specific role that EFhd2 plays in the pathophysiology of neurological disorders is still poorly understood. Recent studies demonstrated that EFhd2 has structural characteristics similar to amyloid proteins found in neurological disorders. Moreover, EFhd2 co-aggregates and interacts with known neuropathological proteins, such as tau, C9orf72, and Lrrk2. These results suggest that EFhd2 may play an important role in the pathophysiology of neurodegenerative diseases. Therefore, the understanding of EFhd2's role in health and disease could lead to decipher molecular mechanisms that become activated in response to neuronal stress and degeneration.

  6. Structure and interdomain interactions of a hybrid domain: a disulphide-rich module of the fibrillin/LTBP superfamily of matrix proteins. (United States)

    Jensen, Sacha A; Iqbal, Sarah; Lowe, Edward D; Redfield, Christina; Handford, Penny A


    The fibrillins and latent transforming growth factor-beta binding proteins (LTBPs) form a superfamily of structurally-related proteins consisting of calcium-binding epidermal growth factor-like (cbEGF) domains interspersed with 8-cysteine-containing transforming growth factor beta-binding protein-like (TB) and hybrid (hyb) domains. Fibrillins are the major components of the extracellular 10-12 nm diameter microfibrils, which mediate a variety of cell-matrix interactions. Here we present the crystal structure of a fibrillin-1 cbEGF9-hyb2-cbEGF10 fragment, solved to 1.8 A resolution. The hybrid domain fold is similar, but not identical, to the TB domain fold seen in previous fibrillin-1 and LTBP-1 fragments. Pairwise interactions with neighboring cbEGF domains demonstrate extensive interfaces, with the hyb2-cbEGF10 interface dependent on Ca(2+) binding. These observations provide accurate constraints for models of fibrillin organization within the 10-12 nm microfibrils and provide further molecular insights into how Ca(2+) binding influences the intermolecular interactions and biomechanical properties of fibrillin-1.

  7. Protein-protein interactions

    DEFF Research Database (Denmark)

    Byron, Olwyn; Vestergaard, Bente


    Responsive formation of protein:protein interaction (PPI) upon diverse stimuli is a fundament of cellular function. As a consequence, PPIs are complex, adaptive entities, and exist in structurally heterogeneous interplays defined by the energetic states of the free and complexed protomers. The bi...

  8. Life under tension: Computational studies of proteins involved in mechanotransduction (United States)

    Sotomayor, Marcos Manuel

    cadherins. Simulations also revealed how calcium ions control cadherin's shape and the availability of key residues involved in cell-cell adhesion, suggesting a conceptual framework for interpreting mutations in cadherin calcium binding motifs causing hereditary deafness. Overall, simulations provided a unique nanoscopic view of the dynamics and function of some of the proteins involved in mechanotransduction.

  9. Molecular cloning, expression profiles and characterization of a novel translationally controlled tumor protein in rubber tree (Hevea brasiliensis). (United States)

    Li, Dejun; Deng, Zhi; Liu, Xianghong; Qin, Bi


    The translationally controlled tumor protein (TCTP) is a multi-functioning protein that carries out vital roles in various life processes. In this study, a new TCTP gene, designated as HbTCTP1, was isolated in Hevea brasiliensis. The full-length complementary DNA (cDNA) of HbTCTP1 contained a maximum open reading frame (ORF) of 507base pair (bp) encoding 168 amino acids. The sequence comparison showed that the deduced HbTCTP1 indicated high identities to plant TCTP proteins, and clustered in the dicot cluster of plant TCTPs. Although HbTCTP1 and human TCTP proteins did not parallel in overall sequence similarity, they indicated highly similar 3D structures with a nearly identical spatial organization of α-helices, β-sheets, and coil regions. Real time reverse-transcription PCR (RT-PCR) analyses showed that HbTCTP1 was expressed throughout different tissues and developmental stages of leaves. Besides being related to tapping panel dryness (TPD), the HbTCTP1 transcripts were regulated by various treatments, including drought, low temperature, high salt, ethrel (ET), wounding, H2O2, and methyl jasmonate (Me-JA) treatments. The recombinant HbTCTP1 fusion protein was shown to protect supercoiled plasmid DNA from damages induced by metal-catalyzed generation of reactive oxygen species. The (45)Ca(2+)-overlay assay showed that HbTCTP1 was a calcium-binding protein. Our results are greatly helpful in understanding the molecular characterization and expression profiles of HbTCTP1, and lay the foundation for further analyzing the function of HbTCTP1 in rubber tree. Copyright © 2012 Elsevier GmbH. All rights reserved.

  10. Complex structure of type VI peptidoglycan muramidase effector and a cognate immunity protein

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Tianyu [Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101 (China); University of Chinese Academy of Sciences, Beijing 100049 (China); Ding, Jinjing; Zhang, Ying; Wang, Da-Cheng, E-mail: [Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101 (China); Liu, Wei, E-mail: [The Third Military Medical University, Chongqing 400038 (China); Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101 (China)


    The structure of the Tse3–Tsi3 complex associated with the bacterial type VI secretion system of P. aeruginosa has been solved and refined at 1.9 Å resolution. The structural basis of the recognition of the muramidase effector and its inactivation by its cognate immunity protein is revealed. The type VI secretion system (T6SS) is a bacterial protein-export machine that is capable of delivering virulence effectors between Gram-negative bacteria. The T6SS of Pseudomonas aeruginosa transports two lytic enzymes, Tse1 and Tse3, to degrade cell-wall peptidoglycan in the periplasm of rival bacteria that are competing for niches via amidase and muramidase activities, respectively. Two cognate immunity proteins, Tsi1 and Tsi3, are produced by the bacterium to inactivate the two antibacterial effectors, thereby protecting its siblings from self-intoxication. Recently, Tse1–Tsi1 has been structurally characterized. Here, the structure of the Tse3–Tsi3 complex is reported at 1.9 Å resolution. The results reveal that Tse3 contains a C-terminal catalytic domain that adopts a soluble lytic transglycosylase (SLT) fold in which three calcium-binding sites were surprisingly observed close to the catalytic Glu residue. The electrostatic properties of the substrate-binding groove are also distinctive from those of known structures with a similar fold. All of these features imply that a unique catalytic mechanism is utilized by Tse3 in cleaving glycosidic bonds. Tsi3 comprises a single domain showing a β-sandwich architecture that is reminiscent of the immunoglobulin fold. Three loops of Tsi3 insert deeply into the groove of Tse3 and completely occlude its active site, which forms the structural basis of Tse3 inactivation. This work is the first crystallographic report describing the three-dimensional structure of the Tse3–Tsi3 effector–immunity pair.

  11. A novel method for preparation of HAMLET-like protein complexes. (United States)

    Permyakov, Sergei E; Knyazeva, Ekaterina L; Leonteva, Marina V; Fadeev, Roman S; Chekanov, Aleksei V; Zhadan, Andrei P; Håkansson, Anders P; Akatov, Vladimir S; Permyakov, Eugene A


    Some natural proteins induce tumor-selective apoptosis. α-Lactalbumin (α-LA), a milk calcium-binding protein, is converted into an antitumor form, called HAMLET/BAMLET, via partial unfolding and association with oleic acid (OA). Besides triggering multiple cell death mechanisms in tumor cells, HAMLET exhibits bactericidal activity against Streptococcus pneumoniae. The existing methods for preparation of active complexes of α-LA with OA employ neutral pH solutions, which greatly limit water solubility of OA. Therefore these methods suffer from low scalability and/or heterogeneity of the resulting α-LA - OA samples. In this study we present a novel method for preparation of α-LA - OA complexes using alkaline conditions that favor aqueous solubility of OA. The unbound OA is removed by precipitation under acidic conditions. The resulting sample, bLA-OA-45, bears 11 OA molecules and exhibits physico-chemical properties similar to those of BAMLET. Cytotoxic activities of bLA-OA-45 against human epidermoid larynx carcinoma and S. pneumoniae D39 cells are close to those of HAMLET. Treatment of S. pneumoniae with bLA-OA-45 or HAMLET induces depolarization and rupture of the membrane. The cells are markedly rescued from death upon pretreatment with an inhibitor of Ca(2+) transport. Hence, the activation mechanisms of S. pneumoniae death are analogous for these two complexes. The developed express method for preparation of active α-LA - OA complex is high-throughput and suited for development of other protein complexes with low-molecular-weight amphiphilic substances possessing valuable cytotoxic properties. Copyright © 2011 Elsevier Masson SAS. All rights reserved.

  12. CML20, an Arabidopsis Calmodulin-like Protein, Negatively Regulates Guard Cell ABA Signaling and Drought Stress Tolerance

    Directory of Open Access Journals (Sweden)

    Xiaomeng Wu


    Full Text Available Guard cells shrink in response to drought and abscisic acid (ABA, which is caused by efflux of ions that in turn reduces stomatal aperture and improves the plant’s ability to retain moisture. Cytosolic free calcium is an essential secondary messenger in guard cell ABA signaling, but the details of this regulatory pathway remain sketchy. Here, the calmodulin-like protein CML20, which has four EF-hand domains and calcium-binding activity in vitro, was found to be a negative regulator of ABA-induced stomatal movement in Arabidopsis. The guard cells of cml20 loss-of-function mutant plants were hypersensitive to both ABA-activated S-type anion currents, and ABA inhibited inward K+ currents than those of wild type. Additional, due to smaller stomatal aperture, cml20 showed less water loss from the leaves than wild type. These phenotypes of CML20 overexpressing plants contrasted with wild type in the opposite direction. In the cml20 mutant, the transcripts of stress responsive genes, such as MYB2, RAB18, ERD10, COR47, and RD29A were up-regulated in response to drought and ABA, while down-regulated of APX2 transcription and higher reactive oxygen species (ROS accumulation. These observations support the CML20, a functional Ca2+ sensor, is a negative regulator in guard cell ABA signaling.

  13. F-Type Lectins: A Highly Diversified Family of Fucose-Binding Proteins with a Unique Sequence Motif and Structural Fold, Involved in Self/Non-Self-Recognition

    Directory of Open Access Journals (Sweden)

    Gerardo R. Vasta


    Full Text Available The F-type lectin (FTL family is one of the most recent to be identified and structurally characterized. Members of the FTL family are characterized by a fucose recognition domain [F-type lectin domain (FTLD] that displays a novel jellyroll fold (“F-type” fold and unique carbohydrate- and calcium-binding sequence motifs. This novel lectin family comprises widely distributed proteins exhibiting single, double, or greater multiples of the FTLD, either tandemly arrayed or combined with other structurally and functionally distinct domains, yielding lectin subunits of pleiotropic properties even within a single species. Furthermore, the extraordinary variability of FTL sequences (isoforms that are expressed in a single individual has revealed genetic mechanisms of diversification in ligand recognition that are unique to FTLs. Functions of FTLs in self/non-self-recognition include innate immunity, fertilization, microbial adhesion, and pathogenesis, among others. In addition, although the F-type fold is distinctive for FTLs, a structure-based search revealed apparently unrelated proteins with minor sequence similarity to FTLs that displayed the FTLD fold. In general, the phylogenetic analysis of FTLD sequences from viruses to mammals reveals clades that are consistent with the currently accepted taxonomy of extant species. However, the surprisingly discontinuous distribution of FTLDs within each taxonomic category suggests not only an extensive structural/functional diversification of the FTLs along evolutionary lineages but also that this intriguing lectin family has been subject to frequent gene duplication, secondary loss, lateral transfer, and functional co-option.

  14. EF-hand proteins and the regulation of actin-myosin interaction in the eutardigrade Hypsibius klebelsbergi (tardigrada). (United States)

    Prasath, Thiruketheeswaran; Greven, Hartmut; D'Haese, Jochen


    Many tardigrade species resist harsh environmental conditions by entering anhydrobiosis or cryobiosis. Desiccation as well as freeze resistance probably leads to changes of the ionic balance that includes the intracellular calcium concentration. In order to search for protein modifications affecting the calcium homoeostasis, we studied the regulatory system controlling actin-myosin interaction of the eutardigrade Hypsibius klebelsbergi and identified full-length cDNA clones for troponin C (TnC, 824 bp), calmodulin (CaM, 1,407 bp), essential myosin light chain (eMLC, 1,015 bp), and regulatory myosin light chain (rMLC, 984 bp) from a cDNA library. All four proteins belong to the EF-hand superfamily typified by a calcium coordinating helix-loop-helix motif. Further, we cloned and obtained recombinant TnC and both MLCs. CaM and TnC revealed four and two potential calcium-binding domains, respectively. Gel mobility shift assays demonstrated calcium-induced conformational transition of TnC. From both MLCs, only the rMLC showed one potential N-terminal EF-hand domain. Additionally, sequence properties suggest phosphorylation of this myosin light chain. Based on our results, we suggest a dual-regulated system at least in somatic muscles for tardigrades with a calcium-dependent tropomyosin-troponin complex bound to the actin filaments and a phosphorylation of the rMLC turning on and off both actin and myosin. Our results indicate no special modifications of the molecular structure and function of the EF-hand proteins in tardigrades. Phylogenetic trees of 131 TnCs, 96 rMLCs, and 62 eMLCs indicate affinities to Ecdysozoa, but also to some other taxa suggesting that our results reflect the complex evolution of these proteins rather than phylogenetic relationships. © 2012 WILEY PERIODICALS, INC.

  15. Complex structure of type VI peptidoglycan muramidase effector and a cognate immunity protein. (United States)

    Wang, Tianyu; Ding, Jinjing; Zhang, Ying; Wang, Da-Cheng; Liu, Wei


    The type VI secretion system (T6SS) is a bacterial protein-export machine that is capable of delivering virulence effectors between Gram-negative bacteria. The T6SS of Pseudomonas aeruginosa transports two lytic enzymes, Tse1 and Tse3, to degrade cell-wall peptidoglycan in the periplasm of rival bacteria that are competing for niches via amidase and muramidase activities, respectively. Two cognate immunity proteins, Tsi1 and Tsi3, are produced by the bacterium to inactivate the two antibacterial effectors, thereby protecting its siblings from self-intoxication. Recently, Tse1-Tsi1 has been structurally characterized. Here, the structure of the Tse3-Tsi3 complex is reported at 1.9 Å resolution. The results reveal that Tse3 contains a C-terminal catalytic domain that adopts a soluble lytic transglycosylase (SLT) fold in which three calcium-binding sites were surprisingly observed close to the catalytic Glu residue. The electrostatic properties of the substrate-binding groove are also distinctive from those of known structures with a similar fold. All of these features imply that a unique catalytic mechanism is utilized by Tse3 in cleaving glycosidic bonds. Tsi3 comprises a single domain showing a β-sandwich architecture that is reminiscent of the immunoglobulin fold. Three loops of Tsi3 insert deeply into the groove of Tse3 and completely occlude its active site, which forms the structural basis of Tse3 inactivation. This work is the first crystallographic report describing the three-dimensional structure of the Tse3-Tsi3 effector-immunity pair.

  16. Nonphosphorylated neurofilament protein is expressed by scattered neurons in the vestibular and precerebellar brainstem. (United States)

    Baizer, Joan S


    Vestibular information is essential for the control of posture, balance, and eye movements. The vestibular nerve projects to the four nuclei of the vestibular nuclear complex (VNC), as well as to several additional brainstem nuclei and the cerebellum. We have found that expression of the calcium-binding proteins calretinin (CR) and calbindin (CB), and the synthetic enzyme for nitric oxide synthase (nNOS) define subdivisions of the medial vestibular nucleus (MVe) and the nucleus prepositus (PrH), in cat, monkey, and human. We have asked if the pattern of expression of nonphosphorylated neurofilament protein (NPNFP) might define additional subdivisions of these or other nuclei that participate in vestibular function. We studied the distribution of cells immunoreactive to NPNFP in the brainstems of 5 cats and one squirrel monkey. Labeled cells were scattered throughout the four nuclei of the VNC, as well as in PrH, the reticular formation (RF) and the external cuneate nucleus. We used double-label immunofluorescence to visualize the distribution of these cells relative to other neurochemically defined subdivisions. NPNFP cells were excluded from the CR and CB regions of the MVe. In PrH, NPNFP and nNOS were not colocalized. Cells in the lateral vestibular nucleus and RF colocalized NPNFP and a marker for glutamatergic neurons. We also found that the cholinergic cells and axons of cranial nerve nuclei 3, 4, 6, 7,10 and 12 colocalize NPNFP. The data suggest that NPNFP is expressed by a subset of glutamatergic projection neurons of the vestibular brainstem. NPNFP may be a marker for those cells that are especially vulnerable to the effects of normal aging, neurological disease or disruption of sensory input.

  17. The rice cold-responsive calcium-dependent protein kinase OsCPK17 is regulated by alternative splicing and post-translational modifications. (United States)

    Cecília Almadanim, M; Gonçalves, Nuno M; Rosa, Margarida T G; Alexandre, Bruno M; Cordeiro, André M; Rodrigues, Mafalda; Saibo, Nelson J M; Soares, Cláudio M; Romão, Célia V; Margarida Oliveira, M; Abreu, Isabel A


    Plant calcium-dependent protein kinases (CDPKs) are key proteins implicated in calcium-mediated signaling pathways of a wide range of biological events in the organism. The action of each particular CDPK is strictly regulated by many mechanisms in order to ensure an accurate signal translation and the activation of the adequate response processes. In this work, we investigated the regulation of a CDPK involved in rice cold stress response, OsCPK17, to better understand its mode of action. We identified two new alternative splicing (AS) mRNA forms of OsCPK17 encoding truncated versions of the protein, missing the CDPK activation domain. We analyzed the expression patterns of all AS variants in rice tissues and examined their subcellular localization in onion epidermal cells. The results indicate that the AS of OsCPK17 putatively originates truncated forms of the protein with distinct functions, and different subcellular and tissue distributions. Additionally, we addressed the regulation of OsCPK17 by post-translational modifications in several in vitro experiments. Our analysis indicated that OsCPK17 activity depends on its structural rearrangement induced by calcium binding, and that the protein can be autophosphorylated. The identified phosphorylation sites mostly populate the OsCPK17 N-terminal domain. Exceptions are phosphosites T107 and S136 in the kinase domain and S558 in the C-terminal domain. These phosphosites seem conserved in CDPKs and may reflect a common regulatory mechanism for this protein family. Copyright © 2017. Published by Elsevier B.V.

  18. Comparative proteomic analysis of early salt stress-responsive proteins in roots of SnRK2 transgenic rice

    Directory of Open Access Journals (Sweden)

    Nam Myung Hee


    is an upstream regulator of stress signaling in rice roots. Enzymes involved in glycolysis, branched amino acid catabolism, dnaK-type molecular chaperone, calcium binding protein, Sal T and glyoxalase are potential targets of OSRK1 in rice roots under salt stress that need to be further investigated.

  19. Expression of genes encoding multi-transmembrane proteins in specific primate taste cell populations.

    Directory of Open Access Journals (Sweden)

    Bryan D Moyer

    Full Text Available BACKGROUND: Using fungiform (FG and circumvallate (CV taste buds isolated by laser capture microdissection and analyzed using gene arrays, we previously constructed a comprehensive database of gene expression in primates, which revealed over 2,300 taste bud-associated genes. Bioinformatics analyses identified hundreds of genes predicted to encode multi-transmembrane domain proteins with no previous association with taste function. A first step in elucidating the roles these gene products play in gustation is to identify the specific taste cell types in which they are expressed. METHODOLOGY/PRINCIPAL FINDINGS: Using double label in situ hybridization analyses, we identified seven new genes expressed in specific taste cell types, including sweet, bitter, and umami cells (TRPM5-positive, sour cells (PKD2L1-positive, as well as other taste cell populations. Transmembrane protein 44 (TMEM44, a protein with seven predicted transmembrane domains with no homology to GPCRs, is expressed in a TRPM5-negative and PKD2L1-negative population that is enriched in the bottom portion of taste buds and may represent developmentally immature taste cells. Calcium homeostasis modulator 1 (CALHM1, a component of a novel calcium channel, along with family members CALHM2 and CALHM3; multiple C2 domains; transmembrane 1 (MCTP1, a calcium-binding transmembrane protein; and anoctamin 7 (ANO7, a member of the recently identified calcium-gated chloride channel family, are all expressed in TRPM5 cells. These proteins may modulate and effect calcium signalling stemming from sweet, bitter, and umami receptor activation. Synaptic vesicle glycoprotein 2B (SV2B, a regulator of synaptic vesicle exocytosis, is expressed in PKD2L1 cells, suggesting that this taste cell population transmits tastant information to gustatory afferent nerve fibers via exocytic neurotransmitter release. CONCLUSIONS/SIGNIFICANCE: Identification of genes encoding multi-transmembrane domain proteins

  20. Defining structural and evolutionary modules in proteins: a community detection approach to explore sub-domain architecture. (United States)

    Hleap, Jose Sergio; Susko, Edward; Blouin, Christian


    Assessing protein modularity is important to understand protein evolution. Still the question of the existence of a sub-domain modular architecture remains. We propose a graph-theory approach with significance and power testing to identify modules in protein structures. In the first step, clusters are determined by optimizing the partition that maximizes the modularity score. Second, each cluster is tested for significance. Significant clusters are referred to as modules. Evolutionary modules are identified by analyzing homologous structures. Dynamic modules are inferred from sets of snapshots of molecular simulations. We present here a methodology to identify sub-domain architecture robustly, biologically meaningful, and statistically supported. The robustness of this new method is tested using simulated data with known modularity. Modules are correctly identified even when there is a low correlation between landmarks within a module. We also analyzed the evolutionary modularity of a data set of α-amylase catalytic domain homologs, and the dynamic modularity of the Niemann-Pick C1 (NPC1) protein N-terminal domain.The α-amylase contains an (α/β)8 barrel (TIM barrel) with the polysaccharides cleavage site and a calcium-binding domain. In this data set we identified four robust evolutionary modules, one of which forms the minimal functional TIM barrel topology.The NPC1 protein is involved in the intracellular lipid metabolism coordinating sterol trafficking. NPC1 N-terminus is the first luminal domain which binds to cholesterol and its oxygenated derivatives. Our inferred dynamic modules in the protein NPC1 are also shown to match functional components of the protein related to the NPC1 disease. A domain compartmentalization can be found and described in correlation space. To our knowledge, there is no other method attempting to identify sub-domain architecture from the correlation among residues. Most attempts made focus on sequence motifs of protein-protein

  1. Total protein (United States)

    ... page: // Total protein To use the sharing features on this page, please enable JavaScript. The total protein test measures the total amount of two classes ...

  2. A tracked approach for automated NMR assignments in proteins (TATAPRO)

    Energy Technology Data Exchange (ETDEWEB)

    Atreya, H.S.; Sahu, S.C.; Chary, K.V.R.; Govil, Girjesh [Tata Institute of Fundamental Research, Department of Chemical Sciences (India)


    experimental data on a calcium binding protein from Entamoeba histolytica (Eh-CaBP, 15 kDa) having substantial internal sequence homology and using published data on four other proteins in the molecular weight range of 18-42 kDa. In all the cases, nearly complete sequence specific resonance assignments (> 95%) are obtained. Furthermore, the reliability of the program has been tested by deleting sets of chemical shifts randomly from the master{sub l}ist created for the test proteins.

  3. Protein Structure (United States)

    Asmus, Elaine Garbarino


    Individual students model specific amino acids and then, through dehydration synthesis, a class of students models a protein. The students clearly learn amino acid structure, primary, secondary, tertiary, and quaternary structure in proteins and the nature of the bonds maintaining a protein's shape. This activity is fun, concrete, inexpensive and…

  4. Latent transforming growth factor β-binding protein-3 and fibulin-1C interact with the extracellular domain of the heparin-binding EGF-like growth factor precursor

    Directory of Open Access Journals (Sweden)

    Eidels Leon


    Full Text Available Abstract Background The membrane-bound cell-surface precursor and soluble forms of heparin-binding epidermal growth factor-like growth factor (HB-EGF contribute to many cellular developmental processes. The widespread occurrence of HB-EGF in cell and tissue types has led to observations of its role in such cellular and tissue events as tumor formation, cell migration, extracellular matrix formation, wound healing, and cell adherence. Several studies have reported the involvement of such extracellular matrix proteins as latent transforming growth factor β-binding protein, TGF-β, and fibulin-1 in some of these processes. To determine whether HB-EGF interacts with extracellular matrix proteins we used the extracellular domain of proHB-EGF in a yeast two-hybrid system to screen a monkey kidney cDNA library. cDNA clones containing nucleotide sequences encoding domains of two proteins were obtained and their derived amino acid sequences were evaluated. Results From ≈ 3 × 106 screened monkey cDNA clones, cDNA clones were recovered that contained nucleotide sequences encoding domains of the monkey latent transforming growth factor-β binding protein-3 (MkLTBP-3 and fibulin-1C protein. The amino acid sequence derived from the MkLTBP-3 gene shared 98.6% identity with human LTBP-3 and 86.7% identity with mouse LTBP-3 amino acid sequences. The amino acid sequence derived from the monkey fibulin-1C gene shared 97.2% identity with human fibulin-1C. Yeast two-hybrid screens indicate that LTBP-3 and fibulin-1C interact with proHB-EGF through their calcium-binding EGF-like modules. Conclusions The interactions of the extracellular domain of proHB-EGF with LTBP-3 and fibulin-1C suggest novel functions for HB-EGF between cell and tissue surfaces.

  5. Protective effects of equol and their polyphenolic isomers against dermal aging: microarray/protein evidence with clinical implications and unique delivery into human skin. (United States)

    Lephart, Edwin D


    Equol is a polyphenolic/isoflavonoid molecule that can be expressed as isomers. However, the characteristics of the equol isomers on dermal gene/protein expression and human skin percutaneous absorption remain unknown. Perform a comprehensive investigation on equol as: R-equol, racemic equol or S-equol to determine their differential expression of skin-related genes, quantify collagen expression and determine percutaneous absorption in human skin. Quantified: (i) gene expression/mRNA levels via gene array technology using human skin equivalents with equol exposure at 1.2% in qPCR experiments, (ii) in vitro collagen expression in human fibroblasts, and (iii) percutaneous absorption by Franz cell techniques. In the qPCR studies, only three genes displayed the greatest significant expression by S-equol, whereas 16 genes displayed the greatest significant levels (either stimulation or inhibition) by R-equol and/or racemic equol, such as extracellular matrix proteins (i.e., collagen and elastin), nerve growth factor, aging genes [FOS, 100 A8 and A9 calcium-binding proteins, 5α-reductase type 1, and matrix metalloproteinases (1, 3, and 9)], and inflammatory genes (e.g., interleukin-1 alpha, interleukin-6, and cyclooxygenase-1). Collagen type I expression in fibroblasts was greater with racemic versus S-equol treatment at 1 and 10 nM. Percutaneous absorption demonstrated high sequestering in keratinocytes with subsequent accumulation/release over time. Overall, these results illustrate the significant differences in mirror-image molecules or isomers of equol where R-equol and/or racemic equol are better molecules for skin gene expression compared to S-equol and the percutaneous absorption of equol represents a unique epidermal reservoir delivery mechanism.

  6. G protein-gated inwardly rectifying potassium channel subunits 1 and 2 are down-regulated in rat dorsal root ganglion neurons and spinal cord after peripheral axotomy. (United States)

    Lyu, Chuang; Mulder, Jan; Barde, Swapnali; Sahlholm, Kristoffer; Zeberg, Hugo; Nilsson, Johanna; Århem, Peter; Hökfelt, Tomas; Fried, Kaj; Shi, Tie-Jun Sten


    Increased nociceptive neuronal excitability underlies chronic pain conditions. Various ion channels, including sodium, calcium and potassium channels have pivotal roles in the control of neuronal excitability. The members of the family of G protein-gated inwardly rectifying potassium (GIRK) channels, GIRK1-4, have been implicated in modulating excitability. Here, we investigated the expression and distribution of GIRK1 and GIRK2 in normal and injured dorsal root ganglia (DRGs) and spinal cord of rats. We found that ~70% of the DRG neurons expressed GIRK1, while only <10% expressed GIRK2. The neurochemical profiles of GIRK1- and GIRK2-immunoreactive neurons were characterized using the neuronal markers calcitonin gene-related peptide, isolectin-B4 and neurofilament-200, and the calcium-binding proteins calbindin D28k, calretinin, parvalbumin and secretagogin. Both GIRK subunits were expressed in DRG neurons with nociceptive characteristics. However, while GIRK1 was widely expressed in several sensory neuronal subtypes, GIRK2 was detected mainly in a group of small C-fiber neurons. In the spinal dorsal horn, GIRK1- and -2-positive cell bodies and processes were mainly observed in lamina II, but also in superficial and deeper layers. Abundant GIRK1-, but not GIRK2-like immunoreactivity, was found in the ventral horn (laminae VI-X). Fourteen days after axotomy, GIRK1 and GIRK2 were down-regulated in DRG neurons at the mRNA and protein levels. Both after axotomy and rhizotomy there was a reduction of GIRK1- and -2-positive processes in the dorsal horn, suggesting a presynaptic localization of these potassium channels. Furthermore, nerve ligation caused accumulation of both subunits on both sides of the lesion, providing evidence for anterograde and retrograde fast axonal transport. Our data support the hypothesis that reduced GIRK function is associated with increased neuronal excitability and causes sensory disturbances in post-injury conditions, including neuropathic

  7. 70-kDa Heat Shock Cognate Protein hsc70 Mediates Calmodulin-dependent Nuclear Import of the Sex-determining Factor SRY* (United States)

    Kaur, Gurpreet; Lieu, Kim G.; Jans, David A.


    We recently showed that the developmentally important family of SOX (SRY (sex determining region on the Y chromosome)-related high mobility group (HMG) box) proteins require the calcium-binding protein calmodulin (CaM) for optimal nuclear accumulation, with clinical mutations in SRY that specifically impair nuclear accumulation via this pathway resulting in XY sex reversal. However, the mechanism by which CaM facilitates nuclear accumulation is unknown. Here, we show, for the first time, that the 70-kDa heat shock cognate protein hsc70 plays a key role in CaM-dependent nuclear import of SRY. Using a reconstituted nuclear import assay, we show that antibodies to hsc70 significantly reduce nuclear accumulation of wild type SRY and mutant derivatives thereof that retain CaM-dependent nuclear import, with an increased rate of nuclear accumulation upon addition of both CaM and hsc70, in contrast to an SRY mutant derivative with impaired CaM binding. siRNA knockdown of hsc70 in intact cells showed similar results, indicating clear dependence upon hsc70 for CaM-dependent nuclear import. Analysis using the technique of fluorescence recovery after photobleaching indicated that hsc70 is required for the maximal rate of SRY nuclear import in living cells but has no impact upon SRY nuclear retention/nuclear dynamics. Finally, we demonstrate direct binding of hsc70 to the SRY·CaM complex, with immunoprecipitation experiments from cell extracts showing association of hsc70 with wild type SRY, but not with a mutant derivative with impaired CaM binding, dependent on Ca2+. Our novel findings strongly implicate hsc70 in CaM-dependent nuclear import of SRY. PMID:23235156

  8. Clinical value of detection on ser um monocyte chemotactant protein-1 and vascular endothelial cadher in levels in patients with acute cerebral infarction

    Directory of Open Access Journals (Sweden)

    Xia Zhou


    Full Text Available Objective: To study the correlation of serum monocyte chemotactant protein-1 (MCP-1 and vascular endothelia cadherin (VE-cadherin levels in patients with acute cerebral infarction, and nerve injury molecules, interleukins and matrix metalloproteinases. Methods: A total of 86 patients with acute cerebral infarction treated in our hospital from April 2012 to October 2015 were selected as the observation group and 50 healthy subjects in the same period treated in our hospital were selected as the control group. The serums were collected and the contents of MCP-1, VE-cadherin, heart-type fatty acid binding protein (H-FABP, S100 calcium binding protein B (S100B, neuron-specific enolase (NSE, interleukin-lb (IL-1b, IL-6, IL-17, IL-18, matrix metalloproteinase-2 (MMP2, MMP3 and MMP9 were measured. Results: The serum contents of MCP-1, VE-cadherin, H-FABP, S100B, NSE, IL-1b, IL- 6, IL-17, IL-18, MMP2, MMP3 and MMP9 in observation group were significantly higher than those of control group. Carotid artery plaque formation and unstable plaque properties will increase the serum contents of MCP-1, VE-cadherin, H-FABP, S100B, NSE, IL-1b, IL-6, IL-17, IL-18, MMP2, MMP3 and MMP9 in patients with cerebral infarction. The serum levels of MCP-1, VE-cadherin and the contents of H-FABP, S100B, NSE, IL-1b, IL-6, IL-17, IL-18, MMP2, MMP3 and MMP9 were positively correlated. Conclusions: The serum levels of VE-cadherin and MCP-1 were significantly increased in patients with acute cerebral infarction. MCP-1 and VE-cadherin can increase the secretion of interleukins and matrix metalloproteinases, which can result in the carotid artery plaque formation, unstable plaque properties and the injury of nerve function.

  9. Circadian Clock Proteins and Melatonin Receptors in Neurons and Glia of the Sapajus apella Cerebellum

    Directory of Open Access Journals (Sweden)

    Leila M. Guissoni Campos


    Full Text Available Oscillations of brain proteins in circadian rhythms are important for determining several cellular and physiological processes in anticipation of daily and seasonal environmental rhythms. In addition to the suprachiasmatic nucleus, the primary central oscillator, the cerebellum shows oscillations in gene and protein expression. The variety of local circuit rhythms that the cerebellar cortex contains influences functions such as motivational processes, regulation of feeding, food anticipation, language, and working memory. The molecular basis of the cerebellar oscillator has been demonstrated by “clock gene” expression within cells of the cerebellar layers. Genetic and epidemiological evidence suggests that disruption of circadian rhythms in humans can lead to many pathological conditions. Despite this importance, data about clock gene and protein expression in the cerebellum of diurnal (day-active species, specifically primates, is currently poorly explored, mainly in regard to cellular identity, as well as the relationship with other molecules also involved in cerebellar functions. These studies could contribute to clarification of the possible mechanisms behind cerebellar rhythmicity. Considering that calcium binding proteins (CaBPs play crucial roles in preserving and modulating cerebellar functions and that clock gene expression can be controlled by afferent projections or paracrine circadian signals such as the hormone melatonin, the present study aimed to describe cellular identities, distribution patterns and day/night expression changes in PER1, PER2, CaBPs, and MT1 and MT2 melatonin receptors in the cerebellar cortex of a diurnal primate using conventional fluorescence and peroxidase-antiperoxidase immunocytochemical techniques. PER1 and PER2 immunoreactive (IR cells were observed in the Purkinje cells of the cerebellum, and MT1 and MT2 receptors were localized around Purkinje cells in the Pj layer in Bergmann cells. This identity

  10. Structural insights into the T6SS effector protein Tse3 and the Tse3-Tsi3 complex from Pseudomonas aeruginosa reveal a calcium-dependent membrane-binding mechanism. (United States)

    Lu, Defen; Shang, Guijun; Zhang, Heqiao; Yu, Qian; Cong, Xiaoyan; Yuan, Jupeng; He, Fengjuan; Zhu, Chunyuan; Zhao, Yanyu; Yin, Kun; Chen, Yuanyuan; Hu, Junqiang; Zhang, Xiaodan; Yuan, Zenglin; Xu, Sujuan; Hu, Wei; Cang, Huaixing; Gu, Lichuan


    The opportunistic pathogen Pseudomonas aeruginosa uses the type VI secretion system (T6SS) to deliver the muramidase Tse3 into the periplasm of rival bacteria to degrade their peptidoglycan (PG). Concomitantly, P. aeruginosa uses the periplasm-localized immunity protein Tsi3 to prevent potential self-intoxication caused by Tse3, and thus gains an edge over rival bacteria in fierce niche competition. Here, we report the crystal structures of Tse3 and the Tse3-Tsi3 complex. Tse3 contains an annexin repeat-like fold at the N-terminus and a G-type lysozyme fold at the C-terminus. One loop in the N-terminal domain (Loop 12) and one helix (α9) from the C-terminal domain together anchor Tse3 and the Tse3-Tsi3 complex to membrane in a calcium-dependent manner in vitro, and this membrane-binding ability is essential for Tse3's activity. In the C-terminal domain, a Y-shaped groove present on the surface likely serves as the PG binding site. Two calcium-binding motifs are also observed in the groove and these are necessary for Tse3 activity. In the Tse3-Tsi3 structure, three loops of Tsi3 insert into the substrate-binding groove of Tse3, and three calcium ions present at the interface of the complex are indispensable for the formation of the Tse3-Tsi3 complex. © 2014 John Wiley & Sons Ltd.

  11. Putative calcium-binding domains of the Caenorhabditis elegans BK channel are dispensable for intoxication and ethanol activation. (United States)

    Davis, S J; Scott, L L; Ordemann, G; Philpo, A; Cohn, J; Pierce-Shimomura, J T


    Alcohol modulates the highly conserved, voltage- and calcium-activated potassium (BK) channel, which contributes to alcohol-mediated behaviors in species from worms to humans. Previous studies have shown that the calcium-sensitive domains, RCK1 and the Ca(2+) bowl, are required for ethanol activation of the mammalian BK channel in vitro. In the nematode Caenorhabditis elegans, ethanol activates the BK channel in vivo, and deletion of the worm BK channel, SLO-1, confers strong resistance to intoxication. To determine if the conserved RCK1 and calcium bowl domains were also critical for intoxication and basal BK channel-dependent behaviors in C. elegans, we generated transgenic worms that express mutated SLO-1 channels predicted to have the RCK1, Ca(2+) bowl or both domains rendered insensitive to calcium. As expected, mutating these domains inhibited basal function of SLO-1 in vivo as neck and body curvature of these mutants mimicked that of the BK null mutant. Unexpectedly, however, mutating these domains singly or together in SLO-1 had no effect on intoxication in C. elegans. Consistent with these behavioral results, we found that ethanol activated the SLO-1 channel in vitro with or without these domains. By contrast, in agreement with previous in vitro findings, C. elegans harboring a human BK channel with mutated calcium-sensing domains displayed resistance to intoxication. Thus, for the worm SLO-1 channel, the putative calcium-sensitive domains are critical for basal in vivo function but unnecessary for in vivo ethanol action. © 2015 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.

  12. Putative calcium-binding domains of the Caenorhabditis elegans BK channel are dispensable for intoxication and ethanol activation (United States)

    Davis, S. J.; Scott, L. L.; Ordemann, G.; Philpo, A.; Cohn, J.; Pierce-Shimomura, J. T.


    Alcohol modulates the highly conserved, voltage- and calcium-activated potassium (BK) channel, which contributes to alcohol-mediated behaviors in species from worms to humans. Previous studies have shown that the calcium-sensitive domains, RCK1 and the Ca2+ bowl, are required for ethanol activation of the mammalian BK channel in vitro. In the nematode Caenorhabditis elegans, ethanol activates the BK channel in vivo, and deletion of the worm BK channel, SLO-1, confers strong resistance to intoxication. To determine if the conserved RCK1 and calcium bowl domains were also critical for intoxication and basal BK channel-dependent behaviors in C. elegans, we generated transgenic worms that express mutated SLO-1 channels predicted to have the RCK1, Ca2+ bowl or both domains rendered insensitive to calcium. As expected, mutating these domains inhibited basal function of SLO-1 in vivo as neck and body curvature of these mutants mimicked that of the BK null mutant. Unexpectedly, however, mutating these domains singly or together in SLO-1 had no effect on intoxication in C. elegans. Consistent with these behavioral results, we found that ethanol activated the SLO-1 channel in vitro with or without these domains. By contrast, in agreement with previous in vitro findings, C. elegans harboring a human BK channel with mutated calcium-sensing domains displayed resistance to intoxication. Thus, for the worm SLO-1 channel, the putative calcium-sensitive domains are critical for basal in vivo function but unnecessary for in vivo ethanol action. PMID:26113050

  13. Calcium binding restores gel formation of succinylated gelatin and reduces brittleness with preservation of the elastically stored energy

    NARCIS (Netherlands)

    Baigts Allende, Diana; Jongh, de H.H.J.


    To better tailor gelatins for textural characteristics in (food) gels, their interactions are destabilized by introduction of electrostatic repulsions and creation of affinity sites for calcium to “lock” intermolecular interactions. For that purpose gelatins with various degrees of succinylation are

  14. Calcium Binding Restores Gel Formation of Succinylated Gelatin and Reduces Brittleness with Preservation of the Elastically Stored Energy. (United States)

    Baigts Allende, Diana; de Jongh, Harmen H J


    To better tailor gelatins for textural characteristics in (food) gels, their interactions are destabilized by introduction of electrostatic repulsions and creation of affinity sites for calcium to "lock" intermolecular interactions. For that purpose gelatins with various degrees of succinylation are obtained. Extensive succinylation hampers helix formation and gel strength is slightly reduced. At high degrees of succinylation the helix propensity, gelling/melting temperatures, concomitant transition enthalpy, and gel strength become calcium-sensitive, and relatively low calcium concentrations largely restore these properties. Although succinylation has a major impact on the brittleness of the gels formed and the addition of calcium makes the material less brittle compared to nonmodified gelatin, the modification has no impact on the energy balance in the gel, where all energy applied is elastically stored in the material. This is explained by the unaffected stress relaxation by the network and high water-holding capacity related to the small mesh sizes in the gels.

  15. Anticoagulant and calcium-binding properties of high molecular weight derivatives of human fibrinogen, produced by plasmin (fragments X)

    NARCIS (Netherlands)

    Nieuwenhuizen, W.; Gravesen, M.


    Early plasmin degradation products (X fragments) of human fibrinogen were prepared in the presence of calcium-ions or EGTA, and purified on Sepharose 6B-CL. X fragments were characterized with respect to amino-terminal amino acids, polypeptide-chain composition, anticlotting properties and

  16. Whey Protein (United States)

    ... protein daily for 2 years does not improve bone density in postmenopausal women with osteoporosis. Weight loss. Most research suggests that taking whey protein alone, along with diet modifications, or while following an exercise plan does not seem to reduce weight for ...

  17. Protein Extractability

    African Journals Online (AJOL)

    limited to high oleic acid oil and water purification property (Katayon et al., 2006; Foid et al., 2001 and. Folkard et al., 1993), whereas it contains up to. 332.5 g of crude protein per kg of sample (Jose et al., 1999). Studies to characterize the interaction effects of pH and salts on the extraction of. PROTEIN EXTRACTABILITY ...

  18. Tau protein

    DEFF Research Database (Denmark)

    Frederiksen, Jette Lautrup Battistini; Kristensen, Kim; Bahl, Jmc


    Background: Tau protein has been proposed as biomarker of axonal damage leading to irreversible neurological impairment in MS. CSF concentrations may be useful when determining risk of progression from ON to MS. Objective: To investigate the association between tau protein concentration and 14......-3-3 protein in the cerebrospinal fluid (CSF) of patients with monosymptomatic optic neuritis (ON) versus patients with monosymptomatic onset who progressed to multiple sclerosis (MS). To evaluate results against data found in a complete literature review. Methods: A total of 66 patients with MS and/or ON from...... the Department of Neurology of Glostrup Hospital, University of Copenhagen, Denmark, were included. CSF samples were analysed for tau protein and 14-3-3 protein, and clinical and paraclinical information was obtained from medical records. Results: The study shows a significantly increased concentration of tau...

  19. Central Role of Protein Kinase A in Promoting Trigeminal Nociception in an In Vivo Model of Temporomandibular Disorders. (United States)

    Koop, Lindsey K; Hawkins, Jordan L; Cornelison, Lauren E; Durham, Paul L


    To investigate cellular changes in the spinal trigeminal nucleus (STN) and trigeminal ganglion (TG) associated with trigeminal nociception mediated by inflammation in the temporomandibular joint (TMJ). Male Sprague-Dawley rats (n = 86) were utilized to investigate cellular and behavioral responses to prolonged TMJ inflammation caused by bilateral injection of Complete Freund's Adjuvant (CFA) in the TMJ capsules. To investigate the cellular effects of protein kinase A (PKA) in the STN, rats were injected intrathecally with the selective PKA inhibitor KT5720 prior to injection of CFA into both TMJ capsules. Levels of calcitonin gene-related peptide (CGRP), active PKA, and ionized calcium-binding adapter molecule 1 (Iba1) in the STN and expression of phosphorylated extracellular regulated kinases (p-ERK) in the TG were determined with immunohistochemistry (n ≥ 3 experiments per test condition). Nocifensive head withdrawal responses to mechanical stimulation of the cutaneous tissue over the TMJ were monitored following CFA injection in the absence or presence of KT5720 (n = 7). Statistical analysis was performed using parametric analysis of variance (ANOVA) tests. Intrathecal injection of KT5720 significantly inhibited the stimulatory effect of CFA on levels of CGRP, PKA, and Iba1 in the STN. In addition, administration of KT5720 decreased the average number of CFA-induced nocifensive withdrawal responses to mechanical stimulation and the CFA-mediated increase in p-ERK expression in the ganglion. These findings provide evidence that elevated PKA activity in the STN promotes cellular events temporally associated with trigeminal nociception caused by prolonged TMJ inflammation.

  20. Protein-Protein Interaction Databases

    DEFF Research Database (Denmark)

    Szklarczyk, Damian; Jensen, Lars Juhl


    of research are explored. Here we present an overview of the most widely used protein-protein interaction databases and the methods they employ to gather, combine, and predict interactions. We also point out the trade-off between comprehensiveness and accuracy and the main pitfall scientists have to be aware...

  1. Dietary Proteins (United States)

    ... because your body doesn't store it the way it stores fats or carbohydrates. How much you need depends on your age, sex, health, and level of physical activity. Most Americans eat enough protein in their diet.


    GABAergic interneurons comprise the bulk of local inhibitory neuronal circuitry in cortex and hippocampus and a subpopulation of these interneurons contain the calcium binding protein, parvalbumin (PV). A previous report indicated that severe hypothyroidism reduced PV immunoreact...

  3. Molecular and Structural Characterization of the Tegumental 20.6-kDa Protein in Clonorchis sinensis as a Potential Druggable Target

    Directory of Open Access Journals (Sweden)

    Yu-Jung Kim


    Full Text Available The tegument, representing the membrane-bound outer surface of platyhelminth parasites, plays an important role for the regulation of the host immune response and parasite survival. A comprehensive understanding of tegumental proteins can provide drug candidates for use against helminth-associated diseases, such as clonorchiasis caused by the liver fluke Clonorchis sinensis. However, little is known regarding the physicochemical properties of C. sinensis teguments. In this study, a novel 20.6-kDa tegumental protein of the C. sinensis adult worm (CsTegu20.6 was identified and characterized by molecular and in silico methods. The complete coding sequence of 525 bp was derived from cDNA clones and encodes a protein of 175 amino acids. Homology search using BLASTX showed CsTegu20.6 identity ranging from 29% to 39% with previously-known tegumental proteins in C. sinensis. Domain analysis indicated the presence of a calcium-binding EF-hand domain containing a basic helix-loop-helix structure and a dynein light chain domain exhibiting a ferredoxin fold. We used a modified method to obtain the accurate tertiary structure of the CsTegu20.6 protein because of the unavailability of appropriate templates. The CsTegu20.6 protein sequence was split into two domains based on the disordered region, and then, the structure of each domain was modeled using I-TASSER. A final full-length structure was obtained by combining two structures and refining the whole structure. A refined CsTegu20.6 structure was used to identify a potential CsTegu20.6 inhibitor based on protein structure-compound interaction analysis. The recombinant proteins were expressed in Escherichia coli and purified by nickel-nitrilotriacetic acid affinity chromatography. In C. sinensis, CsTegu20.6 mRNAs were abundant in adult and metacercariae, but not in the egg. Immunohistochemistry revealed that CsTegu20.6 localized to the surface of the tegument in the adult fluke. Collectively, our results

  4. Protein Crystallization (United States)

    Chernov, Alexander A.


    Nucleation, growth and perfection of protein crystals will be overviewed along with crystal mechanical properties. The knowledge is based on experiments using optical and force crystals behave similar to inorganic crystals, though with a difference in orders of magnitude in growing parameters. For example, the low incorporation rate of large biomolecules requires up to 100 times larger supersaturation to grow protein, rather than inorganic crystals. Nucleation is often poorly reproducible, partly because of turbulence accompanying the mixing of precipitant with protein solution. Light scattering reveals fluctuations of molecular cluster size, its growth, surface energies and increased clustering as protein ages. Growth most often occurs layer-by-layer resulting in faceted crystals. New molecular layer on crystal face is terminated by a step where molecular incorporation occurs. Quantitative data on the incorporation rate will be discussed. Rounded crystals with molecularly disordered interfaces will be explained. Defects in crystals compromise the x-ray diffraction resolution crucially needed to find the 3D atomic structure of biomolecules. The defects are immobile so that birth defects stay forever. All lattice defects known for inorganics are revealed in protein crystals. Contribution of molecular conformations to lattice disorder is important, but not studied. This contribution may be enhanced by stress field from other defects. Homologous impurities (e.g., dimers, acetylated molecules) are trapped more willingly by a growing crystal than foreign protein impurities. The trapped impurities induce internal stress eliminated in crystals exceeding a critical size (part of mni for ferritin, lysozyme). Lesser impurities are trapped from stagnant, as compared to the flowing, solution. Freezing may induce much more defects unless quickly amorphysizing intracrystalline water.

  5. Computational comparison of a calcium-dependent jellyfish protein (apoaequorin) and calmodulin-cholesterol in short-term memory maintenance. (United States)

    Morrill, Gene A; Kostellow, Adele B; Gupta, Raj K


    Memory reconsolidation and maintenance depend on calcium channels and on calcium/calmodulin-dependent kinases regulating protein turnover in the hippocampus. Ingestion of a jellyfish protein, apoaequorin, reportedly protects and/or improves verbal learning in adults and is currently widely advertised for use by the elderly. Apoaequorin is a member of the EF-hand calcium binding family of proteins that includes calmodulin. Calmodulin-1 (148 residues) differs from Apoaequorin (195 residues) in that it contains four rather than three Ca 2+ -binding sites and three rather than four cholesterol-binding (CRAC, CARC) domains. All three cholesterol-binding CARC domains in calmodulin have a high interaction affinity for cholesterol compared to only two high affinity CARC domains in apoaequorin. Both calmodulin and apoaequorin can form dimers with a potential of eight bound Ca 2+ ions and six high affinity-bound cholesterol molecules in calmodulin with six bound Ca 2+ ions and a mixed population of eight cholesterols bound to both CARC and CRAC domains in apoaqueorin. MEMSAT-SVM analysis indicates that both calmodulin and apoaqueorin have a pore-lining region. The Peptide-Cutter algorithm predicts that calmodulin-1 contains 11 trypsin-specific cleavage sites (compared to 21 in apoaqueorin), four of which are potentially blocked by cholesterol and three are within the Ca-binding domains and/or the pore-lining region. Three are clustered between the third and fourth Ca 2+ -binding sites. Only calmodulin pore-lining regions contain Ca 2+ binding sites and as dimers may insert into the plasma membrane of neural cells and act as Ca 2+ channels. In a dietary supplement, bound cholesterol may protect both apoaequorin and calmodulin from proteolysis in the gut as well as facilitate uptake across the blood-brain barrier. Our results suggest that a physiological calmodulin-cholesterol complex, not cholesterol-free jellyfish protein, may better serve as a dietary supplement to

  6. Structural characterization of S100A15 reveals a novel zinc coordination site among S100 proteins and altered surface chemistry with functional implications for receptor binding

    Directory of Open Access Journals (Sweden)

    Murray Jill I


    Full Text Available Abstract Background S100 proteins are a family of small, EF-hand containing calcium-binding signaling proteins that are implicated in many cancers. While the majority of human S100 proteins share 25-65% sequence similarity, S100A7 and its recently identified paralog, S100A15, display 93% sequence identity. Intriguingly, however, S100A7 and S100A15 serve distinct roles in inflammatory skin disease; S100A7 signals through the receptor for advanced glycation products (RAGE in a zinc-dependent manner, while S100A15 signals through a yet unidentified G-protein coupled receptor in a zinc-independent manner. Of the seven divergent residues that differentiate S100A7 and S100A15, four cluster in a zinc-binding region and the remaining three localize to a predicted receptor-binding surface. Results To investigate the structural and functional consequences of these divergent clusters, we report the X-ray crystal structures of S100A15 and S100A7D24G, a hybrid variant where the zinc ligand Asp24 of S100A7 has been substituted with the glycine of S100A15, to 1.7 Å and 1.6 Å resolution, respectively. Remarkably, despite replacement of the Asp ligand, zinc binding is retained at the S100A15 dimer interface with distorted tetrahedral geometry and a chloride ion serving as an exogenous fourth ligand. Zinc binding was confirmed using anomalous difference maps and solution binding studies that revealed similar affinities of zinc for S100A15 and S100A7. Additionally, the predicted receptor-binding surface on S100A7 is substantially more basic in S100A15 without incurring structural rearrangement. Conclusions Here we demonstrate that S100A15 retains the ability to coordinate zinc through incorporation of an exogenous ligand resulting in a unique zinc-binding site among S100 proteins. The altered surface chemistry between S100A7 and S100A15 that localizes to the predicted receptor binding site is likely responsible for the differential recognition of distinct

  7. Aquaporin Protein-Protein Interactions

    Directory of Open Access Journals (Sweden)

    Jennifer Virginia Roche


    Full Text Available Aquaporins are tetrameric membrane-bound channels that facilitate transport of water and other small solutes across cell membranes. In eukaryotes, they are frequently regulated by gating or trafficking, allowing for the cell to control membrane permeability in a specific manner. Protein–protein interactions play crucial roles in both regulatory processes and also mediate alternative functions such as cell adhesion. In this review, we summarize recent knowledge about aquaporin protein–protein interactions; dividing the interactions into three types: (1 interactions between aquaporin tetramers; (2 interactions between aquaporin monomers within a tetramer (hetero-tetramerization; and (3 transient interactions with regulatory proteins. We particularly focus on the structural aspects of the interactions, discussing the small differences within a conserved overall fold that allow for aquaporins to be differentially regulated in an organism-, tissue- and trigger-specific manner. A deep knowledge about these differences is needed to fully understand aquaporin function and regulation in many physiological processes, and may enable design of compounds targeting specific aquaporins for treatment of human disease.

  8. Protein immobilization strategies for protein biochips

    NARCIS (Netherlands)

    Rusmini, F.; Rusmini, Federica; Zhong, Zhiyuan; Feijen, Jan


    In the past few years, protein biochips have emerged as promising proteomic and diagnostic tools for obtaining information about protein functions and interactions. Important technological innovations have been made. However, considerable development is still required, especially regarding protein

  9. Interaction entropy for protein-protein binding (United States)

    Sun, Zhaoxi; Yan, Yu N.; Yang, Maoyou; Zhang, John Z. H.


    Protein-protein interactions are at the heart of signal transduction and are central to the function of protein machine in biology. The highly specific protein-protein binding is quantitatively characterized by the binding free energy whose accurate calculation from the first principle is a grand challenge in computational biology. In this paper, we show how the interaction entropy approach, which was recently proposed for protein-ligand binding free energy calculation, can be applied to computing the entropic contribution to the protein-protein binding free energy. Explicit theoretical derivation of the interaction entropy approach for protein-protein interaction system is given in detail from the basic definition. Extensive computational studies for a dozen realistic protein-protein interaction systems are carried out using the present approach and comparisons of the results for these protein-protein systems with those from the standard normal mode method are presented. Analysis of the present method for application in protein-protein binding as well as the limitation of the method in numerical computation is discussed. Our study and analysis of the results provided useful information for extracting correct entropic contribution in protein-protein binding from molecular dynamics simulations.

  10. Learning about Proteins (United States)

    ... Videos for Educators Search English Español Learning About Proteins KidsHealth / For Kids / Learning About Proteins What's in ... from the foods you eat. Different Kinds of Protein Protein from animal sources, such as meat and ...

  11. A sequence-based two-level method for the prediction of type I secreted RTX proteins. (United States)

    Luo, Jiesi; Li, Wenling; Liu, Zhongyu; Guo, Yanzhi; Pu, Xuemei; Li, Menglong


    Many Gram-negative bacteria use the type I secretion system (T1SS) to translocate a wide range of substrates (type I secreted RTX proteins, T1SRPs) from the cytoplasm across the inner and outer membrane in one step to the extracellular space. Since T1SRPs play an important role in pathogen-host interactions, identifying them is crucial for a full understanding of the pathogenic mechanism of T1SS. However, experimental identification is often time-consuming and expensive. In the post-genomic era, it becomes imperative to predict new T1SRPs using information from the amino acid sequence alone when new proteins are being identified in a high-throughput mode. In this study, we report a two-level method for the first attempt to identify T1SRPs using sequence-derived features and the random forest (RF) algorithm. At the full-length sequence level, the results show that the unique feature of T1SRPs is the presence of variable numbers of the calcium-binding RTX repeats. These RTX repeats have a strong predictive power and so T1SRPs can be well distinguished from non-T1SRPs. At another level, different from that of the secretion signal, we find that a sequence segment located at the last 20-30 C-terminal amino acids may contain important signal information for T1SRP secretion because obvious differences were shown between the corresponding positions of T1SRPs and non-T1SRPs in terms of amino acid and secondary structure compositions. Using five-fold cross-validation, overall accuracies of 97% at the full-length sequence level and 89% at the secretion signal level were achieved through feature evaluation and optimization. Benchmarking on an independent dataset, our method could correctly predict 63 and 66 of 74 T1SRPs at the full-length sequence and secretion signal levels, respectively. We believe that this study will be useful in elucidating the secretion mechanism of T1SS and facilitating hypothesis-driven experimental design and validation.

  12. Efficient protein alignment algorithm for protein search. (United States)

    Lu, Zaixin; Zhao, Zhiyu; Fu, Bin


    Proteins show a great variety of 3D conformations, which can be used to infer their evolutionary relationship and to classify them into more general groups; therefore protein structure alignment algorithms are very helpful for protein biologists. However, an accurate alignment algorithm itself may be insufficient for effective discovering of structural relationships among tens of thousands of proteins. Due to the exponentially increasing amount of protein structural data, a fast and accurate structure alignment tool is necessary to access protein classification and protein similarity search; however, the complexity of current alignment algorithms are usually too high to make a fully alignment-based classification and search practical. We have developed an efficient protein pairwise alignment algorithm and applied it to our protein search tool, which aligns a query protein structure in the pairwise manner with all protein structures in the Protein Data Bank (PDB) to output similar protein structures. The algorithm can align hundreds of pairs of protein structures in one second. Given a protein structure, the tool efficiently discovers similar structures from tens of thousands of structures stored in the PDB always in 2 minutes in a single machine and 20 seconds in our cluster of 6 machines. The algorithm has been fully implemented and is accessible online at our webserver, which is supported by a cluster of computers. Our algorithm can work out hundreds of pairs of protein alignments in one second. Therefore, it is very suitable for protein search. Our experimental results show that it is more accurate than other well known protein search systems in finding proteins which are structurally similar at SCOP family and superfamily levels, and its speed is also competitive with those systems. In terms of the pairwise alignment performance, it is as good as some well known alignment algorithms.

  13. Small heat shock proteins, protein degradation and protein aggregation diseases

    NARCIS (Netherlands)

    Vos, Michel J.; Zijlstra, Marianne P.; Carra, Serena; Sibon, Ody C. M.; Kampinga, Harm H.

    Small heat shock proteins have been characterized in vitro as ATP-independent molecular chaperones that can prevent aggregation of un- or misfolded proteins and assist in their refolding with the help of ATP-dependent chaperone machines (e. g., the Hsp70 proteins). Comparison of the functionality of

  14. The C-terminal portion of BM-40 (SPARC/osteonectin) is an autonomously folding and crystallisable domain that binds calcium and collagen IV. (United States)

    Maurer, P; Hohenadl, C; Hohenester, E; Göhring, W; Timpl, R; Engel, J


    The extracellular glycoprotein BM-40 consists of three domains, an acidic domain I, a follistatin (FS)-like domain II and a calcium-binding EC domain with an EF-hand related motif. BM-40 and several other related proteins (QR1, SC1/hevin, testican and tsc-36/FRP) are members of a novel modular protein family that share the FS domain followed by an EC domain. We have expressed this pair of FS and EC domains (mutant delta I) and the calcium-binding EC domain alone (mutant delta I, II) of human BM-40 as recombinant proteins in human 293 cells. Circular dichroism demonstrated that both mutants were obtained as folded proteins with a distinct three-dimensional conformation. In addition, mutant delta I, II could be readily crystallized and diffraction patterns with a resolution limit of 2.4 A resolution were obtained. Calcium binding to this fragment was ten times weaker (Kd = 0.8 microM) than for the wild-type protein. Identical reversible increases in alpha-helicity upon calcium binding were observed for the 150-residue long mutant delta I, II and for BM-40 (286 residues). A 26-residue synthetic peptide corresponding to the EF-hand related motif exhibited much weaker calcium binding. The apparent dissociation constant decreased with increasing peptide concentration (from Kd 2.4 mM at 1 microM, to Kd 0.3 mM at 100 microM peptide concentration) and calcium binding was accompanied by dimerization of the peptide. This suggests that for strong calcium binding the EF-hand related motif has to be embedded into a larger protein domain that can form an autonomously folding protein module. The EC domain was also shown by surface plasmon resonance assay to be responsible for calcium-dependent binding to collagen IV with an affinity (Kd = 19 microM) only sixfold lower than that of intact human BM-40.

  15. EDITORIAL: Precision proteins Precision proteins (United States)

    Demming, Anna


    Since the birth of modern day medicine, during the times of Hippocrates in ancient Greece, the profession has developed from the rudimentary classification of disease into a rigorous science with an inspiring capability to treat and cure. Scientific methodology has distilled clinical diagnostic tools from the early arts of prognosis, which used to rely as much on revelation and prophecy, as intuition and judgement [1]. Over the past decade, research into the interactions between proteins and nanosystems has provided some ingenious and apt techniques for delving into the intricacies of anatomical systems. In vivo biosensing has emerged as a vibrant field of research, as much of medical diagnosis relies on the detection of substances or an imbalance in the chemicals in the body. The inherent properties of nanoscale structures, such as cantilevers, make them well suited to biosensing applications that demand the detection of molecules at very low concentrations. Measurable deflections in cantilevers functionalised with antibodies provide quantitative indicators of the presence of specific antigens when the two react. Such developments have roused mounting interest in the interactions of proteins with nanostructures, such as carbon nanotubes [3], which have demonstrated great potential as generic biomarkers. Plasmonic properties are also being exploited in sensing applications, such as the molecular sentinel recently devised by researchers in the US. The device uses the plasmonic properties of a silver nanoparticle linked to a Raman labelled hairpin DNA probe to signal changes in the probe geometry resulting from interactions with substances in the environment. Success stories so far include the detection of two specific genes associated with breast cancer [4]. A greater understanding of how RNA interference regulates gene expression has highlighted the potential of using this natural process as another agent for combating disease in personalized medicine. However, the

  16. Protein docking prediction using predicted protein-protein interface

    Directory of Open Access Journals (Sweden)

    Li Bin


    Full Text Available Abstract Background Many important cellular processes are carried out by protein complexes. To provide physical pictures of interacting proteins, many computational protein-protein prediction methods have been developed in the past. However, it is still difficult to identify the correct docking complex structure within top ranks among alternative conformations. Results We present a novel protein docking algorithm that utilizes imperfect protein-protein binding interface prediction for guiding protein docking. Since the accuracy of protein binding site prediction varies depending on cases, the challenge is to develop a method which does not deteriorate but improves docking results by using a binding site prediction which may not be 100% accurate. The algorithm, named PI-LZerD (using Predicted Interface with Local 3D Zernike descriptor-based Docking algorithm, is based on a pair wise protein docking prediction algorithm, LZerD, which we have developed earlier. PI-LZerD starts from performing docking prediction using the provided protein-protein binding interface prediction as constraints, which is followed by the second round of docking with updated docking interface information to further improve docking conformation. Benchmark results on bound and unbound cases show that PI-LZerD consistently improves the docking prediction accuracy as compared with docking without using binding site prediction or using the binding site prediction as post-filtering. Conclusion We have developed PI-LZerD, a pairwise docking algorithm, which uses imperfect protein-protein binding interface prediction to improve docking accuracy. PI-LZerD consistently showed better prediction accuracy over alternative methods in the series of benchmark experiments including docking using actual docking interface site predictions as well as unbound docking cases.

  17. Our interests in protein-protein interactions

    Indian Academy of Sciences (India)

    protein interactions. Evolution of P-P partnerships. Evolution of P-P structures. Evolutionary dynamics of P-P interactions. Dynamics of P-P interaction network. Host-pathogen interactions. CryoEM mapping of gigantic protein assemblies.

  18. Evolution of protein-protein interactions

    Indian Academy of Sciences (India)

    Evolution of protein-protein interactions · Our interests in protein-protein interactions · Slide 3 · Slide 4 · Slide 5 · Slide 6 · Slide 7 · Slide 8 · Slide 9 · Slide 10 · Slide 11 · Slide 12 · Slide 13 · Slide 14 · Slide 15 · Slide 16 · Slide 17 · Slide 18 · Slide 19 · Slide 20.

  19. 24-hour urine protein (United States)

    Urine protein - 24 hour; Chronic kidney disease - urine protein; Kidney failure - urine protein ... Bladder tumor Heart failure High blood pressure during pregnancy ( preeclampsia ) Kidney disease caused by diabetes, high blood pressure, autoimmune disorders, ...

  20. Protein in diet (United States)

    Diet - protein ... Protein foods are broken down into parts called amino acids during digestion. The human body needs a ... to eat animal products to get all the protein you need in your diet. Amino acids are ...

  1. Protein-losing enteropathy (United States)

    ... this page: // Protein-losing enteropathy To use the sharing features on this page, please enable JavaScript. Protein-losing enteropathy is an abnormal loss of protein ...

  2. EF-hands at atomic resolution: The structure of human psoriasin (S100A7) solved by MAD phasing

    DEFF Research Database (Denmark)

    Brodersen, Ditlev Egeskov; Etzerodt, Michael; Madsen, Peder Søndergaard


    The S100 family consists of small acidic proteins, belonging to the EF-hand class of calcium-binding proteins. They are primarily regulatory proteins, involved in cell growth, cell structure regulation and signal transduction. Psoriasin (S100A7) is an 11.7 kDa protein that is highly upregulated......-substituted psoriasin has been determined by multiple anomalous wavelength dispersion (MAD) phasing and refined to atomic resolution (1.05 A). The structure represents the most accurately determined structure of a calcium-binding protein. Although the overall structure of psoriasin is similar to those of other S100...... proteins, several important differences exist, mainly in the N-terminal EF-hand motif that contains a distorted loop and lacks a crucial calcium-binding residue. It is these minor differences that may account for the different specificities among members of this family. CONCLUSIONS: The structure of human...

  3. Nanotechnologies in protein microarrays. (United States)

    Krizkova, Sona; Heger, Zbynek; Zalewska, Marta; Moulick, Amitava; Adam, Vojtech; Kizek, Rene


    Protein microarray technology became an important research tool for study and detection of proteins, protein-protein interactions and a number of other applications. The utilization of nanoparticle-based materials and nanotechnology-based techniques for immobilization allows us not only to extend the surface for biomolecule immobilization resulting in enhanced substrate binding properties, decreased background signals and enhanced reporter systems for more sensitive assays. Generally in contemporarily developed microarray systems, multiple nanotechnology-based techniques are combined. In this review, applications of nanoparticles and nanotechnologies in creating protein microarrays, proteins immobilization and detection are summarized. We anticipate that advanced nanotechnologies can be exploited to expand promising fields of proteins identification, monitoring of protein-protein or drug-protein interactions, or proteins structures.

  4. Protein sequence comparison and protein evolution

    Energy Technology Data Exchange (ETDEWEB)

    Pearson, W.R. [Univ. of Virginia, Charlottesville, VA (United States). Dept. of Biochemistry


    This tutorial was one of eight tutorials selected to be presented at the Third International Conference on Intelligent Systems for Molecular Biology which was held in the United Kingdom from July 16 to 19, 1995. This tutorial examines how the information conserved during the evolution of a protein molecule can be used to infer reliably homology, and thus a shared proteinfold and possibly a shared active site or function. The authors start by reviewing a geological/evolutionary time scale. Next they look at the evolution of several protein families. During the tutorial, these families will be used to demonstrate that homologous protein ancestry can be inferred with confidence. They also examine different modes of protein evolution and consider some hypotheses that have been presented to explain the very earliest events in protein evolution. The next part of the tutorial will examine the technical aspects of protein sequence comparison. Both optimal and heuristic algorithms and their associated parameters that are used to characterize protein sequence similarities are discussed. Perhaps more importantly, they survey the statistics of local similarity scores, and how these statistics can both be used to improve the selectivity of a search and to evaluate the significance of a match. They them examine distantly related members of three protein families, the serine proteases, the glutathione transferases, and the G-protein-coupled receptors (GCRs). Finally, the discuss how sequence similarity can be used to examine internal repeated or mosaic structures in proteins.

  5. Cation binding mode of fully oxidised calmodulin explained by the unfolding of the apostate. (United States)

    Lafitte, Daniel; Tsvetkov, Philippe O; Devred, François; Toci, René; Barras, Frédéric; Briand, Claudette; Makarov, Alexander A; Haiech, Jacques


    Calmodulin is the most ubiquitous calcium binding protein. The protein is very sensitive to oxidation and this modification has pronounced effects on calmodulin function. In this work, we decided to fully oxidise calmodulin in order to study the consequences on cation binding, domain stability, and alpha helicity. Oxidation of methionines unfolds completely the apostate of the protein, which upon calcium binding recovers the major part of its secondary and tertiary structure. However, the unstructuring of the apostate results in a protein that binds calcium to any site in an independent manner, does not bind magnesium and does not possess auxiliary sites anymore.

  6. Protein- protein interaction detection system using fluorescent protein microdomains (United States)

    Waldo, Geoffrey S.; Cabantous, Stephanie


    The invention provides a protein labeling and interaction detection system based on engineered fragments of fluorescent and chromophoric proteins that require fused interacting polypeptides to drive the association of the fragments, and further are soluble and stable, and do not change the solubility of polypeptides to which they are fused. In one embodiment, a test protein X is fused to a sixteen amino acid fragment of GFP (.beta.-strand 10, amino acids 198-214), engineered to not perturb fusion protein solubility. A second test protein Y is fused to a sixteen amino acid fragment of GFP (.beta.-strand 11, amino acids 215-230), engineered to not perturb fusion protein solubility. When X and Y interact, they bring the GFP strands into proximity, and are detected by complementation with a third GFP fragment consisting of GFP amino acids 1-198 (strands 1-9). When GFP strands 10 and 11 are held together by interaction of protein X and Y, they spontaneous association with GFP strands 1-9, resulting in structural complementation, folding, and concomitant GFP fluorescence.

  7. Comparing side chain packing in soluble proteins, protein-protein interfaces, and transmembrane proteins. (United States)

    Gaines, J C; Acebes, S; Virrueta, A; Butler, M; Regan, L; O'Hern, C S


    We compare side chain prediction and packing of core and non-core regions of soluble proteins, protein-protein interfaces, and transmembrane proteins. We first identified or created comparable databases of high-resolution crystal structures of these 3 protein classes. We show that the solvent-inaccessible cores of the 3 classes of proteins are equally densely packed. As a result, the side chains of core residues at protein-protein interfaces and in the membrane-exposed regions of transmembrane proteins can be predicted by the hard-sphere plus stereochemical constraint model with the same high prediction accuracies (>90%) as core residues in soluble proteins. We also find that for all 3 classes of proteins, as one moves away from the solvent-inaccessible core, the packing fraction decreases as the solvent accessibility increases. However, the side chain predictability remains high (80% within 30°) up to a relative solvent accessibility, rSASA≲0.3, for all 3 protein classes. Our results show that ≈40% of the interface regions in protein complexes are "core", that is, densely packed with side chain conformations that can be accurately predicted using the hard-sphere model. We propose packing fraction as a metric that can be used to distinguish real protein-protein interactions from designed, non-binding, decoys. Our results also show that cores of membrane proteins are the same as cores of soluble proteins. Thus, the computational methods we are developing for the analysis of the effect of hydrophobic core mutations in soluble proteins will be equally applicable to analyses of mutations in membrane proteins. © 2018 Wiley Periodicals, Inc.

  8. IGSF9 Family Proteins

    DEFF Research Database (Denmark)

    Hansen, Maria; Walmod, Peter Schledermann


    The Drosophila protein Turtle and the vertebrate proteins immunoglobulin superfamily (IgSF), member 9 (IGSF9/Dasm1) and IGSF9B are members of an evolutionarily ancient protein family. A bioinformatics analysis of the protein family revealed that invertebrates contain only a single IGSF9 family gene......, whereas vertebrates contain two to four genes. In cnidarians, the gene appears to encode a secreted protein, but transmembrane isoforms of the protein have also evolved, and in many species, alternative splicing facilitates the expression of both transmembrane and secreted isoforms. In most species......, the longest isoforms of the proteins have the same general organization as the neural cell adhesion molecule family of cell adhesion molecule proteins, and like this family of proteins, IGSF9 family members are expressed in the nervous system. A review of the literature revealed that Drosophila Turtle...

  9. Peptide segments in protein-protein interfaces

    Indian Academy of Sciences (India)



    Sep 6, 2006 ... contact surface from the rest of the protein surface have been used to identify the interaction sites (Jones and Thornton. 1997; Neuvirth et al 2004). Protein antigenic sites (epitopes that are recognized by antibodies) could be generally confined to continuous motifs of about 8–24 amino acid residues, or may ...

  10. Surface Mediated Protein Disaggregation (United States)

    Radhakrishna, Mithun; Kumar, Sanat K.


    Preventing protein aggregation is of both biological and industrial importance. Biologically these aggregates are known to cause amyloid type diseases like Alzheimer's and Parkinson's disease. Protein aggregation leads to reduced activity of the enzymes in industrial applications. Inter-protein interactions between the hydrophobic residues of the protein are known to be the major driving force for protein aggregation. In the current paper we show how surface chemistry and curvature can be tuned to mitigate these inter-protein interactions. Our results calculated in the framework of the Hydrophobic-Polar (HP) lattice model show that, inter-protein interactions can be drastically reduced by increasing the surface hydrophobicity to a critical value corresponding to the adsorption transition of the protein. At this value of surface hydrophobicity, proteins lose inter-protein contacts to gain surface contacts and thus the surface helps in reducing the inter-protein interactions. Further, we show that the adsorption of the proteins inside hydrophobic pores of optimal sizes are most efficient both in reducing inter-protein contacts and simultaneously retaining most of the native-contacts due to strong protein-surface interactions coupled with stabilization due to the confinement. Department of Energy (Grant No DE-FG02-11ER46811).

  11. Loss of Myeloid Related Protein-8/14 Exacerbates Cardiac Allograft Rejection (United States)

    Shimizu, Koichi; Libby, Peter; Rocha, Viviane Z.; Folco, Eduardo J.; Shubiki, Rica; Grabie, Nir; Jang, Sunyoung; Lichtman, Andrew H.; Shimizu, Ayako; Hogg, Nancy; Simon, Daniel I.; Mitchell, Richard N.; Croce, Kevin


    Background The calcium-binding proteins myeloid-related protein (MRP)-8 (S100A8) and MRP-14 (S100A9) form MRP-8/14 heterodimers (S100A8/A9, calprotectin) that regulate myeloid cell function and inflammatory responses, and serve as early serum markers for monitoring acute allograft rejection. Despite functioning as a pro-inflammatory mediator, the pathophysiological role of MRP-8/14 complexes in cardiovascular disease is incompletely defined. This study investigated the role of MRP-8/14 in cardiac allograft rejection using MRP-14-deficient mice (MRP14-/-) that lack MRP-8/14 complexes. Methods and Results We examined parenchymal rejection (PR) after major histocompatibility complex (MHC) class II allomismatched cardiac transplantation (bm12 donor heart and B6 recipients) in wild-type (WT) and MRP14-/- recipients. Allograft survival averaged 5.9 ± 2.9 weeks (n=10) in MRP14-/- recipients, compared to > 12 weeks (n = 15, p MRP14-/- recipients had significantly higher PR scores (2.8 ± 0.8, n=8) than did WT recipients (0.8 ± 0.8, n=12, pMRP14-/- recipients had significantly increased T-cell and macrophage infiltration, as well as increased mRNA levels of IFN-γ and IFN-γ–associated chemokines (CXCL9, CXCL10, and CXCL11), IL-6, and IL-17, with significantly higher levels of Th17 cells. MRP14-/- recipients also had significantly more lymphocytes in the adjacent paraaortic lymph nodes than did WT recipients (cell number per lymph node: 23.7 ± 0.7 × 105 for MRP14-/- vs. 6.0 ± 0.2 × 105 for WT, p MRP14-/- recipients of bm12 hearts expressed significantly higher levels of the co-stimulatory molecules CD80 and CD86 than did those of WT recipients 2 weeks after transplantation. Mixed leukocyte reactions using allo-EC-primed MRP14-/- DCs resulted in significantly higher antigen-presenting function than reactions using WT DCs. Ovalbumin-primed MRP14-/- DCs augmented proliferation of OT-II CD4+ T cells with increased IL-2 and IFN-γ production. Cardiac allografts of B6 MHC

  12. Physics of protein motility and motor proteins (United States)

    Kolomeisky, Anatoly B.


    Motor proteins are enzymatic molecules that transform chemical energy into mechanical motion and work. They are critically important for supporting various cellular activities and functions. In the last 15 years significant progress in understanding the functioning of motor proteins has been achieved due to revolutionary breakthroughs in single-molecule experimental techniques and strong advances in theoretical modelling. However, microscopic mechanisms of protein motility are still not well explained, and the collective efforts of many scientists are needed in order to solve these complex problems. In this special section the reader will find the latest advances on the difficult road to mapping motor proteins dynamics in various systems. Recent experimental developments have allowed researchers to monitor and to influence the activity of single motor proteins with a high spatial and temporal resolution. It has stimulated significant theoretical efforts to understand the non-equilibrium nature of protein motility phenomena. The latest results from all these advances are presented and discussed in this special section. We would like to thank the scientists from all over the world who have reported their latest research results for this special section. We are also grateful to the staff and editors of Journal of Physics: Condensed Matter for their invaluable help in handling all the administrative and refereeing activities. The field of motor proteins and protein motility is fast moving, and we hope that this collection of articles will be a useful source of information in this highly interdisciplinary area. Physics of protein motility and motor proteins contents Physics of protein motility and motor proteinsAnatoly B Kolomeisky Identification of unique interactions between the flexible linker and the RecA-like domains of DEAD-box helicase Mss116 Yuan Zhang, Mirkó Palla, Andrew Sun and Jung-Chi Liao The load dependence of the physical properties of a molecular motor

  13. Polymer Directed Protein Assemblies

    Directory of Open Access Journals (Sweden)

    Patrick van Rijn


    Full Text Available Protein aggregation and protein self-assembly is an important occurrence in natural systems, and is in some form or other dictated by biopolymers. Very obvious influences of biopolymers on protein assemblies are, e.g., virus particles. Viruses are a multi-protein assembly of which the morphology is dictated by poly-nucleotides namely RNA or DNA. This “biopolymer” directs the proteins and imposes limitations on the structure like the length or diameter of the particle. Not only do these bionanoparticles use polymer-directed self-assembly, also processes like amyloid formation are in a way a result of directed protein assembly by partial unfolded/misfolded biopolymers namely, polypeptides. The combination of proteins and synthetic polymers, inspired by the natural processes, are therefore regarded as a highly promising area of research. Directed protein assembly is versatile with respect to the possible interactions which brings together the protein and polymer, e.g., electrostatic, v.d. Waals forces or covalent conjugation, and possible combinations are numerous due to the large amounts of different polymers and proteins available. The protein-polymer interacting behavior and overall morphology is envisioned to aid in clarifying protein-protein interactions and are thought to entail some interesting new functions and properties which will ultimately lead to novel bio-hybrid materials.

  14. Cytochrome c3 from Desulfovibrio gigas: crystal structure at 1.8 A resolution and evidence for a specific calcium-binding site.


    Matias, P. M.; Morais, J.; Coelho, R.; Carrondo, M. A.; Wilson, K. Van; Dauter, Z.; Sieker, L


    Crystals of the tetraheme cytochrome c3 from sulfate-reducing bacteria Desulfovibrio gigas (Dg) (MW 13 kDa, 111 residues, four heme groups) were obtained and X-ray diffraction data collected to 1.8 A resolution. The structure was solved by the method of molecular replacement and the resulting model refined to a conventional R-factor of 14.9%. The three-dimensional structure shows many similarities to other known crystal structures of tetraheme c3 cytochromes, but it also shows some remarkable...

  15. Kinetic study of the effects of calcium ions on cationic artichoke (Cynara scolymus L.) peroxidase: calcium binding, steady-state kinetics and reactions with hydrogen peroxide. (United States)

    Hiner, Alexander N P; Sidrach, Lara; Chazarra, Soledad; Varón, Ramón; Tudela, José; García-Cánovas, Francisco; Rodríguez-López, José Neptuno


    The apparent catalytic constant (k(cat)) of artichoke (Cynara scolymus L.) peroxidase (AKPC) with 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid) (ABTS) increased 130-fold in the presence of calcium ions (Ca2+) but the affinity (K(m)) of the enzyme for ABTS was 500 times lower than for Ca2+-free AKPC. AKPC is known to exhibit an equilibrium between 6-aquo hexa-coordinate and penta-coordinate forms of the haem iron that is modulated by Ca2+ and affects compound I formation. Measurements of the Ca2+ dissociation constant (K(D)) were complicated by the water-association/dissociation equilibrium yielding a global value more than 1000 times too high. The value for the Ca2+ binding step alone has now been determined to be K(D) approximately 10 nM. AKPC-Ca2+ was more resistant to inactivation by hydrogen peroxide (H(2)O(2)) and exhibited increased catalase activity. An analysis of the complex H(2)O(2) concentration dependent kinetics of Ca2+-free AKPC is presented.

  16. Functional roles of the non-catalytic calcium-binding sites in the N-terminal domain of human peptidylarginine deiminase 4.

    Directory of Open Access Journals (Sweden)

    Yi-Liang Liu

    Full Text Available This study investigated the functional roles of the N-terminal Ca(2+ ion-binding sites, in terms of enzyme catalysis and stability, of peptidylarginine deiminase 4 (PAD4. Amino acid residues located in the N-terminal Ca(2+-binding site of PAD4 were mutated to disrupt the binding of Ca(2+ ions. Kinetic data suggest that Asp155, Asp157 and Asp179, which directly coordinate Ca3 and Ca4, are essential for catalysis in PAD4. For D155A, D157A and D179A, the k(cat/K(m,BAEE values were 0.02, 0.63 and 0.01 s(-1mM(-1 (20.8 s(-1mM(-1 for WT, respectively. Asn153 and Asp176 are directly coordinated with Ca3 and indirectly coordinated with Ca5 via a water molecule. However, N153A displayed low enzymatic activity with a k(cat value of 0.3 s(-1 (13.3 s(-1 for wild-type, whereas D176A retained some catalytic power with a k(cat of 9.7 s(-1. Asp168 is the direct ligand for Ca5, and Ca5 coordination by Glu252 is mediated by two water molecules. However, mutation of these two residues to Ala did not cause a reduction in the k(cat/K(m,BAEE values, which indicates that the binding of Ca5 may not be required for PAD4 enzymatic activity. The possible conformational changes of these PAD4 mutants were examined. Thermal stability analysis of the PAD4 mutants in the absence or presence of Ca(2+ indicated that the conformational stability of the enzyme is highly dependent on Ca(2+ ions. In addition, the results of urea-induced denaturation for the N153, D155, D157 and D179 series mutants further suggest that the binding of Ca(2+ ions in the N-terminal Ca(2+-binding site stabilizes the overall conformational stability of PAD4. Therefore, our data strongly suggest that the N-terminal Ca(2+ ions play critical roles in the full activation of the PAD4 enzyme.

  17. Protein Data Bank (PDB) (United States)

    U.S. Department of Health & Human Services — The Protein Data Bank (PDB) archive is the single worldwide repository of information about the 3D structures of large biological molecules, including proteins and...

  18. Urine protein electrophoresis test (United States)

    Urine protein electrophoresis; UPEP; Multiple myeloma - UPEP; Waldenström macroglobulinemia - UPEP; Amyloidosis - UPEP ... special paper and apply an electric current. The proteins move and form visible bands. These reveal the ...

  19. Protein electrophoresis - serum (United States)

    ... this page: // Protein electrophoresis - serum To use the sharing features on ... JavaScript. This lab test measures the types of protein in the fluid (serum) part of a blood ...

  20. Statistical Properties of Protein-Protein Interfaces

    Directory of Open Access Journals (Sweden)

    Mihaly Mezei


    Full Text Available The properties of 1172 protein complexes (downloaded from the Protein Data Bank (PDB have been studied based on the concept of circular variance as a buriedness indicator and the concept of mutual proximity as a parameter-free definition of contact. The propensities of residues to be in the protein, on the surface or form contact, as well as residue pairs to form contact were calculated. In addition, the concept of circular variance has been used to compare the ruggedness and shape of the contact surface with the overall surface.

  1. Destabilized bioluminescent proteins

    Energy Technology Data Exchange (ETDEWEB)

    Allen, Michael S. (Knoxville, TN); Rakesh, Gupta (New Delhi, IN); Gary, Sayler S. (Blaine, TN)


    Purified nucleic acids, vectors and cells containing a gene cassette encoding at least one modified bioluminescent protein, wherein the modification includes the addition of a peptide sequence. The duration of bioluminescence emitted by the modified bioluminescent protein is shorter than the duration of bioluminescence emitted by an unmodified form of the bioluminescent protein.

  2. CSF total protein (United States)

    CSF total protein is a test to determine the amount of protein in your spinal fluid, also called cerebrospinal fluid (CSF). ... The normal protein range varies from lab to lab, but is typically about 15 to 60 milligrams per deciliter (mg/dL) ...

  3. Proteomic Analysis of Prostate Cancer Field Effect (United States)


    collagen synthesis , and protein translation functions. Those decreased in abundance in CaP were related to cytoskeleton and intermediate filaments...Homo sapiens] ribosomal protein SS [Homo sapiens] diazepam binding inhibitor isoform 2 [Homo sapiens] S1 00 calcium-binding protein A6 [Homo sapiens


    NARCIS (Netherlands)



    Changes in neocortical immunoreactivity (ir) for muscarinic acetylcholine receptors (mAChRs), protein kinase C gamma (PKC gamma), microtubule-associated protein 2 (MAP-2), and the calcium-binding protein parvalbumin (PARV) induced by the performance of a one-trial passive shock avoidance (PSA) task

  5. The plasma level of soluble receptor for advanced glycation end ...

    African Journals Online (AJOL)

    Anna Nashaat Abou-Raya


    Jul 23, 2015 ... cells such as nucleosomes and DNA thereby, increasing the immunogenicity of macrophages through receptors for advanced ... creatinine, 24 h urine proteins, creatinine clearance, protein/creatinine ratio, C3, C4, Anti-nuclear antibody ... for a family of about 20 related calcium binding proteins that are only ...

  6. Protein - Which is Best? (United States)

    Hoffman, Jay R; Falvo, Michael J


    Protein intake that exceeds the recommended daily allowance is widely accepted for both endurance and power athletes. However, considering the variety of proteins that are available much less is known concerning the benefits of consuming one protein versus another. The purpose of this paper is to identify and analyze key factors in order to make responsible recommendations to both the general and athletic populations. Evaluation of a protein is fundamental in determining its appropriateness in the human diet. Proteins that are of inferior content and digestibility are important to recognize and restrict or limit in the diet. Similarly, such knowledge will provide an ability to identify proteins that provide the greatest benefit and should be consumed. The various techniques utilized to rate protein will be discussed. Traditionally, sources of dietary protein are seen as either being of animal or vegetable origin. Animal sources provide a complete source of protein (i.e. containing all essential amino acids), whereas vegetable sources generally lack one or more of the essential amino acids. Animal sources of dietary protein, despite providing a complete protein and numerous vitamins and minerals, have some health professionals concerned about the amount of saturated fat common in these foods compared to vegetable sources. The advent of processing techniques has shifted some of this attention and ignited the sports supplement marketplace with derivative products such as whey, casein and soy. Individually, these products vary in quality and applicability to certain populations. The benefits that these particular proteins possess are discussed. In addition, the impact that elevated protein consumption has on health and safety issues (i.e. bone health, renal function) are also reviewed. Key PointsHigher protein needs are seen in athletic populations.Animal proteins is an important source of protein, however potential health concerns do exist from a diet of protein

  7. Antimicrobial proteins : from old proteins, new tricks


    Smith, Val; Dyrynda, Elisabeth


    This review describes the main types of antimicrobial peptides (AMPs) synthesised by crustaceans, primarily those identified in shrimp, crayfish, crab and lobster. It includes an overview of their range of microbicidal activities and the current landscape of our understanding of their gene expression patterns in different body tissues. It further summarises how their expression might change following various types of immune challenges. Included in the review are proteins or protein fragments ...

  8. Protein utilization in correlation to protein intake. (United States)

    Krajcovicová, M; Dibák, O


    In a 14-day experiment, weaned and adult rats were given ad libitum isocaloric diets with a mounting casein content (5, 10, 15, 25 and 40% by weight) and growth parameters of protein biological value, PER and NPR, and the utilization parameters NPU (body protein) and LPU (liver protein) were determined together with phosphoenolpyruvate carboxykinase (gluconeogenetic enzyme) and pyruvate kinase (glycolytic enzyme) activity in the animals' liver. The decrease in all the biological value parameters in weaned rats on 25% and 40% casein diets and in adult rats on 15%, 25% and 40% casein diets shows that these concentrations are too high for the organism. The decrease in PER and diminished weight and body and liver nitrogen increments in both age groups in animals with a low protein intake is evidence that 5% casein is an inadequate concentration. The optimum diet for weaned rats is thus a 15% casein diet and for adult rats a 10% casein diet, as confirmed by the linear correlation between weight increments, body and liver nitrogen and protein intake and also by gluconeogenetic enzyme activity. Under the given experimental conditions the study is a contribution to the determination of optimum physiological doses of proteins.

  9. Highly thermostable fluorescent proteins (United States)

    Bradbury, Andrew M [Santa Fe, NM; Waldo, Geoffrey S [Santa Fe, NM; Kiss, Csaba [Los Alamos, NM


    Thermostable fluorescent proteins (TSFPs), methods for generating these and other stability-enhanced proteins, polynucleotides encoding such proteins, and assays and method for using the TSFPs and TSFP-encoding nucleic acid molecules are provided. The TSFPs of the invention show extremely enhanced levels of stability and thermotolerance. In one case, for example, a TSFP of the invention is so stable it can be heated to C. for short periods of time without denaturing, and retains 85% of its fluorescence when heated to C. for several minutes. The invention also provides a method for generating stability-enhanced variants of a protein, including but not limited to fluorescent proteins.

  10. Protein Function Prediction. (United States)

    Cruz, Leonardo Magalhães; Trefflich, Sheyla; Weiss, Vinícius Almir; Castro, Mauro Antônio Alves


    Protein function is a concept that can have different interpretations in different biological contexts, and the number and diversity of novel proteins identified by large-scale "omics" technologies poses increasingly new challenges. In this review we explore current strategies used to predict protein function focused on high-throughput sequence analysis, as for example, inference based on sequence similarity, sequence composition, structure, and protein-protein interaction. Various prediction strategies are discussed together with illustrative workflows highlighting the use of some benchmark tools and knowledge bases in the field.

  11. Highly thermostable fluorescent proteins (United States)

    Bradbury, Andrew M [Santa Fe, NM; Waldo, Geoffrey S [Santa Fe, NM; Kiss, Csaba [Los Alamos, NM


    Thermostable fluorescent proteins (TSFPs), methods for generating these and other stability-enhanced proteins, polynucleotides encoding such proteins, and assays and method for using the TSFPs and TSFP-encoding nucleic acid molecules are provided. The TSFPs of the invention show extremely enhanced levels of stability and thermotolerance. In one case, for example, a TSFP of the invention is so stable it can be heated to C. for short periods of time without denaturing, and retains 85% of its fluorescence when heated to C. for several minutes. The invention also provides a method for generating stability-enhanced variants of a protein, including but not limited to fluorescent proteins.

  12. Protein oxidation and peroxidation

    DEFF Research Database (Denmark)

    Davies, Michael Jonathan


    and chain reactions with alcohols and carbonyls as major products; the latter are commonly used markers of protein damage. Direct oxidation of cysteine (and less commonly) methionine residues is a major reaction; this is typically faster than with H2O2, and results in altered protein activity and function....... Unlike H2O2, which is rapidly removed by protective enzymes, protein peroxides are only slowly removed, and catabolism is a major fate. Although turnover of modified proteins by proteasomal and lysosomal enzymes, and other proteases (e.g. mitochondrial Lon), can be efficient, protein hydroperoxides...

  13. Pigment-protein complexes

    Energy Technology Data Exchange (ETDEWEB)

    Siegelman, H W


    The photosynthetically-active pigment protein complexes of procaryotes and eucaryotes include chlorophyll proteins, carotenochlorophyll proteins, and biliproteins. They are either integral components or attached to photosynthetic membranes. Detergents are frequently required to solubilize the pigment-protein complexes. The membrane localization and detergent solubilization strongly suggest that the pigment-protein complexes are bound to the membranes by hydrophobic interactions. Hydrophobic interactions of proteins are characterized by an increase in entropy. Their bonding energy is directly related to temperature and ionic strength. Hydrophobic-interaction chromatography, a relatively new separation procedure, can furnish an important method for the purification of pigment-protein complexes. Phycobilisome purification and properties provide an example of the need to maintain hydrophobic interactions to preserve structure and function.

  14. Protein solubility modeling (United States)

    Agena, S. M.; Pusey, M. L.; Bogle, I. D.


    A thermodynamic framework (UNIQUAC model with temperature dependent parameters) is applied to model the salt-induced protein crystallization equilibrium, i.e., protein solubility. The framework introduces a term for the solubility product describing protein transfer between the liquid and solid phase and a term for the solution behavior describing deviation from ideal solution. Protein solubility is modeled as a function of salt concentration and temperature for a four-component system consisting of a protein, pseudo solvent (water and buffer), cation, and anion (salt). Two different systems, lysozyme with sodium chloride and concanavalin A with ammonium sulfate, are investigated. Comparison of the modeled and experimental protein solubility data results in an average root mean square deviation of 5.8%, demonstrating that the model closely follows the experimental behavior. Model calculations and model parameters are reviewed to examine the model and protein crystallization process. Copyright 1999 John Wiley & Sons, Inc.

  15. Packing in protein cores (United States)

    Gaines, J. C.; Clark, A. H.; Regan, L.; O'Hern, C. S.


    Proteins are biological polymers that underlie all cellular functions. The first high-resolution protein structures were determined by x-ray crystallography in the 1960s. Since then, there has been continued interest in understanding and predicting protein structure and stability. It is well-established that a large contribution to protein stability originates from the sequestration from solvent of hydrophobic residues in the protein core. How are such hydrophobic residues arranged in the core; how can one best model the packing of these residues, and are residues loosely packed with multiple allowed side chain conformations or densely packed with a single allowed side chain conformation? Here we show that to properly model the packing of residues in protein cores it is essential that amino acids are represented by appropriately calibrated atom sizes, and that hydrogen atoms are explicitly included. We show that protein cores possess a packing fraction of φ ≈ 0.56 , which is significantly less than the typically quoted value of 0.74 obtained using the extended atom representation. We also compare the results for the packing of amino acids in protein cores to results obtained for jammed packings from discrete element simulations of spheres, elongated particles, and composite particles with bumpy surfaces. We show that amino acids in protein cores pack as densely as disordered jammed packings of particles with similar values for the aspect ratio and bumpiness as found for amino acids. Knowing the structural properties of protein cores is of both fundamental and practical importance. Practically, it enables the assessment of changes in the structure and stability of proteins arising from amino acid mutations (such as those identified as a result of the massive human genome sequencing efforts) and the design of new folded, stable proteins and protein-protein interactions with tunable specificity and affinity.

  16. Expressed protein ligation for a large dimeric protein

    NARCIS (Netherlands)

    Karagöz, G.E.; Sinnige, T; Hsieh, O.; Rüdiger, S.G.D.


    Expressed protein ligation (EPL) is a protein engineering tool for post-translational ligation of protein or peptide fragments. This technique allows modification of specific parts of proteins, opening possibilities for incorporating probes for biophysical applications such as nuclear magnetic

  17. Toxic proteins in plants. (United States)

    Dang, Liuyi; Van Damme, Els J M


    Plants have evolved to synthesize a variety of noxious compounds to cope with unfavorable circumstances, among which a large group of toxic proteins that play a critical role in plant defense against predators and microbes. Up to now, a wide range of harmful proteins have been discovered in different plants, including lectins, ribosome-inactivating proteins, protease inhibitors, ureases, arcelins, antimicrobial peptides and pore-forming toxins. To fulfill their role in plant defense, these proteins exhibit various degrees of toxicity towards animals, insects, bacteria or fungi. Numerous studies have been carried out to investigate the toxic effects and mode of action of these plant proteins in order to explore their possible applications. Indeed, because of their biological activities, toxic plant proteins are also considered as potentially useful tools in crop protection and in biomedical applications, such as cancer treatment. Genes encoding toxic plant proteins have been introduced into crop genomes using genetic engineering technology in order to increase the plant's resistance against pathogens and diseases. Despite the availability of ample information on toxic plant proteins, very few publications have attempted to summarize the research progress made during the last decades. This review focuses on the diversity of toxic plant proteins in view of their toxicity as well as their mode of action. Furthermore, an outlook towards the biological role(s) of these proteins and their potential applications is discussed. Copyright © 2015 Elsevier Ltd. All rights reserved.


    Directory of Open Access Journals (Sweden)

    Michael J. Falvo


    Full Text Available Protein intake that exceeds the recommended daily allowance is widely accepted for both endurance and power athletes. However, considering the variety of proteins that are available much less is known concerning the benefits of consuming one protein versus another. The purpose of this paper is to identify and analyze key factors in order to make responsible recommendations to both the general and athletic populations. Evaluation of a protein is fundamental in determining its appropriateness in the human diet. Proteins that are of inferior content and digestibility are important to recognize and restrict or limit in the diet. Similarly, such knowledge will provide an ability to identify proteins that provide the greatest benefit and should be consumed. The various techniques utilized to rate protein will be discussed. Traditionally, sources of dietary protein are seen as either being of animal or vegetable origin. Animal sources provide a complete source of protein (i.e. containing all essential amino acids, whereas vegetable sources generally lack one or more of the essential amino acids. Animal sources of dietary protein, despite providing a complete protein and numerous vitamins and minerals, have some health professionals concerned about the amount of saturated fat common in these foods compared to vegetable sources. The advent of processing techniques has shifted some of this attention and ignited the sports supplement marketplace with derivative products such as whey, casein and soy. Individually, these products vary in quality and applicability to certain populations. The benefits that these particular proteins possess are discussed. In addition, the impact that elevated protein consumption has on health and safety issues (i.e. bone health, renal function are also reviewed

  19. Protein kinesis: The dynamics of protein trafficking and stability

    Energy Technology Data Exchange (ETDEWEB)



    The purpose of this conference is to provide a multidisciplinary forum for exchange of state-of-the-art information on protein kinesis. This volume contains abstracts of papers in the following areas: protein folding and modification in the endoplasmic reticulum; protein trafficking; protein translocation and folding; protein degradation; polarity; nuclear trafficking; membrane dynamics; and protein import into organelles.

  20. Protein flexibility as a biosignal. (United States)

    Zhao, Qinyi


    Dynamic properties of a protein are crucial for all protein functions, and those of signaling proteins are closely related to the biological function of living beings. The protein flexibility signal concept can be used to analyze this relationship. Protein flexibility controls the rate of protein conformational change and influences protein function. The modification of protein flexibility results in a change of protein activity. The logical nature of protein flexibility cannot be explained by applying the principles of protein three-dimensional structure theory or conformation concept. Signaling proteins show high protein flexibility. Many properties of signaling can be traced back to the dynamic natures of signaling protein. The action mechanism of volatile anesthetics and universal cellular reactions are related to flexibility in the change of signaling proteins. We conclude that protein dynamics is an enzyme-enhanced process, called dynamicase.

  1. Supramolecular Chemistry Targeting Proteins. (United States)

    van Dun, Sam; Ottmann, Christian; Milroy, Lech-Gustav; Brunsveld, Luc


    The specific recognition of protein surface elements is a fundamental challenge in the life sciences. New developments in this field will form the basis of advanced therapeutic approaches and lead to applications such as sensors, affinity tags, immobilization techniques, and protein-based materials. Synthetic supramolecular molecules and materials are creating new opportunities for protein recognition that are orthogonal to classical small molecule and protein-based approaches. As outlined here, their unique molecular features enable the recognition of amino acids, peptides, and even whole protein surfaces, which can be applied to the modulation and assembly of proteins. We believe that structural insights into these processes are of great value for the further development of this field and have therefore focused this Perspective on contributions that provide such structural data.

  2. Computational Protein Design

    DEFF Research Database (Denmark)

    Johansson, Kristoffer Enøe

    Proteins are the major functional group of molecules in biology. The impact of protein science on medicine and chemical productions is rapidly increasing. However, the greatest potential remains to be realized. The fi eld of protein design has advanced computational modeling from a tool of support...... to a central method that enables new developments. For example, novel enzymes with functions not found in natural proteins have been de novo designed to give enough activity for experimental optimization. This thesis presents the current state-of-the-art within computational design methods together...... with a novel method based on probability theory. With the aim of assembling a complete pipeline for protein design, this work touches upon several aspects of protein design. The presented work is the computational half of a design project where the other half is dedicated to the experimental part...

  3. [Erythrocyte membrane proteins]. (United States)

    Delaunay, J


    Proteins are important constituents of the red blood cell plasma membrane. Several important breakthroughs have occurred in their analysis over the past few years. SDS-polyacrylamide gel electrophoresis lead to the separation of the major proteins and glycoproteins. Location of most of these proteins -- either on the external, the internal or both surfaces of the membrane -- was determined. The strenght of the binding of the protein to the membrane was established. Hydrophobicity of membrane proteins has so far hindered their purification. However, the major glycoprotein (glycophorin A) was isolated and recently sequenced. The description of several membrane-associated enzyme activities has been followed by some understanding of their specific role in the red blood cell physiology. Abnormalities of glycoproteins, Ca2+-ATPase and of membrane protein phosphorylation have been reported under various conditions: sickle cell disease, hereditary spherocytoses, progressive muscular dystrophy.

  4. Algorithms for protein design. (United States)

    Gainza, Pablo; Nisonoff, Hunter M; Donald, Bruce R


    Computational structure-based protein design programs are becoming an increasingly important tool in molecular biology. These programs compute protein sequences that are predicted to fold to a target structure and perform a desired function. The success of a program's predictions largely relies on two components: first, the input biophysical model, and second, the algorithm that computes the best sequence(s) and structure(s) according to the biophysical model. Improving both the model and the algorithm in tandem is essential to improving the success rate of current programs, and here we review recent developments in algorithms for protein design, emphasizing how novel algorithms enable the use of more accurate biophysical models. We conclude with a list of algorithmic challenges in computational protein design that we believe will be especially important for the design of therapeutic proteins and protein assemblies. Copyright © 2016 Elsevier Ltd. All rights reserved.

  5. Mayaro virus proteins

    Directory of Open Access Journals (Sweden)

    J. M. S. Mezencio


    Full Text Available Mayaro virus was grown in BHK-21 cells and purified by centrifugation in a potassium-tartrate gradient (5-50%. The electron microscopy analyses of the purified virus showed an homogeneous population of enveloped particles with 69 ñ 2.3 nm in diameter. Three structural virus proteins were identified and designated pl, p2 and p3. Their average molecular weight were p1, 54 KDa; p2, 50 KDa and p3, 34 KDa. In Mayaro virus infected. Aedes albopictus cells and in BHK-21 infected cells we detected six viral proteins, in wich three of them are the structural virus proteins and the other three were products from processing of precursors of viral proteins, whose molecular weights are 62 KDa, 64 KDa and 110 KDa. The 34 KDa protein was the first viral protein sinthesized at 5 hours post-infection in both cell lines studied.

  6. Pressure cryocooling protein crystals (United States)

    Kim, Chae Un [Ithaca, NY; Gruner, Sol M [Ithaca, NY


    Preparation of cryocooled protein crystal is provided by use of helium pressurizing and cryocooling to obtain cryocooled protein crystal allowing collection of high resolution data and by heavier noble gas (krypton or xenon) binding followed by helium pressurizing and cryocooling to obtain cryocooled protein crystal for collection of high resolution data and SAD phasing simultaneously. The helium pressurizing is carried out on crystal coated to prevent dehydration or on crystal grown in aqueous solution in a capillary.

  7. Protein carbonylation in plants

    DEFF Research Database (Denmark)

    Møller, Ian Max; Havelund, Jesper; Rogowska-Wrzesinska, Adelina


    This chapter provides an overview of the current knowledge on protein carbonylation in plants and its role in plant physiology. It starts with a brief outline of the turnover and production sites of reactive oxygen species (ROS) in plants and the causes of protein carbonylation. This is followed...... by a description of the methods used to study protein carbonylation in plants, which is also very brief as the methods are similar to those used in studies on animals. The chapter also focuses on protein carbonylation in plants in general and in mitochondria and in seeds in particular, as case stories where...

  8. Engineering therapeutic protein disaggregases. (United States)

    Shorter, James


    Therapeutic agents are urgently required to cure several common and fatal neurodegenerative disorders caused by protein misfolding and aggregation, including amyotrophic lateral sclerosis (ALS), Parkinson's disease (PD), and Alzheimer's disease (AD). Protein disaggregases that reverse protein misfolding and restore proteins to native structure, function, and localization could mitigate neurodegeneration by simultaneously reversing 1) any toxic gain of function of the misfolded form and 2) any loss of function due to misfolding. Potentiated variants of Hsp104, a hexameric AAA+ ATPase and protein disaggregase from yeast, have been engineered to robustly disaggregate misfolded proteins connected with ALS (e.g., TDP-43 and FUS) and PD (e.g., α-synuclein). However, Hsp104 has no metazoan homologue. Metazoa possess protein disaggregase systems distinct from Hsp104, including Hsp110, Hsp70, and Hsp40, as well as HtrA1, which might be harnessed to reverse deleterious protein misfolding. Nevertheless, vicissitudes of aging, environment, or genetics conspire to negate these disaggregase systems in neurodegenerative disease. Thus, engineering potentiated human protein disaggregases or isolating small-molecule enhancers of their activity could yield transformative therapeutics for ALS, PD, and AD. © 2016 Shorter. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (

  9. Modular protein domains

    National Research Council Canada - National Science Library

    Cesareni, Giovanni


    ... encodes not only sequence, but somehow explicitly specifies folding, structure, and biological function as well. How, then, can one learn to read this 'language of proteins'? One of the most powerful approaches to 'cracking the protein code' has involved sequence comparisons between and within species, a task now greatly simplified by the ever...

  10. Advances in Protein Precipitation

    NARCIS (Netherlands)

    Golubovic, M.


    Proteins are biological macromolecules, which are among the key components of all living organisms. Proteins are nowadays present in all fields of biotech industry, such as food and feed, synthetic and pharmaceutical industry. They are isolated from their natural sources or produced in different

  11. Amino acids and proteins

    NARCIS (Netherlands)

    van Goudoever, Johannes B.; Vlaardingerbroek, Hester; van den Akker, Chris H.; de Groof, Femke; van der Schoor, Sophie R. D.


    Amino acids and protein are key factors for growth. The neonatal period requires the highest intake in life to meet the demands. Those demands include amino acids for growth, but proteins and amino acids also function as signalling molecules and function as neurotransmitters. Often the nutritional

  12. Poxviral Ankyrin Proteins

    Directory of Open Access Journals (Sweden)

    Michael H. Herbert


    Full Text Available Multiple repeats of the ankyrin motif (ANK are ubiquitous throughout the kingdoms of life but are absent from most viruses. The main exception to this is the poxvirus family, and specifically the chordopoxviruses, with ANK repeat proteins present in all but three species from separate genera. The poxviral ANK repeat proteins belong to distinct orthologue groups spread over different species, and align well with the phylogeny of their genera. This distribution throughout the chordopoxviruses indicates these proteins were present in an ancestral vertebrate poxvirus, and have since undergone numerous duplication events. Most poxviral ANK repeat proteins contain an unusual topology of multiple ANK motifs starting at the N-terminus with a C-terminal poxviral homologue of the cellular F-box enabling interaction with the cellular SCF ubiquitin ligase complex. The subtle variations between ANK repeat proteins of individual poxviruses suggest an array of different substrates may be bound by these protein-protein interaction domains and, via the F-box, potentially directed to cellular ubiquitination pathways and possible degradation. Known interaction partners of several of these proteins indicate that the NF-κB coordinated anti-viral response is a key target, whilst some poxviral ANK repeat domains also have an F-box independent affect on viral host-range.

  13. Multidomain proteins under force. (United States)

    Valle-Orero, Jessica; Rivas-Pardo, Jaime Andrés; Popa, Ionel


    Advancements in single-molecule force spectroscopy techniques such as atomic force microscopy and magnetic tweezers allow investigation of how domain folding under force can play a physiological role. Combining these techniques with protein engineering and HaloTag covalent attachment, we investigate similarities and differences between four model proteins: I10 and I91-two immunoglobulin-like domains from the muscle protein titin, and two α + β fold proteins-ubiquitin and protein L. These proteins show a different mechanical response and have unique extensions under force. Remarkably, when normalized to their contour length, the size of the unfolding and refolding steps as a function of force reduces to a single master curve. This curve can be described using standard models of polymer elasticity, explaining the entropic nature of the measured steps. We further validate our measurements with a simple energy landscape model, which combines protein folding with polymer physics and accounts for the complex nature of tandem domains under force. This model can become a useful tool to help in deciphering the complexity of multidomain proteins operating under force.

  14. NMR of unfolded proteins

    Indian Academy of Sciences (India)

    In the post-genomic era, as more and more genome sequences are becoming known and hectic efforts are underway to decode the information content in them, it is becoming increasingly evident that flexibility in proteins plays a crucial role in many of the biological functions. Many proteins have intrinsic disorder either ...

  15. Stability of Hyperthermophilic Proteins

    DEFF Research Database (Denmark)

    Stiefler-Jensen, Daniel

    in the high stability of hyperthermophilic enzymes. The thesis starts with an introduction to the field of protein and enzyme stability with special focus on the thermophilic and hyperthermophilic enzymes and proteins. After the introduction three original research manuscripts present the experimental data...

  16. Page 1 O Ag O SC (e Ce Journal of Tropical Agriculture, Food ...

    African Journals Online (AJOL)

    the cooking time and also exert other effects through the following mechanism: increasing protein solubility, breaking hydrogen bond between prolcins and collensed tallnis, renoval of calcium binding proteins, phytin and mineral complexes, inducing porous micro structure that allow enzyme-substrate interaction and cases.

  17. How do Plants Absorb Nutrients from the Soil? R -E-S-O-N-A-N-C-E ...

    Indian Academy of Sciences (India)

    utilising solar energy and the inorganic elements available in their surroundings. They obtain their carbon, ... absorb nutrients against a high concentration gradient which would have resulted otherwise due to the natural .... transduction involving the calcium binding protein, calmodulin. Carrier Proteins and Ionic Channels ...

  18. Serum calprotectin as a diagnostic marker of late onset sepsis in full ...

    African Journals Online (AJOL)

    Background: Calprotectin, a complex of two calcium-binding proteins that belong to the S100 protein family, is abundant in the cytosolic fraction of neutrophils. A high level of calprotectin reportedly exists in extracellular fluid during various inflammatory conditions, but its role in neonatal sepsis was investigated only in one ...

  19. Main: 1UHN [RPSD[Archive

    Lifescience Database Archive (English)

    Full Text Available n 2 Name=Cbl2; Arabidopsis Thaliana Molecule: Calcineurin B-Like Protein 2; Chain: A; Fragment: Residues 32-....Shimizu The Crystal Structure Of The Novel Calcium-Binding Protein Atcbl2 From Arabidopsis Thaliana J.Biol.

  20. The role of S100a4 (Mts1) in Apc- and Smad4-driven tumour onset and progression

    DEFF Research Database (Denmark)

    Atlasi, Yaser; Noori, Rubina; Marolin, Ivana


    Introduction S100a4 is a calcium-binding protein belonging to the family of S100-proteins, highly expressed in different stromal cell types. S100A4 has been reported as a prognostic marker in colorectal cancer in association with tumour progression and metastasis. Methods In this study, we analys...

  1. Transfected parvalbumin alters calcium homeostasis in teratocarcinoma PCC7 cells

    DEFF Research Database (Denmark)

    Müller, B K; Kabos, P; Belhage, B


    Indirect evidence supports a protective role of some EF-hand calcium-binding proteins against calcium-induced neurotoxicity. Little is known about how these proteins influence cytosolic calcium levels. After cloning the parvalbumin cDNA into an expression vector, teratocarcinoma cells (PCC7) were...

  2. Protein expression-yeast. (United States)

    Nielsen, Klaus H


    Yeast is an excellent system for the expression of recombinant eukaryotic proteins. Both endogenous and heterologous proteins can be overexpressed in yeast (Phan et al., 2001; Ton and Rao, 2004). Because yeast is easy to manipulate genetically, a strain can be optimized for the expression of a specific protein. Many eukaryotic proteins contain posttranslational modifications that can be performed in yeast but not in bacterial expression systems. In comparison with mammalian cell culture expression systems, growing yeast is both faster and less expensive, and large-scale cultures can be performed using fermentation. While several different yeast expression systems exist, this chapter focuses on the budding yeast Saccharomyces cerevisiae and will briefly describe some options to consider when selecting vectors and tags to be used for protein expression. Throughout this chapter, the expression and purification of yeast eIF3 is shown as an example alongside a general scheme outline. © 2014 Elsevier Inc. All rights reserved.

  3. MicroProteins

    DEFF Research Database (Denmark)

    Eguen, Teinai Ebimienere; Straub, Daniel; Graeff, Moritz


    MicroProteins (miPs) are short, usually single-domain proteins that, in analogy to miRNAs, heterodimerize with their targets and exert a dominant-negative effect. Recent bioinformatic attempts to identify miPs have resulted in a list of potential miPs, many of which lack the defining characterist......MicroProteins (miPs) are short, usually single-domain proteins that, in analogy to miRNAs, heterodimerize with their targets and exert a dominant-negative effect. Recent bioinformatic attempts to identify miPs have resulted in a list of potential miPs, many of which lack the defining...... can extend beyond transcription factors (TFs) to encompass different non-TF proteins that require dimerization for full function....

  4. Protein disulfide engineering. (United States)

    Dombkowski, Alan A; Sultana, Kazi Zakia; Craig, Douglas B


    Improving the stability of proteins is an important goal in many biomedical and industrial applications. A logical approach is to emulate stabilizing molecular interactions found in nature. Disulfide bonds are covalent interactions that provide substantial stability to many proteins and conform to well-defined geometric conformations, thus making them appealing candidates in protein engineering efforts. Disulfide engineering is the directed design of novel disulfide bonds into target proteins. This important biotechnological tool has achieved considerable success in a wide range of applications, yet the rules that govern the stabilizing effects of disulfide bonds are not fully characterized. Contrary to expectations, many designed disulfide bonds have resulted in decreased stability of the modified protein. We review progress in disulfide engineering, with an emphasis on the issue of stability and computational methods that facilitate engineering efforts. Copyright © 2013 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  5. Artificially Engineered Protein Polymers. (United States)

    Yang, Yun Jung; Holmberg, Angela L; Olsen, Bradley D


    Modern polymer science increasingly requires precise control over macromolecular structure and properties for engineering advanced materials and biomedical systems. The application of biological processes to design and synthesize artificial protein polymers offers a means for furthering macromolecular tunability, enabling polymers with dispersities of ∼1.0 and monomer-level sequence control. Taking inspiration from materials evolved in nature, scientists have created modular building blocks with simplified monomer sequences that replicate the function of natural systems. The corresponding protein engineering toolbox has enabled the systematic development of complex functional polymeric materials across areas as diverse as adhesives, responsive polymers, and medical materials. This review discusses the natural proteins that have inspired the development of key building blocks for protein polymer engineering and the function of these elements in material design. The prospects and progress for scalable commercialization of protein polymers are reviewed, discussing both technology needs and opportunities.

  6. Sensitizing properties of proteins

    DEFF Research Database (Denmark)

    Poulsen, Lars K.; Ladics, Gregory S; McClain, Scott


    scientists from academia, government, and industry participated in the symposium. Experts provided overviews on known mechanisms by which proteins in food may cause sensitization, discussed experimental models to predict protein sensitizing potential, and explored whether such experimental techniques may......The scope of allergy risk is diverse considering the myriad ways in which protein allergenicity is affected by physiochemical characteristics of proteins. The complexity created by the matrices of foods and the variability of the human immune system add additional challenges to understanding...... Allergenicity Technical Committee of the International Life Sciences Institute's Health and Environmental Sciences Institute, featured presentations on current methods, test systems, research trends, and unanswered questions in the field of protein sensitization. A diverse group of over 70 interdisciplinary...

  7. Sensitizing properties of proteins

    DEFF Research Database (Denmark)

    Poulsen, Lars K.; Ladics, Gregory S; McClain, Scott


    The scope of allergy risk is diverse considering the myriad ways in which protein allergenicity is affected by physiochemical characteristics of proteins. The complexity created by the matrices of foods and the variability of the human immune system add additional challenges to understanding...... the relationship between sensitization potential and allergy disease. To address these and other issues, an April 2012 international symposium was held in Prague, Czech Republic, to review and discuss the state-of-the-science of sensitizing properties of protein allergens. The symposium, organized by the Protein...... Allergenicity Technical Committee of the International Life Sciences Institute's Health and Environmental Sciences Institute, featured presentations on current methods, test systems, research trends, and unanswered questions in the field of protein sensitization. A diverse group of over 70 interdisciplinary...

  8. [Controversies around diet proteins]. (United States)

    Cichosz, Grazyna; Czeczot, Hanna


    Critical theories regarding proteins of anima origin are still and still popularized, though they are ungrounded from scientific point of view. Predominance of soya proteins over the animal ones in relation to their influence on calcium metabolism, bone break risk or risk of osteoporosis morbidity has not been confirmed in any honest, reliable research experiment. Statement, that sulphur amino acids influence disadvantageously on calcium metabolism of human organism and bone status, is completely groundless, the more so as presence of sulphur amino acids in diet (animal proteins are their best source) is the condition of endogenic synthesis of glutathione, the key antioxidant of the organism, and taurine stimulating brain functioning. Deficiency of proteins in the diet produce weakness of intellectual effectiveness and immune response. There is no doubt that limitation of consumption of animal proteins of standard value is not good for health.

  9. Conditional Function of Autoaggregative Protein Cah and Common cah Mutations in Shiga Toxin-Producing Escherichia coli. (United States)

    Carter, Michelle Qiu; Brandl, Maria T; Kudva, Indira T; Katani, Robab; Moreau, Matthew R; Kapur, Vivek


    Cah is a calcium-binding autotransporter protein involved in autoaggregation and biofilm formation. Although cah is widespread in Shiga toxin-producing Escherichia coli (STEC), we detected mutations in cah at a frequency of 31.3% in this pathogen. In STEC O157:H7 super-shedder strain SS17, a large deletion results in a smaller coding sequence, lacking the C-terminal 71 amino acids compared with Cah in STEC O157:H7 strain EDL933. We examined the function of Cah in biofilm formation and host colonization to better understand selective pressures for cah mutations. EDL933-Cah played a conditional role in biofilm formation in vitro: it enhanced E. coli DH5α biofilm formation on glass surfaces under agitated culture conditions that prevented autoaggregation, but inhibited biofilm formation under hydrostatic conditions that facilitated autoaggregation. This function appeared to be strain-dependent since Cah-mediated biofilm formation was diminished when an EDL933-cah was expressed in SS17. Deletion of cah in EDL933 enhanced bacterial attachment to spinach leaves and altered the adherence pattern of EDL933 to bovine recto-anal junction squamous epithelial (RSE) cells. In contrast, in trans-expression of EDL933-cah in SS17 increased its attachment to leaf surfaces, and in DH5α, enhanced its adherence to RSE cells. Hence the ecological function of Cah appears to be modulated by environmental conditions and other bacterial strain-specific properties. Considering the prevalence of cah in STEC and its role in attachment and biofilm formation, cah mutations might be selected in ecological niches where inactivation of Cah would result in an increased fitness in STEC during colonization of plants or animal hosts.ImportanceShiga toxin-producing Escherichia coli (STEC) harbors genes encoding diverse adhesins and many of these are known to play an important role in bacterial attachment and host colonization. We demonstrated here that the autotransporter protein Cah confers E. coli

  10. Coarse-grain modelling of protein-protein interactions

    NARCIS (Netherlands)

    Baaden, Marc; Marrink, Siewert J.


    Here, we review recent advances towards the modelling of protein-protein interactions (PPI) at the coarse-grained (CG) level, a technique that is now widely used to understand protein affinity, aggregation and self-assembly behaviour. PPI models of soluble proteins and membrane proteins are

  11. Swaps in protein sequences. (United States)

    Fliess, Amit; Motro, Benny; Unger, Ron


    An important question in protein evolution is to what extent proteins may have undergone swaps (switches of domain or fragment order) during evolution. Such events might have occurred in several forms: Swaps of short fragments, swaps of structural and functional motifs, or recombination of domains in multidomain proteins. This question is important for the theoretical understanding of the evolution of proteins, and has practical implications for using swaps as a design tool in protein engineering. In order to analyze the question systematically, we conducted a large scale survey of possible swaps and permutations among all pairs of protein from the Swissport database. A swap is defined as a specific kind of sequence mutation between two proteins in which two fragments that appear in both sequences have different relative order in the two sequences. For example, aXbYc and dYeXf are defined as a swap, where X and Y represent sequence fragments that switched their order. Identifying such swaps is difficult using standard sequence comparison packages. One of the main problems in the analysis stems from the fact that many sequences contain repeats, which may be identified as false-positive swaps. We have used two different approaches to detect pairs of proteins with swaps. The first approach is based on the predefined list of domains in Pfam. We identified all the proteins that share at least two domains and analyzed their relative order, looking for pairs in which the order of these domains was switched. We designed an algorithm to distinguish between real swaps and duplications. In the second approach, we used Blast to detect pairs of proteins that share several fragments. Then, we used an automatic procedure to select pairs that are likely to contain swaps. Those pairs were analyzed visually, using a graphical tool, to eliminate duplications. Combining these approaches, about 140 different cases of swaps in the Swissprot database were found (after eliminating

  12. Anchored design of protein-protein interfaces.

    Directory of Open Access Journals (Sweden)

    Steven M Lewis

    Full Text Available Few existing protein-protein interface design methods allow for extensive backbone rearrangements during the design process. There is also a dichotomy between redesign methods, which take advantage of the native interface, and de novo methods, which produce novel binders.Here, we propose a new method for designing novel protein reagents that combines advantages of redesign and de novo methods and allows for extensive backbone motion. This method requires a bound structure of a target and one of its natural binding partners. A key interaction in this interface, the anchor, is computationally grafted out of the partner and into a surface loop on the design scaffold. The design scaffold's surface is then redesigned with backbone flexibility to create a new binding partner for the target. Careful choice of a scaffold will bring experimentally desirable characteristics into the new complex. The use of an anchor both expedites the design process and ensures that binding proceeds against a known location on the target. The use of surface loops on the scaffold allows for flexible-backbone redesign to properly search conformational space.This protocol was implemented within the Rosetta3 software suite. To demonstrate and evaluate this protocol, we have developed a benchmarking set of structures from the PDB with loop-mediated interfaces. This protocol can recover the correct loop-mediated interface in 15 out of 16 tested structures, using only a single residue as an anchor.

  13. Antimicrobial proteins: From old proteins, new tricks. (United States)

    Smith, Valerie J; Dyrynda, Elisabeth A


    This review describes the main types of antimicrobial peptides (AMPs) synthesised by crustaceans, primarily those identified in shrimp, crayfish, crab and lobster. It includes an overview of their range of microbicidal activities and the current landscape of our understanding of their gene expression patterns in different body tissues. It further summarises how their expression might change following various types of immune challenges. The review further considers proteins or protein fragments from crustaceans that have antimicrobial properties but are more usually associated with other biological functions, or are derived from such proteins. It discusses how these unconventional AMPs might be generated at, or delivered to, sites of infection and how they might contribute to crustacean host defence in vivo. It also highlights recent work that is starting to reveal the extent of multi-functionality displayed by some decapod AMPs, particularly their participation in other aspects of host protection. Examples of such activities include proteinase inhibition, phagocytosis, antiviral activity and haematopoiesis. Copyright © 2015 Elsevier Ltd. All rights reserved.

  14. Multidomain proteins under force (United States)

    Valle-Orero, Jessica; Andrés Rivas-Pardo, Jaime; Popa, Ionel


    Advancements in single-molecule force spectroscopy techniques such as atomic force microscopy and magnetic tweezers allow investigation of how domain folding under force can play a physiological role. Combining these techniques with protein engineering and HaloTag covalent attachment, we investigate similarities and differences between four model proteins: I10 and I91—two immunoglobulin-like domains from the muscle protein titin, and two α + β fold proteins—ubiquitin and protein L. These proteins show a different mechanical response and have unique extensions under force. Remarkably, when normalized to their contour length, the size of the unfolding and refolding steps as a function of force reduces to a single master curve. This curve can be described using standard models of polymer elasticity, explaining the entropic nature of the measured steps. We further validate our measurements with a simple energy landscape model, which combines protein folding with polymer physics and accounts for the complex nature of tandem domains under force. This model can become a useful tool to help in deciphering the complexity of multidomain proteins operating under force.

  15. Protein oxidation in aquatic foods

    DEFF Research Database (Denmark)

    Baron, Caroline P.


    The chapter discusses general considerations about protein oxidation and reviews the mechanisms involved in protein oxidation and consequences of protein oxidation on fish proteins. It presents two case studies, the first deals with protein and lipid oxidation in frozen rainbow trout......, and the second with oxidation in salted herring. The mechanisms responsible for initiation of protein oxidation are unclear, but it is generally accepted that free radical species initiating lipid oxidation can also initiate protein oxidation. The chapter focuses on interaction between protein and lipid...... oxidation. The protein carbonyl group measurement is the widely used method for estimating protein oxidation in foods and has been used in fish muscle. The chapter also talks about the impact of protein oxidation on protein functionality, fish muscle texture, and food nutritional value. Protein oxidation...

  16. Sound of proteins

    DEFF Research Database (Denmark)


    In my group we work with Molecular Dynamics to model several different proteins and protein systems. We submit our modelled molecules to changes in temperature, changes in solvent composition and even external pulling forces. To analyze our simulation results we have so far used visual inspection...... and statistical analysis of the resulting molecular trajectories (as everybody else!). However, recently I started assigning a particular sound frequency to each amino acid in the protein, and by setting the amplitude of each frequency according to the movement amplitude we can "hear" whenever two aminoacids...

  17. PDP: protein domain parser. (United States)

    Alexandrov, Nickolai; Shindyalov, Ilya


    We have developed a program for automatic identification of domains in protein three-dimensional structures. Performance of the program was assessed by three different benchmarks: (i) by comparison with the expert-curated SCOP database of structural domains; (ii) by comparison with a collection of manual domain assignments; and (iii) by comparison with a set of 55 proteins, frequently used as a benchmark for automatic domain assignment. In all these benchmarks PDP identified domains correctly in more than 80% of proteins.

  18. Alpha Shapes and Proteins

    DEFF Research Database (Denmark)

    Winter, Pawel; Sterner, Henrik; Sterner, Peter


    We provide a unified description of (weighted) alpha shapes, beta shapes and the corresponding simplicialcomplexes. We discuss their applicability to various protein-related problems. We also discuss filtrations of alpha shapes and touch upon related persistence issues.We claim that the full...... potential of alpha-shapes and related geometrical constructs in protein-related problems yet remains to be realized and verified. We suggest parallel algorithms for (weighted) alpha shapes, and we argue that future use of filtrations and kinetic variants for larger proteins will need such implementation....

  19. Designing microcapsules based on protein fibrils and protein - polysaccharide complexes

    NARCIS (Netherlands)

    Hua, K.N.P.


    Keywords: encapsulation, microcapsule, protein, fibril, protein-polysaccharide complex, controlled release, interfacial rheology, lysozyme, ovalbumin This thesis describes the design of encapsulation systems using mesostructures from proteins and polysaccharides. The approach was to first

  20. Polymers for Protein Conjugation

    Directory of Open Access Journals (Sweden)

    Gianfranco Pasut


    Full Text Available Polyethylene glycol (PEG at the moment is considered the leading polymer for protein conjugation in view of its unique properties, as well as to its low toxicity in humans, qualities which have been confirmed by its extensive use in clinical practice. Other polymers that are safe, biodegradable and custom-designed have, nevertheless, also been investigated as potential candidates for protein conjugation. This review will focus on natural polymers and synthetic linear polymers that have been used for protein delivery and the results associated with their use. Genetic fusion approaches for the preparation of protein-polypeptide conjugates will be also reviewed and compared with the best known chemical conjugation ones.

  1. Electron transfer in proteins

    DEFF Research Database (Denmark)

    Farver, O; Pecht, I


    Electron migration between and within proteins is one of the most prevalent forms of biological energy conversion processes. Electron transfer reactions take place between active centers such as transition metal ions or organic cofactors over considerable distances at fast rates and with remarkable...... specificity. The electron transfer is attained through weak electronic interaction between the active sites, so that considerable research efforts are centered on resolving the factors that control the rates of long-distance electron transfer reactions in proteins. These factors include (in addition......-containing proteins. These proteins serve almost exclusively in electron transfer reactions, and as it turns out, their metal coordination sites are endowed with properties uniquely optimized for their function....

  2. Protein Colloidal Aggregation Project (United States)

    Oliva-Buisson, Yvette J. (Compiler)


    To investigate the pathways and kinetics of protein aggregation to allow accurate predictive modeling of the process and evaluation of potential inhibitors to prevalent diseases including cataract formation, chronic traumatic encephalopathy, Alzheimer's Disease, Parkinson's Disease and others.

  3. Interactive protein manipulation

    Energy Technology Data Exchange (ETDEWEB)


    We describe an interactive visualization and modeling program for the creation of protein structures ''from scratch''. The input to our program is an amino acid sequence -decoded from a gene- and a sequence of predicted secondary structure types for each amino acid-provided by external structure prediction programs. Our program can be used in the set-up phase of a protein structure prediction process; the structures created with it serve as input for a subsequent global internal energy minimization, or another method of protein structure prediction. Our program supports basic visualization methods for protein structures, interactive manipulation based on inverse kinematics, and visualization guides to aid a user in creating ''good'' initial structures.

  4. Parallel Computational Protein Design. (United States)

    Zhou, Yichao; Donald, Bruce R; Zeng, Jianyang


    Computational structure-based protein design (CSPD) is an important problem in computational biology, which aims to design or improve a prescribed protein function based on a protein structure template. It provides a practical tool for real-world protein engineering applications. A popular CSPD method that guarantees to find the global minimum energy solution (GMEC) is to combine both dead-end elimination (DEE) and A* tree search algorithms. However, in this framework, the A* search algorithm can run in exponential time in the worst case, which may become the computation bottleneck of large-scale computational protein design process. To address this issue, we extend and add a new module to the OSPREY program that was previously developed in the Donald lab (Gainza et al., Methods Enzymol 523:87, 2013) to implement a GPU-based massively parallel A* algorithm for improving protein design pipeline. By exploiting the modern GPU computational framework and optimizing the computation of the heuristic function for A* search, our new program, called gOSPREY, can provide up to four orders of magnitude speedups in large protein design cases with a small memory overhead comparing to the traditional A* search algorithm implementation, while still guaranteeing the optimality. In addition, gOSPREY can be configured to run in a bounded-memory mode to tackle the problems in which the conformation space is too large and the global optimal solution cannot be computed previously. Furthermore, the GPU-based A* algorithm implemented in the gOSPREY program can be combined with the state-of-the-art rotamer pruning algorithms such as iMinDEE (Gainza et al., PLoS Comput Biol 8:e1002335, 2012) and DEEPer (Hallen et al., Proteins 81:18-39, 2013) to also consider continuous backbone and side-chain flexibility.

  5. Protein Nitrogen Determination (United States)

    Nielsen, S. Suzanne

    The protein content of foods can be determined by numerous methods. The Kjeldahl method and the nitrogen combustion (Dumas) method for protein analysis are based on nitrogen determination. Both methods are official for the purposes of nutrition labeling of foods. While the Kjeldahl method has been used widely for over a hundred years, the recent availability of automated instrumentation for the Dumas method in many cases is replacing use of the Kjeldahl method.

  6. Disease specific protein corona (United States)

    Rahman, M.; Mahmoudi, M.


    It is now well accepted that upon their entrance into the biological environments, the surface of nanomaterials would be covered by various biomacromolecules (e.g., proteins and lipids). The absorption of these biomolecules, so called `protein corona', onto the surface of (nano)biomaterials confers them a new `biological identity'. Although the formation of protein coronas on the surface of nanoparticles has been widely investigated, there are few reports on the effect of various diseases on the biological identity of nanoparticles. As the type of diseases may tremendously changes the composition of the protein source (e.g., human plasma/serum), one can expect that amount and composition of associated proteins in the corona composition may be varied, in disease type manner. Here, we show that corona coated silica and polystyrene nanoparticles (after interaction with in the plasma of the healthy individuals) could induce unfolding of fibrinogen, which promotes release of the inflammatory cytokines. However, no considerable releases of inflammatory cytokines were observed for corona coated graphene sheets. In contrast, the obtained corona coated silica and polystyrene nanoparticles from the hypofibrinogenemia patients could not induce inflammatory cytokine release where graphene sheets do. Therefore, one can expect that disease-specific protein coronas can provide a novel approach for applying nanomedicine to personalized medicine, improving diagnosis and treatment of different diseases tailored to the specific conditions and circumstances.

  7. Fast protein folding kinetics (United States)

    Gelman, Hannah; Gruebele, Martin


    Fast folding proteins have been a major focus of computational and experimental study because they are accessible to both techniques: they are small and fast enough to be reasonably simulated with current computational power, but have dynamics slow enough to be observed with specially developed experimental techniques. This coupled study of fast folding proteins has provided insight into the mechanisms which allow some proteins to find their native conformation well less than 1 ms and has uncovered examples of theoretically predicted phenomena such as downhill folding. The study of fast folders also informs our understanding of even “slow” folding processes: fast folders are small, relatively simple protein domains and the principles that govern their folding also govern the folding of more complex systems. This review summarizes the major theoretical and experimental techniques used to study fast folding proteins and provides an overview of the major findings of fast folding research. Finally, we examine the themes that have emerged from studying fast folders and briefly summarize their application to protein folding in general as well as some work that is left to do. PMID:24641816

  8. The effect of protein-protein and protein-membrane interactions on membrane fouling in ultrafiltration

    NARCIS (Netherlands)

    Huisman, I.H.; Prádanos, P.; Hernández, A.


    It was studied how protein-protein and protein-membrane interactions influence the filtration performance during the ultrafiltration of protein solutions over polymeric membranes. This was done by measuring flux, streaming potential, and protein transmission during filtration of bovine serum albumin

  9. Protein: MPA1 [TP Atlas

    Lifescience Database Archive (English)

    Full Text Available MPA1 TLR signaling molecules RSAD2 CIG5 Radical S-adenosyl methionine domain-containing protein 2 Cytomegalo...virus-induced gene 5 protein, Viperin, Virus inhibitory protein, endoplasmic reticu

  10. Protein hydrolysates in sports nutrition

    Directory of Open Access Journals (Sweden)

    Manninen Anssi H


    Full Text Available Abstract It has been suggested that protein hydrolysates providing mainly di- and tripeptides are superior to intact (whole proteins and free amino acids in terms of skeletal muscle protein anabolism. This review provides a critical examination of protein hydrolysate studies conducted in healthy humans with special reference to sports nutrition. The effects of protein hydrolysate ingestion on blood amino acid levels, muscle protein anabolism, body composition, exercise performance and muscle glycogen resynthesis are discussed.


    Directory of Open Access Journals (Sweden)

    J.C.Q. Carvalho


    Full Text Available The theoretical explanation of biological concepts, associated with the use of teaching games andmodels, intensify the comprehension and increase students interest, stimulating them to participateactively on the teaching-learning process. The sta of dissemination from Centro de BiotecnologiaMolecular Estrutural (CBME, in partnership with the Centro de Divulgac~ao Cientca e Cultural(CDCC, presents, in this work, a new educational resource denoted: Protein Synthesis Game. Theapproach of the game involves the cytological aspects of protein synthesis, directed to high schoolstudents. Students are presented to day-by-day facts related to the function of a given protein in thehuman body. Such task leads players to the goal of solving out a problem through synthesizing aspecied protein. The game comprises: (1 a board illustrated with the transversal section of animalcell, with its main structures and organelles and sequences of hypothetical genes; (2 cards with thedescription of steps and other structures required for protein synthesis in eukaryotic cells; (3 piecesrepresenting nucleotides, polynucleotides, ribosome, amino acids, and polypeptide chains. In order toplay the game, students take cards that sequentially permit them to acquire the necessary pieces forproduction of the protein described in each objective. Players must move the pieces on the board andsimulate the steps of protein synthesis. The dynamic of the game allows students to easily comprehendprocesses of transcription and translation. This game was presented to dierent groups of high schoolteachers and students. Their judgments have been heard and indicated points to be improved, whichhelped us with the game development. Furthermore, the opinions colleted were always favorable forthe application of this game as a teaching resource in classrooms.

  12. Bioinformatics and moonlighting proteins

    Directory of Open Access Journals (Sweden)

    Sergio eHernández


    Full Text Available Multitasking or moonlighting is the capability of some proteins to execute two or more biochemical functions. Usually, moonlighting proteins are experimentally revealed by serendipity. For this reason, it would be helpful that Bioinformatics could predict this multifunctionality, especially because of the large amounts of sequences from genome projects. In the present work, we analyse and describe several approaches that use sequences, structures, interactomics and current bioinformatics algorithms and programs to try to overcome this problem. Among these approaches are: a remote homology searches using Psi-Blast, b detection of functional motifs and domains, c analysis of data from protein-protein interaction databases (PPIs, d match the query protein sequence to 3D databases (i.e., algorithms as PISITE, e mutation correlation analysis between amino acids by algorithms as MISTIC. Programs designed to identify functional motif/domains detect mainly the canonical function but usually fail in the detection of the moonlighting one, Pfam and ProDom being the best methods. Remote homology search by Psi-Blast combined with data from interactomics databases (PPIs have the best performance. Structural information and mutation correlation analysis can help us to map the functional sites. Mutation correlation analysis can only be used in very specific situations –it requires the existence of multialigned family protein sequences - but can suggest how the evolutionary process of second function acquisition took place. The multitasking protein database MultitaskProtDB (, previously published by our group, has been used as a benchmark for the all of the analyses.

  13. Direct protein-protein conjugation by genetically introducing bioorthogonal functional groups into proteins. (United States)

    Kim, Sanggil; Ko, Wooseok; Sung, Bong Hyun; Kim, Sun Chang; Lee, Hyun Soo


    Proteins often function as complex structures in conjunction with other proteins. Because these complex structures are essential for sophisticated functions, developing protein-protein conjugates has gained research interest. In this study, site-specific protein-protein conjugation was performed by genetically incorporating an azide-containing amino acid into one protein and a bicyclononyne (BCN)-containing amino acid into the other. Three to four sites in each of the proteins were tested for conjugation efficiency, and three combinations showed excellent conjugation efficiency. The genetic incorporation of unnatural amino acids (UAAs) is technically simple and produces the mutant protein in high yield. In addition, the conjugation reaction can be conducted by simple mixing, and does not require additional reagents or linker molecules. Therefore, this method may prove very useful for generating protein-protein conjugates and protein complexes of biochemical significance. Copyright © 2016. Published by Elsevier Ltd.

  14. Benchtop Detection of Proteins (United States)

    Scardelletti, Maximilian C.; Varaljay, Vanessa


    A process, and a benchtop-scale apparatus for implementing the process, have been developed to detect proteins associated with specific microbes in water. The process and apparatus may also be useful for detection of proteins in other, more complex liquids. There may be numerous potential applications, including monitoring lakes and streams for contamination, testing of blood and other bodily fluids in medical laboratories, and testing for microbial contamination of liquids in restaurants and industrial food-processing facilities. A sample can be prepared and analyzed by use of this process and apparatus within minutes, whereas an equivalent analysis performed by use of other processes and equipment can often take hours to days. The process begins with the conjugation of near-infrared-fluorescent dyes to antibodies that are specific to a particular protein. Initially, the research has focused on using near-infrared dyes to detect antigens or associated proteins in solution, which has proven successful vs. microbial cells, and streamlining the technique in use for surface protein detection on microbes would theoretically render similar results. However, it is noted that additional work is needed to transition protein-based techniques to microbial cell detection. Consequently, multiple such dye/antibody pairs could be prepared to enable detection of multiple selected microbial species, using a different dye for each species. When excited by near-infrared light of a suitable wavelength, each dye fluoresces at a unique longer wavelength that differs from those of the other dyes, enabling discrimination among the various species. In initial tests, the dye/antibody pairs are mixed into a solution suspected of containing the selected proteins, causing the binding of the dye/antibody pairs to such suspect proteins that may be present. The solution is then run through a microcentrifuge that includes a membrane that acts as a filter in that it retains the dye/antibody/protein

  15. Self-Assembling Protein Microarrays (United States)

    Ramachandran, Niroshan; Hainsworth, Eugenie; Bhullar, Bhupinder; Eisenstein, Samuel; Rosen, Benjamin; Lau, Albert Y.; C. Walter, Johannes; LaBaer, Joshua


    Protein microarrays provide a powerful tool for the study of protein function. However, they are not widely used, in part because of the challenges in producing proteins to spot on the arrays. We generated protein microarrays by printing complementary DNAs onto glass slides and then translating target proteins with mammalian reticulocyte lysate. Epitope tags fused to the proteins allowed them to be immobilized in situ. This obviated the need to purify proteins, avoided protein stability problems during storage, and captured sufficient protein for functional studies. We used the technology to map pairwise interactions among 29 human DNA replication initiation proteins, recapitulate the regulation of Cdt1 binding to select replication proteins, and map its geminin-binding domain.

  16. Changes in protein composition and protein phosphorylation during ...

    African Journals Online (AJOL)

    Changes in protein profiles and protein phosphorylation were studied in various stages of germinating somatic and zygotic embryos. Many proteins, which were expressed in cotyledonary stage somatic embryos, were also present in the zygotic embryos obtained from mature dry seed. The intensity of 22 kDa protein was ...

  17. Electrochemical nanomoulding through proteins (United States)

    Allred, Daniel B.

    The continued improvements in performance of modern electronic devices are directly related to the manufacturing of smaller, denser features on surfaces. Electrochemical fabrication has played a large role in continuing this trend due to its low cost and ease of scaleability toward ever smaller dimensions. This work introduces the concept of using proteins, essentially monodisperse complex polymers whose three-dimensional structures are fixed by their encoded amino acid sequences, as "moulds" around which nanostructures can be built by electrochemical fabrication. Bacterial cell-surface layer proteins, or "S-layer" proteins, from two organisms---Deinococcus radiodurans and Sporosarcina ureae---were used as the "moulds" for electrochemical fabrication. The proteins are easily purified as micron-sized sheets of periodic molecular complexes with 18-nm hexagonal and 13-nm square unit cell lattices, respectively. Direct imaging by transmission electron microscopy on ultrathin noble metal films without sample preparation eliminates potential artifacts to the high surface energy substrates necessary for high nucleation densities. Characterization involved imaging, electron diffraction, spectroscopy, and three-dimensional reconstruction. The S-layer protein of D. radiodurans was further subjected to an atomic force microscope based assay to determine the integrity of its structure and long-range order and was found to be useful for fabrication from around pH 3 to 12.

  18. Protein Denaturation in Foam. (United States)

    Clarkson; Cui; Darton


    The aim of this study was to elucidate the mechanism by which protein molecules become denatured in foam. It was found that damage to the protein is mainly due to surface denaturation at the gas-liquid interface. A fraction of the molecules adsorbed do not refold to their native state when they desorb. The degree of denaturation was found to correlate directly with the interfacial exposure, which, for mobile or partially mobile interfaces, is increased by drainage. Experiments with two different proteins showed that, under the conditions of the tests, around 10% of BSA molecules which had adsorbed at the surface remained denatured when they desorbed. For pepsin the figure was around 75%. Oxidation, which was previously thought to be a major cause of protein damage in foam, was found to be minimal. Neither do the high shear stresses in the liquid bulk encountered during bubble bursting cause denaturation, because energy is dissipated at a much greater length scale than that of the protein molecule. Copyright 1999 Academic Press.

  19. Protein (Cyanobacteria): 654346314 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  20. Protein (Cyanobacteria): 654344406 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  1. Polarizable protein packing

    KAUST Repository

    Ng, Albert H.


    To incorporate protein polarization effects within a protein combinatorial optimization framework, we decompose the polarizable force field AMOEBA into low order terms. Including terms up to the third-order provides a fair approximation to the full energy while maintaining tractability. We represent the polarizable packing problem for protein G as a hypergraph and solve for optimal rotamers with the FASTER combinatorial optimization algorithm. These approximate energy models can be improved to high accuracy [root mean square deviation (rmsd) < 1 kJ mol -1] via ridge regression. The resulting trained approximations are used to efficiently identify new, low-energy solutions. The approach is general and should allow combinatorial optimization of other many-body problems. © 2011 Wiley Periodicals, Inc. J Comput Chem, 2011 Copyright © 2011 Wiley Periodicals, Inc.

  2. Thermodynamics of Protein Aggregation (United States)

    Osborne, Kenneth L.; Barz, Bogdan; Bachmann, Michael; Strodel, Birgit

    Amyloid protein aggregation characterizes many neurodegenerative disorders, including Alzheimer's, Parkinson's, and Creutz- feldt-Jakob disease. Evidence suggests that amyloid aggregates may share similar aggregation pathways, implying simulation of full-length amyloid proteins is not necessary for understanding amyloid formation. In this study we simulate GNNQQNY, the N-terminal prion-determining domain of the yeast protein Sup35 to investigate the thermodynamics of structural transitions during aggregation. We use a coarse-grained model with replica-exchange molecular dynamics to investigate the association of 3-, 6-, and 12-chain GNNQQNY systems and we determine the aggregation pathway by studying aggregation states of GN- NQQNY. We find that the aggregation of the hydrophilic GNNQQNY sequence is mainly driven by H-bond formation, leading to the formation of /3-sheets from the very beginning of the assembly process. Condensation (aggregation) and ordering take place simultaneously, which is underpinned by the occurrence of a single heat capacity peak only.

  3. Thermal hysteresis proteins. (United States)

    Barrett, J


    Extreme environments present a wealth of biochemical adaptations. Thermal hysteresis proteins (THPs) have been found in vertebrates, invertebrates, plants, bacteria and fungi and are able to depress the freezing point of water (in the presence of ice crystals) in a non-colligative manner by binding to the surface of nascent ice crystals. The THPs comprise a disparate group of proteins with a variety of tertiary structures and often no common sequence similarities or structural motifs. Different THPs bind to different faces of the ice crystal, and no single mechanism has been proposed to account for THP ice binding affinity and specificity. Experimentally THPs have been used in the cryopreservation of tissues and cells and to induce cold tolerance in freeze susceptible organisms. THPs represent a remarkable example of parallel and convergent evolution with different proteins being adapted for an anti-freeze role.

  4. Accessory Proteins at ERES

    DEFF Research Database (Denmark)

    Klinkenberg, Rafael David

    proteins. Together these components co‐operate in cargo‐selection as well as forming, loading and releasing budding vesicles from specific regions on the membrane surface of the ER. Coat components furthermore convey vesicle targeting towards the Golgi. However, not much is known about the mechanisms...... that regulate the COPII assembly at the vesicle bud site. This thesis provides the first regulatory mechanism of COPII assembly in relation to ER‐membrane lipid‐signal recognition by the accessory protein p125A (Sec23IP). The aim of the project was to characterize p125A function by dissecting two main domains...... in the protein; a putative lipid‐associating domain termed the DDHD domain that is defined by the four amino acid motif that gives the domain its name; and a ubiquitously found domain termed Sterile α‐motif (SAM), which is mostly associated with oligomerization and polymerization. We first show, that the DDHD...

  5. Matricellular proteins and biomaterials. (United States)

    Morris, Aaron H; Kyriakides, Themis R


    Biomaterials are essential to modern medicine as components of reconstructive implants, implantable sensors, and vehicles for localized drug delivery. Advances in biomaterials have led to progression from simply making implants that are nontoxic to making implants that are specifically designed to elicit particular functions within the host. The interaction of implants and the extracellular matrix during the foreign body response is a growing area of concern for the field of biomaterials, because it can lead to implant failure. Expression of matricellular proteins is modulated during the foreign body response and these proteins interact with biomaterials. The design of biomaterials to specifically alter the levels of matricellular proteins surrounding implants provides a new avenue for the design and fabrication of biomimetic biomaterials. Copyright © 2014 International Society of Matrix Biology. Published by Elsevier B.V. All rights reserved.

  6. Trisulfides in Proteins

    DEFF Research Database (Denmark)

    Nielsen, Rasmus W.; Tachibana, Christine; Hansen, Niels Erik


    Trisulfides and other oligosulfides are widely distributed in the biological world. In plants, e.g., garlic, trisulfides are associated with potentially beneficial properties. However, an extra neutral sulfur atom covalently bound between the two sulfur atoms of a pair of cysteines is not a commo...... post-translational modification, and the number of proteins in which a trisulfide has been unambiguously identified is small. Nevertheless, we believe that its prevalence may be underestimated, particularly with the increasing evidence for significant pools of sulfides in living tissues...... and their possible roles in cellular metabolism. This review focuses on examples of proteins that are known to contain a trisulfide bridge, and gives an overview of the chemistry of trisulfide formation, and the methods by which it is detected in proteins....

  7. Epistasis in protein evolution (United States)

    Starr, Tyler N.


    Abstract The structure, function, and evolution of proteins depend on physical and genetic interactions among amino acids. Recent studies have used new strategies to explore the prevalence, biochemical mechanisms, and evolutionary implications of these interactions—called epistasis—within proteins. Here we describe an emerging picture of pervasive epistasis in which the physical and biological effects of mutations change over the course of evolution in a lineage‐specific fashion. Epistasis can restrict the trajectories available to an evolving protein or open new paths to sequences and functions that would otherwise have been inaccessible. We describe two broad classes of epistatic interactions, which arise from different physical mechanisms and have different effects on evolutionary processes. Specific epistasis—in which one mutation influences the phenotypic effect of few other mutations—is caused by direct and indirect physical interactions between mutations, which nonadditively change the protein's physical properties, such as conformation, stability, or affinity for ligands. In contrast, nonspecific epistasis describes mutations that modify the effect of many others; these typically behave additively with respect to the physical properties of a protein but exhibit epistasis because of a nonlinear relationship between the physical properties and their biological effects, such as function or fitness. Both types of interaction are rampant, but specific epistasis has stronger effects on the rate and outcomes of evolution, because it imposes stricter constraints and modulates evolutionary potential more dramatically; it therefore makes evolution more contingent on low‐probability historical events and leaves stronger marks on the sequences, structures, and functions of protein families. PMID:26833806

  8. Protein biosynthesis in mitochondria. (United States)

    Kuzmenko, A V; Levitskii, S A; Vinogradova, E N; Atkinson, G C; Hauryliuk, V; Zenkin, N; Kamenski, P A


    Translation, that is biosynthesis of polypeptides in accordance with information encoded in the genome, is one of the most important processes in the living cell, and it has been in the spotlight of international research for many years. The mechanisms of protein biosynthesis in bacteria and in the eukaryotic cytoplasm are now understood in great detail. However, significantly less is known about translation in eukaryotic mitochondria, which is characterized by a number of unusual features. In this review, we summarize current knowledge about mitochondrial translation in different organisms while paying special attention to the aspects of this process that differ from cytoplasmic protein biosynthesis.

  9. Water-transporting proteins

    DEFF Research Database (Denmark)

    Zeuthen, Thomas


    Transport through lipids and aquaporins is osmotic and entirely driven by the difference in osmotic pressure. Water transport in cotransporters and uniporters is different: Water can be cotransported, energized by coupling to the substrate flux by a mechanism closely associated with protein...... is not clear. It is associated with the substrate movements in aqueous pathways within the protein; a conventional unstirred layer mechanism can be ruled out, due to high rates of diffusion in the cytoplasm. The physiological roles of the various modes of water transport are reviewed in relation to epithelial...

  10. Cold gelation of globular proteins

    NARCIS (Netherlands)

    Alting, A.C.


    Keywords : globular proteins, whey protein, ovalbumin, cold gelation, disulfide bonds, texture, gel hardnessProtein gelation in food products is important to obtain desirable sensory and textural properties. Cold gelation is a novel method to produce protein-based gels. It is a two step process in

  11. The Formation of Protein Structure

    DEFF Research Database (Denmark)

    Bohr, Jakob; Bohr, Henrik; Brunak, Søren


    Dynamically induced curvature owing to long-range excitations along the backbones of protein molecules with non-linear elastic properties may control the folding of proteins.......Dynamically induced curvature owing to long-range excitations along the backbones of protein molecules with non-linear elastic properties may control the folding of proteins....

  12. A simple dependence between protein evolution rate and the number of protein-protein interactions

    Directory of Open Access Journals (Sweden)

    Hirsh Aaron E


    Full Text Available Abstract Background It has been shown for an evolutionarily distant genomic comparison that the number of protein-protein interactions a protein has correlates negatively with their rates of evolution. However, the generality of this observation has recently been challenged. Here we examine the problem using protein-protein interaction data from the yeast Saccharomyces cerevisiae and genome sequences from two other yeast species. Results In contrast to a previous study that used an incomplete set of protein-protein interactions, we observed a highly significant correlation between number of interactions and evolutionary distance to either Candida albicans or Schizosaccharomyces pombe. This study differs from the previous one in that it includes all known protein interactions from S. cerevisiae, and a larger set of protein evolutionary rates. In both evolutionary comparisons, a simple monotonic relationship was found across the entire range of the number of protein-protein interactions. In agreement with our earlier findings, this relationship cannot be explained by the fact that proteins with many interactions tend to be important to yeast. The generality of these correlations in other kingdoms of life unfortunately cannot be addressed at this time, due to the incompleteness of protein-protein interaction data from organisms other than S. cerevisiae. Conclusions Protein-protein interactions tend to slow the rate at which proteins evolve. This may be due to structural constraints that must be met to maintain interactions, but more work is needed to definitively establish the mechanism(s behind the correlations we have observed.

  13. Modelling of proteins in membranes

    DEFF Research Database (Denmark)

    Sperotto, Maria Maddalena; May, S.; Baumgaertner, A.


    This review describes some recent theories and simulations of mesoscopic and microscopic models of lipid membranes with embedded or attached proteins. We summarize results supporting our understanding of phenomena for which the activities of proteins in membranes are expected to be significantly...... affected by the lipid environment. Theoretical predictions are pointed out, and compared to experimental findings, if available. Among others, the following phenomena are discussed: interactions of interfacially adsorbed peptides, pore-forming amphipathic peptides, adsorption of charged proteins onto...... oppositely charged lipid membranes, lipid-induced tilting of proteins embedded in lipid bilayers, protein-induced bilayer deformations, protein insertion and assembly, and lipid-controlled functioning of membrane proteins....

  14. Protein degradation systems in platelets. (United States)

    Kraemer, B F; Weyrich, A S; Lindemann, S


    Protein synthesis and degradation are essential processes that allow cells to survive and adapt to their surrounding milieu. In nucleated cells, the degradation and/or cleavage of proteins is required to eliminate aberrant proteins. Cells also degrade proteins as a mechanism for cell signalling and complex cellular functions. Although the last decade has convincingly shown that platelets synthesise proteins, the roles of protein degradation in these anucleate cytoplasts are less clear. Here we review what is known about protein degradation in platelets placing particular emphasis on the proteasome and the cysteine protease calpain.

  15. Truly Absorbed Microbial Protein Synthesis, Rumen Bypass Protein, Endogenous Protein, and Total Metabolizable Protein from Starchy and Protein-Rich Raw Materials

    NARCIS (Netherlands)

    Parand, Ehsan; Vakili, Alireza; Mesgaran, Mohsen Danesh; Duinkerken, Van Gert; Yu, Peiqiang


    This study was carried out to measure truly absorbed microbial protein synthesis, rumen bypass protein, and endogenous protein loss, as well as total metabolizable protein, from starchy and protein-rich raw feed materials with model comparisons. Predictions by the DVE2010 system as a more

  16. Protein requirements of Penaeid shrimp.


    Kanazawa, A


    Proteins are indispensable nutrients for growth and maintenance of live of all animals. The optimum protein levels in diets for shrimps are different among the various species. Squid meal is an effective protein source for many penaeids. The effects of dietary protein, lipid, and carbohydrate levels on the growth and survival of larvae of Penaeus japonicus were examined by feeding trials using purified diet with carrageenan as a binder. As a result, the effects of protein levels on growth and...

  17. Protein oxidation and ageing

    DEFF Research Database (Denmark)

    Linton, S; Davies, Michael Jonathan; Dean, R T


    of redox-active metal ions that could catalyse oxidant formation. As a result of this decrease in antioxidant defences, and increased rate of ROS formation, it is possible that the impact of ROS increases with age. ROS are known to oxidise biological macromolecules, with proteins an important target...

  18. Thermodynamics of meat proteins

    NARCIS (Netherlands)

    Sman, van der R.G.M.


    We describe the water activity of meat, being a mixture of proteins, salts and water, by the Free-Volume-Flory–Huggins (FVFH) theory augmented with the equation. Earlier, the FVFH theory is successfully applied to describe the thermodynamics to glucose homopolymers like starch, dextrans and

  19. Protein digestion in ruminants

    African Journals Online (AJOL)

    Animal Nutrition, Animal and Dairy Science Research Institute, Irene, 1675Republic of South Africa. Although the protein requirement of domestic ruminants may be calculated from a simple one-compartment model, this approach ignores factors such as microbial fermentation in the rumen and the non-equality of feed.

  20. Protein Sorting Prediction

    DEFF Research Database (Denmark)

    Nielsen, Henrik


    Many computational methods are available for predicting protein sorting in bacteria. When comparing them, it is important to know that they can be grouped into three fundamentally different approaches: signal-based, global-property-based and homology-based prediction. In this chapter, the strengt...

  1. Allosteric Regulation of Proteins

    Indian Academy of Sciences (India)

    ... Lecture Workshops · Refresher Courses · Symposia · Live Streaming. Home; Journals; Resonance – Journal of Science Education; Volume 22; Issue 1. Allosteric Regulation of Proteins: A Historical Perspective on the Development of Concepts and Techniques. General Article Volume 22 Issue 1 January 2017 pp 37-50 ...

  2. Markers of protein oxidation

    DEFF Research Database (Denmark)

    Headlam, Henrietta A; Davies, Michael Jonathan


    Exposure of proteins to radicals in the presence of O2 gives both side-chain oxidation and backbone fragmentation. These processes can be interrelated, with initial side-chain oxidation giving rise to backbone damage via transfer reactions. We have shown previously that alkoxyl radicals formed on...

  3. Protein digestion in ruminants

    African Journals Online (AJOL)

    acids absorbed into the circulation of the animal. Ideally, therefore, the biological value of a feed protein should be determined from the amount and type of amino acid appearing in the portal circulation of the animal, and not simplythe dissappearance of amino acids from the tract. Ruminant digestion may be more easily ...

  4. Antifreeze Proteins of Bacteria

    Indian Academy of Sciences (India)

    Home; Journals; Resonance – Journal of Science Education; Volume 12; Issue 12. Antifreeze Proteins of Bacteria. M K Chattopadhyay. General Article Volume 12 Issue 12 December 2007 pp 25-30. Fulltext. Click here to view fulltext PDF. Permanent link: Keywords.

  5. NMR of unfolded proteins

    Indian Academy of Sciences (India)



    Jan 3, 2005 ... deposition of data and advanced search on the pattern of PDB.12. Detailed characterization of the unfolded state and consequent identification of the folding initiation sites in a given protein provide valuable insight into its folding mechanism.18 Well-formed or transient residual structures in the unfolded ...

  6. Protein Requirements during Aging

    Directory of Open Access Journals (Sweden)

    Glenda Courtney-Martin


    Full Text Available Protein recommendations for elderly, both men and women, are based on nitrogen balance studies. They are set at 0.66 and 0.8 g/kg/day as the estimated average requirement (EAR and recommended dietary allowance (RDA, respectively, similar to young adults. This recommendation is based on single linear regression of available nitrogen balance data obtained at test protein intakes close to or below zero balance. Using the indicator amino acid oxidation (IAAO method, we estimated the protein requirement in young adults and in both elderly men and women to be 0.9 and 1.2 g/kg/day as the EAR and RDA, respectively. This suggests that there is no difference in requirement on a gender basis or on a per kg body weight basis between younger and older adults. The requirement estimates however are ~40% higher than the current protein recommendations on a body weight basis. They are also 40% higher than our estimates in young men when calculated on the basis of fat free mass. Thus, current recommendations may need to be re-assessed. Potential rationale for this difference includes a decreased sensitivity to dietary amino acids and increased insulin resistance in the elderly compared with younger individuals.

  7. Protein: CAD [Trypanosomes Database

    Lifescience Database Archive (English)

    Full Text Available CAD carbamoyl-phosphate synthetase 2, aspartate transcarbamylase, and dihydroorotaseCAD... trifunctional proteincarbamoylphosphate synthetase 2/aspartate transcarbamylase/dihydroorotasemultifunctional protein CAD... H.sapiens 47458828 18105007 790 P27708 CAD_(gene)|| 114010 2p22-p21 hsa00250|hsa00240 ...

  8. Conditional function of autoaggregative protien cah and common cah mutations in Shiga toxin-producing Escherichia coli (United States)

    Cah is a calcium-binding autotransporter protein involved in autoaggregation and biofilm formation. Although cah is widespread in Shiga toxin-producing Escherichia coli (STEC), we detected mutations in cah at a frequency of 31.3% in this pathogen. In STEC O157:H7 super-shedder strain SS17, a large d...

  9. The characterization of a novel S100A1 binding site in the N-terminus of TRPM1

    Czech Academy of Sciences Publication Activity Database

    Jirků, M.; Lánský, Z.; Bednárová, Lucie; Šulc, M.; Monincová, Lenka; Majer, Pavel; Vyklický, L.; Vondrášek, Jiří; Teisinger, J.; Boušová, Kristýna


    Roč. 78, Sep (2016), s. 186-193 ISSN 1357-2725 Institutional support: RVO:61388963 Keywords : TRPM1 channel * binding site * calcium-binding protein S100A1 * steady-state fluorescence anisotropy * molecular modeling * circular dichroism Subject RIV: CE - Biochemistry Impact factor: 3.505, year: 2016

  10. Interaction of an S100A9 gene variant with saturated fat and carbohydrates to modulate insulin resistance in 3 populations of different ancestries (United States)

    BACKGROUND: S100 calcium binding protein A9 (S100A9) has previously been identified as a type 2 diabetes (T2D) gene. However, this finding requires independent validation and more in depth analyses in other populations and ancestries. OBJECTIVES: We aimed to replicate the associations between an S10...

  11. Measuring protein breakdown in individual proteins in vivo

    DEFF Research Database (Denmark)

    Holm, Lars; Kjær, Michael


    be used to determine the breakdown rate of specific proteins and, therefore, do not keep up to the preceding methodological demands in physiological research. A newly developed approach to determine the fractional breakdown rate of single proteins seems promising. Its conceptual advantage......PURPOSE OF REVIEW: To outline different approaches of how protein breakdown can be quantified and to present a new approach to determine the fractional breakdown rate of individual slow turnover proteins in vivo. RECENT FINDINGS: None of the available methods for determining protein breakdown can...... is that the proteins of interest are the site of measurement. Hence, the application initially demands the proteins to be labeled with stable isotopically labeled amino acids. Subsequently, the loss of label from the proteins will be dependent on the protein breakdown rate when no labeled amino acids...

  12. Interaction between plate make and protein in protein crystallisation screening.

    Directory of Open Access Journals (Sweden)

    Gordon J King

    Full Text Available BACKGROUND: Protein crystallisation screening involves the parallel testing of large numbers of candidate conditions with the aim of identifying conditions suitable as a starting point for the production of diffraction quality crystals. Generally, condition screening is performed in 96-well plates. While previous studies have examined the effects of protein construct, protein purity, or crystallisation condition ingredients on protein crystallisation, few have examined the effect of the crystallisation plate. METHODOLOGY/PRINCIPAL FINDINGS: We performed a statistically rigorous examination of protein crystallisation, and evaluated interactions between crystallisation success and plate row/column, different plates of same make, different plate makes and different proteins. From our analysis of protein crystallisation, we found a significant interaction between plate make and the specific protein being crystallised. CONCLUSIONS/SIGNIFICANCE: Protein crystal structure determination is the principal method for determining protein structure but is limited by the need to produce crystals of the protein under study. Many important proteins are difficult to crystallize, so that identification of factors that assist crystallisation could open up the structure determination of these more challenging targets. Our findings suggest that protein crystallisation success may be improved by matching a protein with its optimal plate make.

  13. Minireview: protein arginine methylation of nonhistone proteins in transcriptional regulation. (United States)

    Lee, Young-Ho; Stallcup, Michael R


    Endocrine regulation frequently culminates in altered transcription of specific genes. The signal transduction pathways, which transmit the endocrine signal from cell surface to the transcription machinery, often involve posttranslational modifications of proteins. Although phosphorylation has been by far the most widely studied protein modification, recent studies have indicated important roles for other types of modification, including protein arginine methylation. Ten different protein arginine methyltransferase (PRMT) family members have been identified in mammalian cells, and numerous substrates are being identified for these PRMTs. Whereas major attention has been focused on the methylation of histones and its role in chromatin remodeling and transcriptional regulation, there are many nonhistone substrates methylated by PRMTs. This review primarily focuses on recent progress on the roles of the nonhistone protein methylation in transcription. Protein methylation of coactivators, transcription factors, and signal transducers, among other proteins, plays important roles in transcriptional regulation. Protein methylation may affect protein-protein interaction, protein-DNA or protein-RNA interaction, protein stability, subcellular localization, or enzymatic activity. Thus, protein arginine methylation is critical for regulation of transcription and potentially for various physiological/pathological processes.

  14. HIV protein sequence hotspots for crosstalk with host hub proteins.

    Directory of Open Access Journals (Sweden)

    Mahdi Sarmady

    Full Text Available HIV proteins target host hub proteins for transient binding interactions. The presence of viral proteins in the infected cell results in out-competition of host proteins in their interaction with hub proteins, drastically affecting cell physiology. Functional genomics and interactome datasets can be used to quantify the sequence hotspots on the HIV proteome mediating interactions with host hub proteins. In this study, we used the HIV and human interactome databases to identify HIV targeted host hub proteins and their host binding partners (H2. We developed a high throughput computational procedure utilizing motif discovery algorithms on sets of protein sequences, including sequences of HIV and H2 proteins. We identified as HIV sequence hotspots those linear motifs that are highly conserved on HIV sequences and at the same time have a statistically enriched presence on the sequences of H2 proteins. The HIV protein motifs discovered in this study are expressed by subsets of H2 host proteins potentially outcompeted by HIV proteins. A large subset of these motifs is involved in cleavage, nuclear localization, phosphorylation, and transcription factor binding events. Many such motifs are clustered on an HIV sequence in the form of hotspots. The sequential positions of these hotspots are consistent with the curated literature on phenotype altering residue mutations, as well as with existing binding site data. The hotspot map produced in this study is the first global portrayal of HIV motifs involved in altering the host protein network at highly connected hub nodes.

  15. Fragments of protein A eluted during protein A affinity chromatography. (United States)

    Carter-Franklin, Jayme N; Victa, Corazon; McDonald, Paul; Fahrner, Robert


    Protein A affinity chromatography is a common method for process scale purification of monoclonal antibodies. During protein A affinity chromatography, protein A ligand co-elutes with the antibody (commonly called leaching), which is a potential disadvantage since the leached protein A may need to be cleared for pharmaceutical antibodies. To determine the mechanism of protein A leaching and characterize the leached protein A, we fluorescently labeled the protein A ligand in situ on protein A affinity chromatography media. We found that intact protein A leaches when loading either purified antibody or unpurified antibody in harvested cell culture fluid (HCCF), and that additionally fragments of protein A leach when loading HCCF. The leaching of protein A fragments can be reduced by EDTA, suggesting that proteinases contribute to the generation of protein A fragments. We found that protein A fragments larger than about 6000 Da can be measured by enzyme linked immunosorbent assay, and that they can be more difficult to clear than whole protein A by cation-exchange chromatography.

  16. Inferring protein function by domain context similarities in protein-protein interaction networks

    Directory of Open Access Journals (Sweden)

    Sun Zhirong


    Full Text Available Abstract Background Genome sequencing projects generate massive amounts of sequence data but there are still many proteins whose functions remain unknown. The availability of large scale protein-protein interaction data sets makes it possible to develop new function prediction methods based on protein-protein interaction (PPI networks. Although several existing methods combine multiple information resources, there is no study that integrates protein domain information and PPI networks to predict protein functions. Results The domain context similarity can be a useful index to predict protein function similarity. The prediction accuracy of our method in yeast is between 63%-67%, which outperforms the other methods in terms of ROC curves. Conclusion This paper presents a novel protein function prediction method that combines protein domain composition information and PPI networks. Performance evaluations show that this method outperforms existing methods.

  17. High quality protein microarray using in situ protein purification

    Directory of Open Access Journals (Sweden)

    Fleischmann Robert D


    Full Text Available Abstract Background In the postgenomic era, high throughput protein expression and protein microarray technologies have progressed markedly permitting screening of therapeutic reagents and discovery of novel protein functions. Hexa-histidine is one of the most commonly used fusion tags for protein expression due to its small size and convenient purification via immobilized metal ion affinity chromatography (IMAC. This purification process has been adapted to the protein microarray format, but the quality of in situ His-tagged protein purification on slides has not been systematically evaluated. We established methods to determine the level of purification of such proteins on metal chelate-modified slide surfaces. Optimized in situ purification of His-tagged recombinant proteins has the potential to become the new gold standard for cost-effective generation of high-quality and high-density protein microarrays. Results Two slide surfaces were examined, chelated Cu2+ slides suspended on a polyethylene glycol (PEG coating and chelated Ni2+ slides immobilized on a support without PEG coating. Using PEG-coated chelated Cu2+ slides, consistently higher purities of recombinant proteins were measured. An optimized wash buffer (PBST composed of 10 mM phosphate buffer, 2.7 mM KCl, 140 mM NaCl and 0.05% Tween 20, pH 7.4, further improved protein purity levels. Using Escherichia coli cell lysates expressing 90 recombinant Streptococcus pneumoniae proteins, 73 proteins were successfully immobilized, and 66 proteins were in situ purified with greater than 90% purity. We identified several antigens among the in situ-purified proteins via assays with anti-S. pneumoniae rabbit antibodies and a human patient antiserum, as a demonstration project of large scale microarray-based immunoproteomics profiling. The methodology is compatible with higher throughput formats of in vivo protein expression, eliminates the need for resin-based purification and circumvents

  18. Metabolism of minor isoforms of prion proteins: Cytosolic prion protein and transmembrane prion protein


    Song, Zhiqi; Zhao, Deming; Yang, Lifeng


    Transmissible spongiform encephalopathy or prion disease is triggered by the conversion from cellular prion protein to pathogenic prion protein. Growing evidence has concentrated on prion protein configuration changes and their correlation with prion disease transmissibility and pathogenicity. In vivo and in vitro studies have shown that several cytosolic forms of prion protein with specific topological structure can destroy intracellular stability and contribute to prion protein pathogenicit...

  19. Shape complementarity and hydrogen bond preferences in protein-protein interfaces: implications for antibody modeling and protein-protein docking. (United States)

    Kuroda, Daisuke; Gray, Jeffrey J


    Characterizing protein-protein interfaces and the hydrogen bonds is a first step to better understand proteins' structures and functions toward high-resolution protein design. However, there are few large-scale surveys of hydrogen bonds of interfaces. In addition, previous work of shape complementarity of protein complexes suggested that lower shape complementarity in antibody-antigen interfaces is related to their evolutionary origin. Using 6637 non-redundant protein-protein interfaces, we revealed peculiar features of various protein complex types. In contrast to previous findings, the shape complementarity of antibody-antigen interfaces resembles that of the other interface types. These results highlight the importance of hydrogen bonds during evolution of protein interfaces and rectify the prevailing belief that antibodies have lower shape complementarity. Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail:

  20. Dairy Proteins and Energy Balance

    DEFF Research Database (Denmark)

    Bendtsen, Line Quist

    High protein diets affect energy balance beneficially through decreased hunger, enhanced satiety and increased energy expenditure. Dairy products are a major source of protein. Dairy proteins are comprised of two classes, casein (80%) and whey proteins (20%), which are both of high quality......, but casein is absorbed slowly and whey is absorbed rapidly. The present PhD study investigated the effects of total dairy proteins, whey, and casein, on energy balance and the mechanisms behind any differences in the effects of the specific proteins. The results do not support the hypothesis that dairy...... proteins, whey or casein are more beneficial than other protein sources in the regulation of energy balance, and suggest that dairy proteins, whey or casein seem to play only a minor role, if any, in the prevention and treatment of obesity....

  1. Discovering Protein-Protein Interactions Using Nucleic Acid Programmable Protein Arrays. (United States)

    Tang, Yanyang; Qiu, Ji; Machner, Matthias; LaBaer, Joshua


    We have developed a protocol enabling the study of protein-protein interactions (PPIs) at the proteome level using in vitro-synthesized proteins. Assay preparation requires molecular cloning of the query gene into a vector that supports in vitro transcription/translation (IVTT) and appends a HaloTag to the query protein of interest. In parallel, protein microarrays are prepared by printing plasmids encoding glutathione S-transferase (GST)-tagged target proteins onto a carrier matrix/glass slide coated with antibody directed against GST. At the time of the experiment, the query protein and the target protein are produced separately through IVTT. The query protein is then applied to nucleic acid programmable protein arrays (NAPPA) that display thousands of freshly produced target proteins captured by anti-GST antibody. Interactions between the query and immobilized target proteins are detected through addition of a fluorophore-labeled HaloTag ligand. Our protocol allows the elucidation of PPIs in a high-throughput fashion using proteins produced in vitro, obviating the scientific challenges, high cost, and laborious work, as well as concerns about protein stability, which are usually present in protocols using conventional protein arrays. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley & Sons, Inc.

  2. Circular dichroism spectroscopy of fluorescent proteins

    NARCIS (Netherlands)

    Visser, N.V.; Hink, M.A.; Borst, J.W.; Krogt, van der G.N.M.; Visser, A.J.W.G.


    Circular dichroism (CD) spectra have been obtained from several variants of green fluorescent protein: blue fluorescent protein (BFP), enhanced cyan fluorescent protein (CFP), enhanced green fluorescent protein (GFP), enhanced yellow fluorescent protein (YFP), all from Aequorea victoria, and the red

  3. Competitive Protein Adsorption - Multilayer Adsorption and Surface Induced Protein Aggregation

    DEFF Research Database (Denmark)

    Holmberg, Maria; Hou, Xiaolin


    In this study, competitive adsorption of albumin and IgG (immunoglobulin G) from human serum solutions and protein mixtures onto polymer surfaces is studied by means of radioactive labeling. By using two different radiolabels (125I and 131I), albumin and IgG adsorption to polymer surfaces...... is monitored simultaneously and the influence from the presence of other human serum proteins on albumin and IgG adsorption, as well as their mutual influence during adsorption processes, is investigated. Exploring protein adsorption by combining analysis of competitive adsorption from complex solutions...... of high concentration with investigation of single protein adsorption and interdependent adsorption between two specific proteins enables us to map protein adsorption sequences during competitive protein adsorption. Our study shows that proteins can adsorb in a multilayer fashion onto the polymer surfaces...

  4. Protein Functionalized Nanodiamond Arrays

    Directory of Open Access Journals (Sweden)

    Liu YL


    Full Text Available Abstract Various nanoscale elements are currently being explored for bio-applications, such as in bio-images, bio-detection, and bio-sensors. Among them, nanodiamonds possess remarkable features such as low bio-cytotoxicity, good optical property in fluorescent and Raman spectra, and good photostability for bio-applications. In this work, we devise techniques to position functionalized nanodiamonds on self-assembled monolayer (SAMs arrays adsorbed on silicon and ITO substrates surface using electron beam lithography techniques. The nanodiamond arrays were functionalized with lysozyme to target a certain biomolecule or protein specifically. The optical properties of the nanodiamond-protein complex arrays were characterized by a high throughput confocal microscope. The synthesized nanodiamond-lysozyme complex arrays were found to still retain their functionality in interacting with E. coli.

  5. Problems in Protein Biosynthesis (United States)

    Lengyel, Peter


    Outline of the steps in protein synthesis. Nature of the genetic code. The use of synthetic oligo- and polynucleotides in deciphering the code. Structure of the code: relatedness of synonym codons. The wobble hypothesis. Chain initiation and N-formyl-methionine. Chain termination and nonsense codons. Mistakes in translation: ambiguity in vitro. Suppressor mutations resulting in ambiguity. Limitations in the universality of the code. Attempts to determine the particular codons used by a species. Mechanisms of suppression, caused by (a) abnormal aminoacyl-tRNA, (b) ribosomal malfunction. Effect of streptomycin. The problem of "reading" a nucleic acid template. Different ribosomal mutants and DNA polymerase mutants might cause different mistakes. The possibility of involvement of allosteric proteins in template reading. PMID:5338560

  6. Accessory Proteins at ERES

    DEFF Research Database (Denmark)

    Klinkenberg, Rafael David

    distribution of mSec16B. We further dissect both mSec16A and mSec16B, and show that the region in human mSec16B encompassing residues 35‐194 and the region in human mSec16A comprising residues 1096‐1190 maintain membrane binding irrespective of the removal of membrane associating proteins by salt wash...... or proteolytic digestion. However, neither mSec16B (35‐194) nor mSec16A (1096‐1190) maintain ERES targeting. These findings support previous observations of the need for the membrane binding regions to be expressed in cis with a Central Conserved Domain (CCD) in both proteins to convey ERES targeting....

  7. Porcine prion protein amyloid


    Hammarstr?m, Per; Nystr?m, Sofie


    ABSTRACT Mammalian prions are composed of misfolded aggregated prion protein (PrP) with amyloid-like features. Prions are zoonotic disease agents that infect a wide variety of mammalian species including humans. Mammals and by-products thereof which are frequently encountered in daily life are most important for human health. It is established that bovine prions (BSE) can infect humans while there is no such evidence for any other prion susceptible species in the human food chain (sheep, goat...

  8. Engineering ancestral protein hyperstability. (United States)

    Romero-Romero, M Luisa; Risso, Valeria A; Martinez-Rodriguez, Sergio; Ibarra-Molero, Beatriz; Sanchez-Ruiz, Jose M


    Many experimental analyses and proposed scenarios support that ancient life was thermophilic. In congruence with this hypothesis, proteins encoded by reconstructed sequences corresponding to ancient phylogenetic nodes often display very high stability. Here, we show that such 'reconstructed ancestral hyperstability' can be further engineered on the basis of a straightforward approach that uses exclusively information afforded by the ancestral reconstruction process itself. Since evolution does not imply continuous progression, screening of the mutations between two evolutionarily related resurrected ancestral proteins may identify mutations that further stabilize the most stable one. To explore this approach, we have used a resurrected thioredoxin corresponding to the last common ancestor of the cyanobacterial, Deinococcus and Thermus groups (LPBCA thioredoxin), which has a denaturation temperature of ∼123°C. This high value is within the top 0.1% of the denaturation temperatures in the ProTherm database and, therefore, achieving further stabilization appears a priori as a challenging task. Nevertheless, experimental comparison with a resurrected thioredoxin corresponding to the last common ancestor of bacteria (denaturation temperature of ∼115°C) immediately identifies three mutations that increase the denaturation temperature of LPBCA thioredoxin to ∼128°C. Comparison between evolutionarily related resurrected ancestral proteins thus emerges as a simple approach to expand the capability of ancestral reconstruction to search sequence space for extreme protein properties of biotechnological interest. The fact that ancestral sequences for many phylogenetic nodes can be derived from a single alignment of modern sequences should contribute to the general applicability of this approach. © 2016 The Author(s); published by Portland Press Limited on behalf of the Biochemical Society.

  9. Immunoprecipitation-based analysis of protein-protein interactions. (United States)

    Speth, Corinna; Toledo-Filho, Luis A A; Laubinger, Sascha


    Several techniques allow the detection of protein-protein interactions. In vivo co-immunoprecipitation (Co-IP) studies are an important complement to other commonly used techniques such as yeast two-hybrid or fluorescence complementation, as they reveal interactions between functional proteins at physiological relevant concentrations. Here, we describe an in vivo Co-IP approach using either GFP affinity matrix or specific antibodies to purify proteins of interests and their interacting partners.

  10. Neutron protein crystallography

    Energy Technology Data Exchange (ETDEWEB)

    Niimura, Nobuo [Japan Atomic Energy Research Inst., Tokai, Ibaraki (Japan). Tokai Research Establishment


    X-ray diffraction of single crystal has enriched the knowledge of various biological molecules such as proteins, DNA, t-RNA, viruses, etc. It is difficult to make structural analysis of hydrogen atoms in a protein using X-ray crystallography, whereas neutron diffraction seems usable to directly determine the location of those hydrogen atoms. Here, neutron diffraction method was applied to structural analysis of hen egg-white lysozyme. Since the crystal size of a protein to analyze is generally small (5 mm{sup 3} at most), the neutron beam at the sample position in monochromator system was set to less than 5 x 5 mm{sup 2} and beam divergence to 0.4 degree or less. Neutron imaging plate with {sup 6}Li or Gd mixed with photostimulated luminescence material was used and about 2500 Bragg reflections were recorded in one crystal setting. A total of 38278 reflections for 2.0 A resolution were collected in less than 10 days. Thus, stereo views of Trp-111 omit map around the indol ring of Trp-111 was presented and the three-dimensional arrangement of 696H and 264D atoms in the lysozyme molecules was determined using the omit map. (M.N.)

  11. Biophysical characterization of Ca2+-binding of S100A5 and Ca2+-induced interaction with RAGE. (United States)

    Kim, Iktae; Lee, Ko On; Yun, Young-Joo; Jeong, Jea Yeon; Kim, Eun-Hee; Cheong, Haekap; Ryu, Kyoung-Seok; Kim, Nak-Kyoon; Suh, Jeong-Yong


    S100A5 is a calcium-binding protein of S100 family, which represents a major ligand to the receptor for advanced glycation end product (RAGE), a pattern recognition receptor engaged in diverse pathological processes. Here we have characterized calcium binding of S100A5 and the complex formation between S100A5 and RAGE using calorimetry and NMR spectroscopy. S100A5 binds to calcium ions in a sequential manner with the equilibrium dissociation constants (K D ) of 1.3 μM and 3.5 μM, which corresponds to the calcium-binding at the C-terminal and N-terminal EF-hands. Upon calcium binding, S100A5 interacts with the V domain of RAGE (RAGE-v) to form a heterotrimer (K D ∼5.9 μM) that is distinct among the S100 family proteins. Chemical shift perturbation data from NMR titration experiments indicates that S100A5 employs the periphery of the dimer interface to interact with RAGE-v. Distinct binding mode and stoichiometry of RAGE against different S100 family proteins could be important to modulate diverse RAGE signaling. Copyright © 2016 Elsevier Inc. All rights reserved.

  12. Metabolism of minor isoforms of prion proteins: Cytosolic prion protein and transmembrane prion protein (United States)

    Song, Zhiqi; Zhao, Deming; Yang, Lifeng


    Transmissible spongiform encephalopathy or prion disease is triggered by the conversion from cellular prion protein to pathogenic prion protein. Growing evidence has concentrated on prion protein configuration changes and their correlation with prion disease transmissibility and pathogenicity. In vivo and in vitro studies have shown that several cytosolic forms of prion protein with specific topological structure can destroy intracellular stability and contribute to prion protein pathogenicity. In this study, the latest molecular chaperone system associated with endoplasmic reticulum-associated protein degradation, the endoplasmic reticulum resident protein quality-control system and the ubiquitination proteasome system, is outlined. The molecular chaperone system directly correlates with the prion protein degradation pathway. Understanding the molecular mechanisms will help provide a fascinating avenue for further investigations on prion disease treatment and prion protein-induced neurodegenerative diseases. PMID:25206608

  13. Understanding Protein Non-Folding (United States)

    Uversky, Vladimir N.; Dunker, A. Keith


    This review describes the family of intrinsically disordered proteins, members of which fail to form rigid 3-D structures under physiological conditions, either along their entire lengths or only in localized regions. Instead, these intriguing proteins/regions exist as dynamic ensembles within which atom positions and backbone Ramachandran angles exhibit extreme temporal fluctuations without specific equilibrium values. Many of these intrinsically disordered proteins are known to carry out important biological functions which, in fact, depend on the absence of specific 3-D structure. The existence of such proteins does not fit the prevailing structure-function paradigm, which states that unique 3-D structure is a prerequisite to function. Thus, the protein structure-function paradigm has to be expanded to include intrinsically disordered proteins and alternative relationships among protein sequence, structure, and function. This shift in the paradigm represents a major breakthrough for biochemistry, biophysics and molecular biology, as it opens new levels of understanding with regard to the complex life of proteins. This review will try to answer the following questions: How were intrinsically disordered proteins discovered? Why don't these proteins fold? What is so special about intrinsic disorder? What are the functional advantages of disordered proteins/regions? What is the functional repertoire of these proteins? What are the relationships between intrinsically disordered proteins and human diseases? PMID:20117254

  14. Regulation of protein function by ‘microProteins'


    Staudt, Annica-Carolin; Wenkel, Stephan


    Elegant post-translational regulation is achieved by ‘microProteins', which form homotypic dimers with their targets and act through the dominant–negative suppression of protein complex function. The recent identification of new microProteins suggests their role is general and has evolved in both the plant and animal kingdoms.

  15. Digestion of protein and protein gels in simulated gastric environment

    NARCIS (Netherlands)

    Luo, Q.; Boom, R.M.; Janssen, A.E.M.


    Despite the increasing attention to food digestion research, food scientists still need to better understand the underlying mechanisms of digestion. Most in vitro studies on protein digestion are based on experiments with protein solutions. In this study, the digestion of egg white protein and whey

  16. Molecular simulations of lipid-mediated protein-protein interactions

    NARCIS (Netherlands)

    de Meyer, F.J.M.; Venturoli, M.; Smit, B.


    Recent experimental results revealed that lipid-mediated interactions due to hydrophobic forces may be important in determining the protein topology after insertion in the membrane, in regulating the protein activity, in protein aggregation and in signal transduction. To gain insight into the

  17. The interface of protein structure, protein biophysics, and molecular evolution (United States)

    Liberles, David A; Teichmann, Sarah A; Bahar, Ivet; Bastolla, Ugo; Bloom, Jesse; Bornberg-Bauer, Erich; Colwell, Lucy J; de Koning, A P Jason; Dokholyan, Nikolay V; Echave, Julian; Elofsson, Arne; Gerloff, Dietlind L; Goldstein, Richard A; Grahnen, Johan A; Holder, Mark T; Lakner, Clemens; Lartillot, Nicholas; Lovell, Simon C; Naylor, Gavin; Perica, Tina; Pollock, David D; Pupko, Tal; Regan, Lynne; Roger, Andrew; Rubinstein, Nimrod; Shakhnovich, Eugene; Sjölander, Kimmen; Sunyaev, Shamil; Teufel, Ashley I; Thorne, Jeffrey L; Thornton, Joseph W; Weinreich, Daniel M; Whelan, Simon


    Abstract The interface of protein structural biology, protein biophysics, molecular evolution, and molecular population genetics forms the foundations for a mechanistic understanding of many aspects of protein biochemistry. Current efforts in interdisciplinary protein modeling are in their infancy and the state-of-the art of such models is described. Beyond the relationship between amino acid substitution and static protein structure, protein function, and corresponding organismal fitness, other considerations are also discussed. More complex mutational processes such as insertion and deletion and domain rearrangements and even circular permutations should be evaluated. The role of intrinsically disordered proteins is still controversial, but may be increasingly important to consider. Protein geometry and protein dynamics as a deviation from static considerations of protein structure are also important. Protein expression level is known to be a major determinant of evolutionary rate and several considerations including selection at the mRNA level and the role of interaction specificity are discussed. Lastly, the relationship between modeling and needed high-throughput experimental data as well as experimental examination of protein evolution using ancestral sequence resurrection and in vitro biochemistry are presented, towards an aim of ultimately generating better models for biological inference and prediction. PMID:22528593

  18. Utilization of soya protein as an alternative protein source in ...

    African Journals Online (AJOL)

    In contrast, no significant differences were found in feed and protein utilization parameters. For carcass trait, ash, crude fat, and energy varied significantly with soya protein incorporation in fish diet. Concerning organoleptic characteristics, odour and texture in mouth were not affected by incorporation of soya protein in diet.

  19. Protein engineering techniques gateways to synthetic protein universe

    CERN Document Server

    Poluri, Krishna Mohan


    This brief provides a broad overview of protein-engineering research, offering a glimpse of the most common experimental methods. It also presents various computational programs with applications that are widely used in directed evolution, computational and de novo protein design. Further, it sheds light on the advantages and pitfalls of existing methodologies and future perspectives of protein engineering techniques.

  20. Recent excitements in protein NMR: Large proteins and biologically ...

    Indian Academy of Sciences (India)

    The advent of Transverse Relaxation Optimized SpectroscopY (TROSY) and perdeuteration allowed biomolecularNMR spectroscopists to overcome the size limitation barrier (~20 kDa) in de novo structure determination of proteins.The utility of these techniques was immediately demonstrated on large proteins and protein ...

  1. Protein stress and stress proteins: implications in aging and disease

    Indian Academy of Sciences (India)


    Apr 2, 2007 ... Environmantal stress induces damage that activates an adaptive response in any organism. The cellular stress response is based on the induction of cytoprotective proteins, the so called stress or heat shock proteins. The stress response as well as stress proteins are ubiquitous, highly conserved ...

  2. Protein: MPA1 [TP Atlas

    Lifescience Database Archive (English)

    Full Text Available feron stimulator, Mediator of IRF3 activation, Stimulator of interferon genes protein 9606 Homo sapiens Q86WV6 340061 ... ...MPA1 TLR signaling molecules TMEM173 ERIS, MITA, STING Transmembrane protein 173 Endoplasmic reticulum inter

  3. Epitope tagging of recombinant proteins. (United States)

    Brizzard, B; Chubet, R


    Epitope tagging is a method of expressing proteins whereby an epitope for a specific monoclonal antibody is fused to a target protein using recombinant DNA techniques. The fusion gene is cloned into an appropriate expression vector for the experimental cell type and host cells are transfected. The fusion protein can then be detected and/or purified using a monoclonal antibody specific for the epitope tag. This unit presents protocols for detection and purification of proteins tagged with a particular epitope, the FLAG tag, although the same general approach can be applied to other epitope tags. The protocols in this unit employ the anti-FLAG M2 antibody to detect and purify FLAG-tagged proteins. The methods presented are immunoprecipitation of FLAG fusion proteins from cells using an anti-FLAG M2 affinity gel, detection of FLAG fusion proteins by western blotting, and purification of FLAG fusion proteins by anti-FLAG M2 affinity chromatography.

  4. Protein: FBA3 [TP Atlas

    Lifescience Database Archive (English)

    Full Text Available FBA3 Atg1 kinase complex TOR1 DRR1 Serine/threonine-protein kinase TOR1 Dominant rapamycin... resistance protein 1, Phosphatidylinositol kinase homolog TOR1, Target of rapamycin kinase 1 559292

  5. Functional aspects of protein flexibility

    DEFF Research Database (Denmark)

    Teilum, Kaare; Olsen, Johan G; Kragelund, Birthe B


    Proteins are dynamic entities, and they possess an inherent flexibility that allows them to function through molecular interactions within the cell, among cells and even between organisms. Appreciation of the non-static nature of proteins is emerging, but to describe and incorporate...... this into an intuitive perception of protein function is challenging. Flexibility is of overwhelming importance for protein function, and the changes in protein structure during interactions with binding partners can be dramatic. The present review addresses protein flexibility, focusing on protein-ligand interactions....... The thermodynamics involved are reviewed, and examples of structure-function studies involving experimentally determined flexibility descriptions are presented. While much remains to be understood about protein flexibility, it is clear that it is encoded within their amino acid sequence and should be viewed...

  6. Protein Linked to Atopic Dermatitis (United States)

    ... Research Matters January 14, 2013 Protein Linked to Atopic Dermatitis Normal skin from a mouse (left) shows no ... that lack of a certain protein may trigger atopic dermatitis, the most common type of eczema. The finding ...

  7. Protein-ECE MEtallopincer Hybrids

    NARCIS (Netherlands)

    Kruithof, C.A.


    Modification of proteins with metal complexes is a promising and a relatively new field which conceals many challenges and potential applications. The field is a balance of contributions from the biological (protein engineering, bioconjugation) and chemical sciences (organic, inorganic and

  8. Leptospira Protein Expression During Infection (United States)

    We are characterizing protein expression in vivo during experimental leptospirosis using immunofluorescence microscopy. Coding regions for several proteins were identified through analysis of Leptospira interrogans serovar Copenhageni and L. borgpetersenii serovar Hardjo genomes. In addition, codi...

  9. Yeast Interacting Proteins Database: YJL199C, YJL199C [Yeast Interacting Proteins Database

    Lifescience Database Archive (English)

    Full Text Available d in closely related Saccharomyces species; protein detected in large-scale protein-protein interaction studies...cies; protein detected in large-scale protein-protein interaction studies Rows with this prey as prey (4) Ro...n; not conserved in closely related Saccharomyces species; protein detected in large-scale protein-protein interaction studies... species; protein detected in large-scale protein-protein interaction studies Rows with this prey as prey Ro

  10. Protein: MPA6 [TP Atlas

    Lifescience Database Archive (English)

    Full Text Available MPA6 Adionectin and its receptors Adipoq Acdc, Acrp30, Apm1 Adiponectin 30 kDa adipocyte complement-relate...d protein, Adipocyte complement-related 30 kDa protein, Adipocyte, C1q and collagen domain-containing, Adipocyte-specific protein AdipoQ 10090 Mus musculus 11450 Q60994 1C28, 1C3H Q60994 18446001, 19788607 ...

  11. Dipolar response of hydrated proteins


    Matyushov, Dmitry V.


    The paper presents an analytical theory and numerical simulations of the dipolar response of hydrated proteins. The effective dielectric constant of the solvated protein, representing the average dipole moment induced at the protein by a uniform external field, shows a remarkable variation among the proteins studied by numerical simulations. It changes from 0.5 for ubiquitin to 640 for cytochrome c. The former value implies a negative dipolar susceptibility of ubiquitin, that is a dia-electri...

  12. Protein corona: Opportunities and challenges (United States)

    Zanganeh, Saeid; Spitler, Ryan; Erfanzadeh, Mohsen; Alkilany, Alaaldin M.; Mahmoudi, Morteza


    In contact with biological fluids diverse type of biomolecules (e.g., proteins) adsorb onto nanoparticles forming protein corona. Surface properties of the coated nanoparticles, in terms of type and amount of associated proteins, dictate their interactions with biological systems and thus biological fate, therapeutic efficiency and toxicity. In this perspective, we will focus on the recent advances and pitfalls in the protein corona field. PMID:26783938

  13. The papillomavirus E2 proteins. (United States)

    McBride, Alison A


    The papillomavirus E2 proteins are pivotal to the viral life cycle and have well characterized functions in transcriptional regulation, initiation of DNA replication and partitioning the viral genome. The E2 proteins also function in vegetative DNA replication, post-transcriptional processes and possibly packaging. This review describes structural and functional aspects of the E2 proteins and their binding sites on the viral genome. It is intended to be a reference guide to this viral protein. Published by Elsevier Inc.

  14. Protein corona: Opportunities and challenges. (United States)

    Zanganeh, Saeid; Spitler, Ryan; Erfanzadeh, Mohsen; Alkilany, Alaaldin M; Mahmoudi, Morteza


    In contact with biological fluids diverse type of biomolecules (e.g., proteins) adsorb onto nanoparticles forming protein corona. Surface properties of the coated nanoparticles, in terms of type and amount of associated proteins, dictate their interactions with biological systems and thus biological fate, therapeutic efficiency and toxicity. In this perspective, we will focus on the recent advances and pitfalls in the protein corona field. Copyright © 2016 Elsevier Ltd. All rights reserved.

  15. A Novel Approach for Protein-Named Entity Recognition and Protein-Protein Interaction Extraction

    Directory of Open Access Journals (Sweden)

    Meijing Li


    Full Text Available Many researchers focus on developing protein-named entity recognition (Protein-NER or PPI extraction systems. However, the studies about these two topics cannot be merged well; then existing PPI extraction systems’ Protein-NER still needs to improve. In this paper, we developed the protein-protein interaction extraction system named PPIMiner based on Support Vector Machine (SVM and parsing tree. PPIMiner consists of three main models: natural language processing (NLP model, Protein-NER model, and PPI discovery model. The Protein-NER model, which is named ProNER, identifies the protein names based on two methods: dictionary-based method and machine learning-based method. ProNER is capable of identifying more proteins than dictionary-based Protein-NER model in other existing systems. The final discovered PPIs extracted via PPI discovery model are represented in detail because we showed the protein interaction types and the occurrence frequency through two different methods. In the experiments, the result shows that the performances achieved by our ProNER and PPI discovery model are better than other existing tools. PPIMiner applied this protein-named entity recognition approach and parsing tree based PPI extraction method to improve the performance of PPI extraction. We also provide an easy-to-use interface to access PPIs database and an online system for PPIs extraction and Protein-NER.

  16. Proteins: Chemistry, Characterization, and Quality

    NARCIS (Netherlands)

    Sforza, S.; Tedeschi, T.; Wierenga, P.A.


    Proteins are one of the major macronutrients in food, and several traditional food commodities are good sources of proteins (meat, egg, milk and dairy products, fish, and soya). Proteins are polymers made by 20 different amino acids. They might undergo desired or undesired chemical or enzymatic

  17. Protein: MPA3 [TP Atlas

    Lifescience Database Archive (English)

    Full Text Available MPA3 NADPH oxidase regulators NOXO1 P41NOX, SH3PXD5 NOXO1 NADPH oxidase organizer 1... NADPH oxidase regulatory protein, Nox organizer 1, Nox-organizing protein 1, SH3 and PX domain-containing protein 5 9606 Homo sapiens Q8NFA2 124056 2L73 ...

  18. Protein: MPA3 [TP Atlas

    Lifescience Database Archive (English)

    Full Text Available 1 47 kDa autosomal chronic granulomatous disease protein, 47 kDa neutrophil oxidase factor, NCF-47K, Neutro...phil NADPH oxidase factor 1, Nox organizer 2, Nox-organizing protein 2, SH3 and PX domain-containing protein

  19. Protein: MPB1 [TP Atlas

    Lifescience Database Archive (English)

    Full Text Available MPB1 Related chemokines IL8 CXCL8 Interleukin_8 Interleukin-8 C-X-C motif chemokine... 8, Emoctakin, Granulocyte chemotactic protein 1, Monocyte-derived neutrophil chemotactic factor, Monocyte-d...erived neutrophil-activating peptide, Neutrophil-activating protein 1, Protein 3-10C, T-cell chemotactic fac

  20. Protein: FBA4 [TP Atlas

    Lifescience Database Archive (English)

    Full Text Available ng kinase assembly factor MAT1 CDK7/cyclin-H assembly factor, Cyclin-G1-interacting protein, Menage a trois, RING finger 66, RING finger protein MAT1, p35, p36 9606 Homo sapiens P51948 4331 1G25 4331 P51948 ...

  1. Photoreceptor proteins from purple bacteria

    NARCIS (Netherlands)

    Hendriks, J.; van der Horst, M.A.; Chua, T.K.; Ávila Pérez, M.; van Wilderen, L.J.; Alexandre, M.T.A.; Groot, M.-L.; Kennis, J.T.M.; Hellingwerf, K.J.; Hunter, C.N.; Daldal, F.; Thurnauer, M.C.; Beatty, J.T.


    Purple bacteria contain representatives of four of the six main families of photoreceptor proteins: phytochromes, BLUF domain containing proteins, xanthopsins (i.e., photoactive yellow proteins), and phototropins (containing one or more light, oxygen, or voltage (LOV) domains). Most of them have a

  2. Protein quality of pig diets

    NARCIS (Netherlands)

    Hulshof, Tetske


    The increasing world population and per capita income imposes a risk for protein scarcity. It is, therefore, necessary to use current ingredients more efficiently which includes the accurate assessment of protein quality before inclusion in animal diets. Protein quality is defined in this thesis as

  3. Modeling complexes of modeled proteins. (United States)

    Anishchenko, Ivan; Kundrotas, Petras J; Vakser, Ilya A


    Structural characterization of proteins is essential for understanding life processes at the molecular level. However, only a fraction of known proteins have experimentally determined structures. This fraction is even smaller for protein-protein complexes. Thus, structural modeling of protein-protein interactions (docking) primarily has to rely on modeled structures of the individual proteins, which typically are less accurate than the experimentally determined ones. Such "double" modeling is the Grand Challenge of structural reconstruction of the interactome. Yet it remains so far largely untested in a systematic way. We present a comprehensive validation of template-based and free docking on a set of 165 complexes, where each protein model has six levels of structural accuracy, from 1 to 6 Å C α RMSD. Many template-based docking predictions fall into acceptable quality category, according to the CAPRI criteria, even for highly inaccurate proteins (5-6 Å RMSD), although the number of such models (and, consequently, the docking success rate) drops significantly for models with RMSD > 4 Å. The results show that the existing docking methodologies can be successfully applied to protein models with a broad range of structural accuracy, and the template-based docking is much less sensitive to inaccuracies of protein models than the free docking. Proteins 2017; 85:470-478. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  4. Structuring high-protein foods

    NARCIS (Netherlands)

    Purwanti, N.


    Increased protein consumption gives rise to various health benefits. High-protein intake can lead to muscle development, body weight control and suppression of sarcopenia progression. However, increasing the protein content in food products leads to textural changes over time. These changes result

  5. Functional Foods Containing Whey Proteins (United States)

    Whey proteins, modified whey proteins, and whey components are useful as nutrients or supplements for health maintenance. Extrusion modified whey proteins can easily fit into new products such as beverages, confectionery items (e.g., candies), convenience foods, desserts, baked goods, sauces, and in...

  6. Protein Quantitation Using Mass Spectrometry (United States)

    Zhang, Guoan; Ueberheide, Beatrix M.; Waldemarson, Sofia; Myung, Sunnie; Molloy, Kelly; Eriksson, Jan; Chait, Brian T.; Neubert, Thomas A.; Fenyö, David


    Mass spectrometry is a method of choice for quantifying low-abundance proteins and peptides in many biological studies. Here, we describe a range of computational aspects of protein and peptide quantitation, including methods for finding and integrating mass spectrometric peptide peaks, and detecting interference to obtain a robust measure of the amount of proteins present in samples. PMID:20835801

  7. Biophysics of protein evolution and evolutionary protein biophysics (United States)

    Sikosek, Tobias; Chan, Hue Sun


    The study of molecular evolution at the level of protein-coding genes often entails comparing large datasets of sequences to infer their evolutionary relationships. Despite the importance of a protein's structure and conformational dynamics to its function and thus its fitness, common phylogenetic methods embody minimal biophysical knowledge of proteins. To underscore the biophysical constraints on natural selection, we survey effects of protein mutations, highlighting the physical basis for marginal stability of natural globular proteins and how requirement for kinetic stability and avoidance of misfolding and misinteractions might have affected protein evolution. The biophysical underpinnings of these effects have been addressed by models with an explicit coarse-grained spatial representation of the polypeptide chain. Sequence–structure mappings based on such models are powerful conceptual tools that rationalize mutational robustness, evolvability, epistasis, promiscuous function performed by ‘hidden’ conformational states, resolution of adaptive conflicts and conformational switches in the evolution from one protein fold to another. Recently, protein biophysics has been applied to derive more accurate evolutionary accounts of sequence data. Methods have also been developed to exploit sequence-based evolutionary information to predict biophysical behaviours of proteins. The success of these approaches demonstrates a deep synergy between the fields of protein biophysics and protein evolution. PMID:25165599

  8. Protein-protein interactions and cancer: targeting the central dogma. (United States)

    Garner, Amanda L; Janda, Kim D


    Between 40,000 and 200,000 protein-protein interactions have been predicted to exist within the human interactome. As these interactions are of a critical nature in many important cellular functions and their dysregulation is causal of disease, the modulation of these binding events has emerged as a leading, yet difficult therapeutic arena. In particular, the targeting of protein-protein interactions relevant to cancer is of fundamental importance as the tumor-promoting function of several aberrantly expressed proteins in the cancerous state is directly resultant of its ability to interact with a protein-binding partner. Of significance, these protein complexes play a crucial role in each of the steps of the central dogma of molecular biology, the fundamental processes of genetic transmission. With the many important discoveries being made regarding the mechanisms of these genetic process, the identification of new chemical probes are needed to better understand and validate the druggability of protein-protein interactions related to the central dogma. In this review, we provide an overview of current small molecule-based protein-protein interaction inhibitors for each stage of the central dogma: transcription, mRNA splicing and translation. Importantly, through our analysis we have uncovered a lack of necessary probes targeting mRNA splicing and translation, thus, opening up the possibility for expansion of these fields.

  9. The Proteins API: accessing key integrated protein and genome information. (United States)

    Nightingale, Andrew; Antunes, Ricardo; Alpi, Emanuele; Bursteinas, Borisas; Gonzales, Leonardo; Liu, Wudong; Luo, Jie; Qi, Guoying; Turner, Edd; Martin, Maria


    The Proteins API provides searching and programmatic access to protein and associated genomics data such as curated protein sequence positional annotations from UniProtKB, as well as mapped variation and proteomics data from large scale data sources (LSS). Using the coordinates service, researchers are able to retrieve the genomic sequence coordinates for proteins in UniProtKB. This, the LSS genomics and proteomics data for UniProt proteins is programmatically only available through this service. A Swagger UI has been implemented to provide documentation, an interface for users, with little or no programming experience, to 'talk' to the services to quickly and easily formulate queries with the services and obtain dynamically generated source code for popular programming languages, such as Java, Perl, Python and Ruby. Search results are returned as standard JSON, XML or GFF data objects. The Proteins API is a scalable, reliable, fast, easy to use RESTful services that provides a broad protein information resource for users to ask questions based upon their field of expertise and allowing them to gain an integrated overview of protein annotations available to aid their knowledge gain on proteins in biological processes. The Proteins API is available at ( © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  10. The Proteins API: accessing key integrated protein and genome information (United States)

    Antunes, Ricardo; Alpi, Emanuele; Gonzales, Leonardo; Liu, Wudong; Luo, Jie; Qi, Guoying; Turner, Edd


    Abstract The Proteins API provides searching and programmatic access to protein and associated genomics data such as curated protein sequence positional annotations from UniProtKB, as well as mapped variation and proteomics data from large scale data sources (LSS). Using the coordinates service, researchers are able to retrieve the genomic sequence coordinates for proteins in UniProtKB. This, the LSS genomics and proteomics data for UniProt proteins is programmatically only available through this service. A Swagger UI has been implemented to provide documentation, an interface for users, with little or no programming experience, to ‘talk’ to the services to quickly and easily formulate queries with the services and obtain dynamically generated source code for popular programming languages, such as Java, Perl, Python and Ruby. Search results are returned as standard JSON, XML or GFF data objects. The Proteins API is a scalable, reliable, fast, easy to use RESTful services that provides a broad protein information resource for users to ask questions based upon their field of expertise and allowing them to gain an integrated overview of protein annotations available to aid their knowledge gain on proteins in biological processes. The Proteins API is available at ( PMID:28383659

  11. Characterization of protein-protein interactions by isothermal titration calorimetry. (United States)

    Velazquez-Campoy, Adrian; Leavitt, Stephanie A; Freire, Ernesto


    Isothermal titration calorimetry (ITC) is a powerful technique to study both protein-ligand and protein-protein interactions. This methods chapter is devoted to describing protein-protein interactions, in particular, the association between two different proteins and the self-association of a protein into homodimers. ITC is the only technique that determines directly the thermodynamic parameters of a given reaction: DeltaG, DeltaH, DeltaS, and DeltaCP. Isothermal titration calorimeters have evolved over the years and one of the latest models is the VP-ITC produced by Microcal, Inc. In this chapter we will be describing the general procedure for performing an ITC experiment as well as for the specific cases of porcine pancreatic trypsin binding to soybean trypsin inhibitor and the dissociation of bovine pancreatic alpha-chymotrypsin.

  12. Understanding Protein-Protein Interactions Using Local Structural Features

    DEFF Research Database (Denmark)

    Planas-Iglesias, Joan; Bonet, Jaume; García-García, Javier


    Protein-protein interactions (PPIs) play a relevant role among the different functions of a cell. Identifying the PPI network of a given organism (interactome) is useful to shed light on the key molecular mechanisms within a biological system. In this work, we show the role of structural features...... (loops and domains) to comprehend the molecular mechanisms of PPIs. A paradox in protein-protein binding is to explain how the unbound proteins of a binary complex recognize each other among a large population within a cell and how they find their best docking interface in a short timescale. We use...... interacting and non-interacting protein pairs to classify the structural features that sustain the binding (or non-binding) behavior. Our study indicates that not only the interacting region but also the rest of the protein surface are important for the interaction fate. The interpretation...

  13. Protein subcellular localization assays using split fluorescent proteins (United States)

    Waldo, Geoffrey S [Santa Fe, NM; Cabantous, Stephanie [Los Alamos, NM


    The invention provides protein subcellular localization assays using split fluorescent protein systems. The assays are conducted in living cells, do not require fixation and washing steps inherent in existing immunostaining and related techniques, and permit rapid, non-invasive, direct visualization of protein localization in living cells. The split fluorescent protein systems used in the practice of the invention generally comprise two or more self-complementing fragments of a fluorescent protein, such as GFP, wherein one or more of the fragments correspond to one or more beta-strand microdomains and are used to "tag" proteins of interest, and a complementary "assay" fragment of the fluorescent protein. Either or both of the fragments may be functionalized with a subcellular targeting sequence enabling it to be expressed in or directed to a particular subcellular compartment (i.e., the nucleus).

  14. Diffusion of Integral Membrane Proteins in Protein-Rich Membranes

    DEFF Research Database (Denmark)

    Javanainen, Matti; Martinez-Seara, Hector; Metzler, Ralf


    of being protein-poor, native cell membranes are extremely crowded with proteins. On the basis of extensive molecular simulations, we here demonstrate that protein crowding of the membrane at physiological levels leads to deviations from the SD relation and to the emergence of a stronger Stokes......-like dependence D ∝ 1/R. We propose that this 1/R law mainly arises due to geometrical factors: smaller proteins are able to avoid confinement effects much better than their larger counterparts. The results highlight that the lateral dynamics in the crowded setting found in native membranes is radically different......The lateral diffusion of embedded proteins along lipid membranes in protein-poor conditions has been successfully described in terms of the Saffman-Delbrück (SD) model, which predicts that the protein diffusion coefficient D is weakly dependent on its radius R as D ∝ ln(1/R). However, instead...

  15. Protein from methanol

    Energy Technology Data Exchange (ETDEWEB)

    Rosenzweig, M.; Ushio, S.


    The biosynthesis of proteins from methanol produced from natural gas can provide an attractive alternative to the already commercially proven technique of protein synthesis from gas oil and n-paraffin feedstocks if current pilot-plant tests in England and Japan prove successful. The methanol route also provides other advantages as a protein feedstock: it is water soluble, contains no polycyclic aromatic compounds, and requires less oxygen than methane. Its lower boiling point helps ease the separation of feedstock from the product stream. Finally, it will require lower investment costs. Both ICI and Mitsubishi Gas Chemical Co. are large methanol producers. ICI already has a 1000 ton/yr plant operating at Teeside, England, and expects to decide on a 100,000 m ton/yr plant later this year. Mitsubishi is constructing a large-scale pilot plant scheduled to come onstream this year. ICI will use a Pseudomona bacterium at 98.6/sup 0/F (37/sup 0/C) in the fermenter. Mitsubishi has not yet decided on a yeast or a bacteria, and is searching for a strain capable of withstanding up to 115/sup 0/F (46/sup 0/C). In the more advanced ICI process, methanol will be mixed with phosphoric acid, potassium sulfate, sodium chloride, and traces of iron, copper, zinc, and molybdenum; diluted with water; passed through a sterilization tank; and fermented at pH 7 in a pressure cycle fermenter. The product stream, containing a 3 percent suspension of cellular dry matter, is taken near the top of the fermenter riser, then passed through a flotation vessel and a centrifuge to pack the cell concentration to 20 percent. Water is recycled. Whatever methanol remains in the fermenter product stream is either used up by the microorganisms in subsequent processing or vaporized in the dryer. (auth)

  16. NMR Studies of Protein Hydration and Protein-Ligand Interactions (United States)

    Chong, Yuan

    Water on the surface of a protein is called hydration water. Hydration water is known to play a crucial role in a variety of biological processes including protein folding, enzymatic activation, and drug binding. Although the significance of hydration water has been recognized, the underlying mechanism remains far from being understood. This dissertation employs a unique in-situ nuclear magnetic resonance (NMR) technique to study the mechanism of protein hydration and the role of hydration in alcohol-protein interactions. Water isotherms in proteins are measured at different temperatures via the in-situ NMR technique. Water is found to interact differently with hydrophilic and hydrophobic groups on the protein. Water adsorption on hydrophilic groups is hardly affected by the temperature, while water adsorption on hydrophobic groups strongly depends on the temperature around 10 C, below which the adsorption is substantially reduced. This effect is induced by the dramatic decrease in the protein flexibility below 10 C. Furthermore, nanosecond to microsecond protein dynamics and the free energy, enthalpy, and entropy of protein hydration are studied as a function of hydration level and temperature. A crossover at 10 C in protein dynamics and thermodynamics is revealed. The effect of water at hydrophilic groups on protein dynamics and thermodynamics shows little temperature dependence, whereas water at hydrophobic groups has stronger effect above 10 C. In addition, I investigate the role of water in alcohol binding to the protein using the in-situ NMR detection. The isotherms of alcohols are first measured on dry proteins, then on proteins with a series of controlled hydration levels. The free energy, enthalpy, and entropy of alcohol binding are also determined. Two distinct types of alcohol binding are identified. On the one hand, alcohols can directly bind to a few specific sites on the protein. This type of binding is independent of temperature and can be

  17. Protein-Protein Interactions: Structurally Conserved Residues Distinguish between Binding Sites and Exposed Protein Surfaces

    National Research Council Canada - National Science Library

    Buyong Ma; Tal Elkayam; Haim Wolfson; Ruth Nussinov


    Polar residue hot spots have been observed at protein-protein binding sites. Here we show that hot spots occur predominantly at the interfaces of macromolecular complexes, distinguishing binding sites from the remainder of the surface...

  18. Information contained in protein shapes (United States)

    Sundaram, K.; Viswanadhan, V. N.; Macelroy, R. D.


    The sequence of local conformations at C-alpha atoms of a protein has been considered as an informational message string. The total self-information contents and self-information per letter have been evaluated for 83 globular proteins whose structures are known from X-ray crystallography. The derived information contents provide a method of quantitating structural specificity of proteins. This method of analysis enables repeating, intricate structural features to be recognized. Among the globular proteins whose structures have been solved, high potential iron protein stands out with the largest three-letter dependence.

  19. Protein-protein interaction network-based detection of functionally similar proteins within species. (United States)

    Song, Baoxing; Wang, Fen; Guo, Yang; Sang, Qing; Liu, Min; Li, Dengyun; Fang, Wei; Zhang, Deli


    Although functionally similar proteins across species have been widely studied, functionally similar proteins within species showing low sequence similarity have not been examined in detail. Identification of these proteins is of significant importance for understanding biological functions, evolution of protein families, progression of co-evolution, and convergent evolution and others which cannot be obtained by detection of functionally similar proteins across species. Here, we explored a method of detecting functionally similar proteins within species based on graph theory. After denoting protein-protein interaction networks using graphs, we split the graphs into subgraphs using the 1-hop method. Proteins with functional similarities in a species were detected using a method of modified shortest path to compare these subgraphs and to find the eligible optimal results. Using seven protein-protein interaction networks and this method, some functionally similar proteins with low sequence similarity that cannot detected by sequence alignment were identified. By analyzing the results, we found that, sometimes, it is difficult to separate homologous from convergent evolution. Evaluation of the performance of our method by gene ontology term overlap showed that the precision of our method was excellent. Copyright © 2012 Wiley Periodicals, Inc.

  20. Tetramer formation in Arabidopsis MADS domain proteins: analysis of a protein-protein interaction network

    NARCIS (Netherlands)

    Espinosa-Soto, C.; Immink, R.G.H.; Angenent, G.C.; Alvarez-Buylla, E.R.; Folter, de S.


    Background: MADS domain proteins are transcription factors that coordinate several important developmental processes in plants. These proteins interact with other MADS domain proteins to form dimers, and it has been proposed that they are able to associate as tetrameric complexes that regulate

  1. Discover Protein Complexes in Protein-Protein Interaction Networks Using Parametric Local Modularity

    Directory of Open Access Journals (Sweden)

    Tan Kai


    Full Text Available Abstract Background Recent advances in proteomic technologies have enabled us to create detailed protein-protein interaction maps in multiple species and in both normal and diseased cells. As the size of the interaction dataset increases, powerful computational methods are required in order to effectively distil network models from large-scale interactome data. Results We present an algorithm, miPALM (Module Inference by Parametric Local Modularity, to infer protein complexes in a protein-protein interaction network. The algorithm uses a novel graph theoretic measure, parametric local modularity, to identify highly connected sub-networks as candidate protein complexes. Using gold standard sets of protein complexes and protein function and localization annotations, we show our algorithm achieved an overall improvement over previous algorithms in terms of precision, recall, and biological relevance of the predicted complexes. We applied our algorithm to predict and characterize a set of 138 novel protein complexes in S. cerevisiae. Conclusions miPALM is a novel algorithm for detecting protein complexes from large protein-protein interaction networks with improved accuracy than previous methods. The software is implemented in Matlab and is freely available at

  2. Detection of protein complex from protein-protein interaction network using Markov clustering (United States)

    Ochieng, P. J.; Kusuma, W. A.; Haryanto, T.


    Detection of complexes, or groups of functionally related proteins, is an important challenge while analysing biological networks. However, existing algorithms to identify protein complexes are insufficient when applied to dense networks of experimentally derived interaction data. Therefore, we introduced a graph clustering method based on Markov clustering algorithm to identify protein complex within highly interconnected protein-protein interaction networks. Protein-protein interaction network was first constructed to develop geometrical network, the network was then partitioned using Markov clustering to detect protein complexes. The interest of the proposed method was illustrated by its application to Human Proteins associated to type II diabetes mellitus. Flow simulation of MCL algorithm was initially performed and topological properties of the resultant network were analysed for detection of the protein complex. The results indicated the proposed method successfully detect an overall of 34 complexes with 11 complexes consisting of overlapping modules and 20 non-overlapping modules. The major complex consisted of 102 proteins and 521 interactions with cluster modularity and density of 0.745 and 0.101 respectively. The comparison analysis revealed MCL out perform AP, MCODE and SCPS algorithms with high clustering coefficient (0.751) network density and modularity index (0.630). This demonstrated MCL was the most reliable and efficient graph clustering algorithm for detection of protein complexes from PPI networks.

  3. Human cancer protein-protein interaction network: a structural perspective.

    Directory of Open Access Journals (Sweden)

    Gozde Kar


    Full Text Available Protein-protein interaction networks provide a global picture of cellular function and biological processes. Some proteins act as hub proteins, highly connected to others, whereas some others have few interactions. The dysfunction of some interactions causes many diseases, including cancer. Proteins interact through their interfaces. Therefore, studying the interface properties of cancer-related proteins will help explain their role in the interaction networks. Similar or overlapping binding sites should be used repeatedly in single interface hub proteins, making them promiscuous. Alternatively, multi-interface hub proteins make use of several distinct binding sites to bind to different partners. We propose a methodology to integrate protein interfaces into cancer interaction networks (ciSPIN, cancer structural protein interface network. The interactions in the human protein interaction network are replaced by interfaces, coming from either known or predicted complexes. We provide a detailed analysis of cancer related human protein-protein interfaces and the topological properties of the cancer network. The results reveal that cancer-related proteins have smaller, more planar, more charged and less hydrophobic binding sites than non-cancer proteins, which may indicate low affinity and high specificity of the cancer-related interactions. We also classified the genes in ciSPIN according to phenotypes. Within phenotypes, for breast cancer, colorectal cancer and leukemia, interface properties were found to be discriminating from non-cancer interfaces with an accuracy of 71%, 67%, 61%, respectively. In addition, cancer-related proteins tend to interact with their partners through distinct interfaces, corresponding mostly to multi-interface hubs, which comprise 56% of cancer-related proteins, and constituting the nodes with higher essentiality in the network (76%. We illustrate the interface related affinity properties of two cancer-related hub

  4. Molecular principles of human virus protein-protein interactions. (United States)

    Halehalli, Rachita Ramachandra; Nagarajaram, Hampapathalu Adimurthy


    Viruses, from the human protein-protein interaction network perspective, target hubs, bottlenecks and interconnected nodes enriched in certain biological pathways. However, not much is known about the general characteristic features of the human proteins interacting with viral proteins (referred to as hVIPs) as well as the motifs and domains utilized by human-virus protein-protein interactions (referred to as Hu-Vir PPIs). Our study has revealed that hVIPs are mostly disordered proteins, whereas viral proteins are mostly ordered proteins. Protein disorder in viral proteins and hVIPs varies from one subcellular location to another. In any given viral-human PPI pair, at least one of the two proteins is structurally disordered suggesting that disorder associated conformational flexibility as one of the characteristic features of virus-host interaction. Further analyses reveal that hVIPs are (i) slowly evolving proteins, (ii) associated with high centrality scores in human-PPI network, (iii) involved in multiple pathways, (iv) enriched in eukaryotic linear motifs (ELMs) associated with protein modification, degradation and regulatory processes, (v) associated with high number of splice variants and (vi) expressed abundantly across multiple tissues. These aforementioned findings suggest that conformational flexibility, spatial diversity, abundance and slow evolution are the characteristic features of the human proteins targeted by viral proteins. Hu-Vir PPIs are mostly mediated via domain-motif interactions (DMIs) where viral proteins employ motifs that mimic host ELMs to bind to domains in human proteins. DMIs are shared among viruses belonging to different families indicating a possible convergent evolution of these motifs to help viruses to adopt common strategies to subvert host cellular pathways. Hu-Vir PPI data, DDI and DMI data for human-virus PPI can be downloaded from Supplementary data are

  5. Introduction to protein crystallization. (United States)

    McPherson, Alexander; Gavira, Jose A


    Protein crystallization was discovered by chance about 150 years ago and was developed in the late 19th century as a powerful purification tool and as a demonstration of chemical purity. The crystallization of proteins, nucleic acids and large biological complexes, such as viruses, depends on the creation of a solution that is supersaturated in the macromolecule but exhibits conditions that do not significantly perturb its natural state. Supersaturation is produced through the addition of mild precipitating agents such as neutral salts or polymers, and by the manipulation of various parameters that include temperature, ionic strength and pH. Also important in the crystallization process are factors that can affect the structural state of the macromolecule, such as metal ions, inhibitors, cofactors or other conventional small molecules. A variety of approaches have been developed that combine the spectrum of factors that effect and promote crystallization, and among the most widely used are vapor diffusion, dialysis, batch and liquid-liquid diffusion. Successes in macromolecular crystallization have multiplied rapidly in recent years owing to the advent of practical, easy-to-use screening kits and the application of laboratory robotics. A brief review will be given here of the most popular methods, some guiding principles and an overview of current technologies.

  6. Bioactive proteins from pipefishes

    Directory of Open Access Journals (Sweden)

    E. Rethna Priya


    Full Text Available Objective: To screen antimicrobial potence of some pipefish species collected from Tuticorin coastal environment. Methods: Antimicrobial activity of pipefishes in methanol extract was investigated against 10 bacterial and 10 fungal human pathogenic strains. Results: Among the tested strains, in Centriscus scutatus, pipefish showed maximum zone of inhibition against Vibrio cholerae (8 mm and minimum in the sample of Hippichthys cyanospilos against Klebseilla pneumoniae (2 mm. In positive control, maximum zone of inhibition was recorded in Vibrio cholerae (9 mm and minimum in Klebseilla pneumoniae, and Salmonella paratyphi (5 mm. Chemical investigation indicated the presence of peptides as evidenced by ninhydrin positive spots on thin layer chromatography and presence of peptide. In SDS PAGE, in Centriscus scutatus, four bands were detected in the gel that represented the presence of proteins in the range nearly 25.8-75 kDa. In Hippichthys cyanospilos, five bands were detected in the gel that represented the presence of proteins in the range nearly 20.5-78 kDa. The result of FT-IR spectrum revealed that the pipe fishes extracts compriseed to have peptide derivatives as their predominant chemical groups. Conclusions: It can be conclude that this present investigation suggests the tested pipe fishes will be a potential source of natural bioactive compounds.

  7. Bioactive proteins from pipefishes

    Directory of Open Access Journals (Sweden)

    E. Rethna Priya


    Full Text Available Objective: To screen antimicrobial potence of some pipefish species collected from Tuticorin coastal environment. Methods: Antimicrobial activity of pipefishes in methanol extract was investigated against 10 bacterial and 10 fungal human pathogenic strains. Results: Among the tested strains, in Centriscus scutatus, pipefish showed maximum zone of inhibition against Vibrio cholerae (8 mm and minimum in the sample of Hippichthys cyanospilos against Klebseilla pneumoniae (2 mm. In positive control, maximum zone of inhibition was recorded in Vibrio cholerae (9 mm and minimum in Klebseilla pneumoniae, and Salmonella paratyphi (5 mm. Chemical investigation indicated the presence of peptides as evidenced by ninhydrin positive spots on thin layer chromatography and presence of peptide. In SDS PAGE, in Centriscus scutatus, four bands were detected in the gel that represented the presence of proteins in the range nearly 25.8-75 kDa. In Hippichthys cyanospilos, five bands were detected in the gel that represented the presence of proteins in the range nearly 20.5-78 kDa. The result of FT-IR spectrum revealed that the pipe fishes extracts compriseed to have peptide derivatives as their predominant chemical groups. Conclusions: It can be conclude that this present investigation suggests the tested pipe fishes will be a potential source of natural bioactive compounds.

  8. Protein Chemical Shift Prediction

    CERN Document Server

    Larsen, Anders S


    The protein chemical shifts holds a large amount of information about the 3-dimensional structure of the protein. A number of chemical shift predictors based on the relationship between structures resolved with X-ray crystallography and the corresponding experimental chemical shifts have been developed. These empirical predictors are very accurate on X-ray structures but tends to be insensitive to small structural changes. To overcome this limitation it has been suggested to make chemical shift predictors based on quantum mechanical(QM) calculations. In this thesis the development of the QM derived chemical shift predictor Procs14 is presented. Procs14 is based on 2.35 million density functional theory(DFT) calculations on tripeptides and contains corrections for hydrogen bonding, ring current and the effect of the previous and following residue. Procs14 is capable at performing predictions for the 13CA, 13CB, 13CO, 15NH, 1HN and 1HA backbone atoms. In order to benchmark Procs14, a number of QM NMR calculatio...

  9. Water-transporting proteins. (United States)

    Zeuthen, Thomas


    Transport through lipids and aquaporins is osmotic and entirely driven by the difference in osmotic pressure. Water transport in cotransporters and uniporters is different: Water can be cotransported, energized by coupling to the substrate flux by a mechanism closely associated with protein. In the K(+)/Cl(-) and the Na(+)/K(+)/2Cl(-) cotransporters, water is entirely cotransported, while water transport in glucose uniporters and Na(+)-coupled transporters of nutrients and neurotransmitters takes place by both osmosis and cotransport. The molecular mechanism behind cotransport of water is not clear. It is associated with the substrate movements in aqueous pathways within the protein; a conventional unstirred layer mechanism can be ruled out, due to high rates of diffusion in the cytoplasm. The physiological roles of the various modes of water transport are reviewed in relation to epithelial transport. Epithelial water transport is energized by the movements of ions, but how the coupling takes place is uncertain. All epithelia can transport water uphill against an osmotic gradient, which is hard to explain by simple osmosis. Furthermore, genetic removal of aquaporins has not given support to osmosis as the exclusive mode of transport. Water cotransport can explain the coupling between ion and water transport, a major fraction of transepithelial water transport and uphill water transport. Aquaporins enhance water transport by utilizing osmotic gradients and cause the osmolarity of the transportate to approach isotonicity.

  10. Mathematical methods for protein science

    Energy Technology Data Exchange (ETDEWEB)

    Hart, W.; Istrail, S.; Atkins, J. [Sandia National Labs., Albuquerque, NM (United States)


    Understanding the structure and function of proteins is a fundamental endeavor in molecular biology. Currently, over 100,000 protein sequences have been determined by experimental methods. The three dimensional structure of the protein determines its function, but there are currently less than 4,000 structures known to atomic resolution. Accordingly, techniques to predict protein structure from sequence have an important role in aiding the understanding of the Genome and the effects of mutations in genetic disease. The authors describe current efforts at Sandia to better understand the structure of proteins through rigorous mathematical analyses of simple lattice models. The efforts have focused on two aspects of protein science: mathematical structure prediction, and inverse protein folding.

  11. Metagenomics and the protein universe (United States)

    Godzik, Adam


    Metagenomics sequencing projects have dramatically increased our knowledge of the protein universe and provided over one-half of currently known protein sequences; they have also introduced a much broader phylogenetic diversity into the protein databases. The full analysis of metagenomic datasets is only beginning, but it has already led to the discovery of thousands of new protein families, likely representing novel functions specific to given environments. At the same time, a deeper analysis of such novel families, including experimental structure determination of some representatives, suggests that most of them represent distant homologs of already characterized protein families, and thus most of the protein diversity present in the new environments are due to functional divergence of the known protein families rather than the emergence of new ones. PMID:21497084

  12. The Papillomavirus E2 proteins

    Energy Technology Data Exchange (ETDEWEB)

    McBride, Alison A., E-mail:


    The papillomavirus E2 proteins are pivotal to the viral life cycle and have well characterized functions in transcriptional regulation, initiation of DNA replication and partitioning the viral genome. The E2 proteins also function in vegetative DNA replication, post-transcriptional processes and possibly packaging. This review describes structural and functional aspects of the E2 proteins and their binding sites on the viral genome. It is intended to be a reference guide to this viral protein. - Highlights: • Overview of E2 protein functions. • Structural domains of the papillomavirus E2 proteins. • Analysis of E2 binding sites in different genera of papillomaviruses. • Compilation of E2 associated proteins. • Comparison of key mutations in distinct E2 functions.

  13. Protein folding and wring resonances

    DEFF Research Database (Denmark)

    Bohr, Jakob; Bohr, Henrik; Brunak, Søren


    The polypeptide chain of a protein is shown to obey topological contraints which enable long range excitations in the form of wring modes of the protein backbone. Wring modes of proteins of specific lengths can therefore resonate with molecular modes present in the cell. It is suggested...... that protein folding takes place when the amplitude of a wring excitation becomes so large that it is energetically favorable to bend the protein backbone. The condition under which such structural transformations can occur is found, and it is shown that both cold and hot denaturation (the unfolding...... of proteins) are natural consequences of the suggested wring mode model. Native (folded) proteins are found to possess an intrinsic standing wring mode....

  14. Advantages of proteins being disordered. (United States)

    Liu, Zhirong; Huang, Yongqi


    The past decade has witnessed great advances in our understanding of protein structure-function relationships in terms of the ubiquitous existence of intrinsically disordered proteins (IDPs) and intrinsically disordered regions (IDRs). The structural disorder of IDPs/IDRs enables them to play essential functions that are complementary to those of ordered proteins. In addition, IDPs/IDRs are persistent in evolution. Therefore, they are expected to possess some advantages over ordered proteins. In this review, we summarize and survey nine possible advantages of IDPs/IDRs: economizing genome/protein resources, overcoming steric restrictions in binding, achieving high specificity with low affinity, increasing binding rate, facilitating posttranslational modifications, enabling flexible linkers, preventing aggregation, providing resistance to non-native conditions, and allowing compatibility with more available sequences. Some potential advantages of IDPs/IDRs are not well understood and require both experimental and theoretical approaches to decipher. The connection with protein design is also briefly discussed. © 2014 The Protein Society.

  15. Protein Adsorption in Three Dimensions (United States)

    Vogler, Erwin A.


    Recent experimental and theoretical work clarifying the physical chemistry of blood-protein adsorption from aqueous-buffer solution to various kinds of surfaces is reviewed and interpreted within the context of biomaterial applications, especially toward development of cardiovascular biomaterials. The importance of this subject in biomaterials surface science is emphasized by reducing the “protein-adsorption problem” to three core questions that require quantitative answer. An overview of the protein-adsorption literature identifies some of the sources of inconsistency among many investigators participating in more than five decades of focused research. A tutorial on the fundamental biophysical chemistry of protein adsorption sets the stage for a detailed discussion of the kinetics and thermodynamics of protein adsorption, including adsorption competition between two proteins for the same adsorbent immersed in a binary-protein mixture. Both kinetics and steady-state adsorption can be rationalized using a single interpretive paradigm asserting that protein molecules partition from solution into a three-dimensional (3D) interphase separating bulk solution from the physical-adsorbent surface. Adsorbed protein collects in one-or-more adsorbed layers, depending on protein size, solution concentration, and adsorbent surface energy (water wettability). The adsorption process begins with the hydration of an adsorbent surface brought into contact with an aqueous-protein solution. Surface hydration reactions instantaneously form a thin, pseudo-2D interface between the adsorbent and protein solution. Protein molecules rapidly diffuse into this newly-formed interface, creating a truly 3D interphase that inflates with arriving proteins and fills to capacity within milliseconds at mg/mL bulk-solution concentrations CB. This inflated interphase subsequently undergoes time-dependent (minutes-to-hours) decrease in volume VI by expulsion of either-or-both interphase water and

  16. Protein function prediction via graph kernels

    National Research Council Canada - National Science Library

    Borgwardt, Karsten M; Ong, Cheng Soon; Schönauer, Stefan; Vishwanathan, S V N; Smola, Alex J; Kriegel, Hans-Peter


    Computational approaches to protein function prediction infer protein function by finding proteins with similar sequence, structure, surface clefts, chemical properties, amino acid motifs, interaction...

  17. Protein oxidation in aging and the removal of oxidized proteins. (United States)

    Höhn, Annika; König, Jeannette; Grune, Tilman


    Reactive oxygen species (ROS) are generated constantly within cells at low concentrations even under physiological conditions. During aging the levels of ROS can increase due to a limited capacity of antioxidant systems and repair mechanisms. Proteins are among the main targets for oxidants due to their high rate constants for several reactions with ROS and their abundance in biological systems. Protein damage has an important influence on cellular viability since most protein damage is non-repairable, and has deleterious consequences on protein structure and function. In addition, damaged and modified proteins can form cross-links and provide a basis for many senescence-associated alterations and may contribute to a range of human pathologies. Two proteolytic systems are responsible to ensure the maintenance of cellular functions: the proteasomal (UPS) and the lysosomal system. Those degrading systems provide a last line of antioxidative protection, removing irreversible damaged proteins and recycling amino acids for the continuous protein synthesis. But during aging, both systems are affected and their proteolytic activity declines significantly. Here we highlight the recent advantages in the understanding of protein oxidation and the fate of these damaged proteins during aging. This article is part of a Special Issue entitled: Posttranslational Protein modifications in biology and Medicine. Copyright © 2013 Elsevier B.V. All rights reserved.

  18. Protein-protein interaction based on pairwise similarity

    Directory of Open Access Journals (Sweden)

    Zaki Nazar


    Full Text Available Abstract Background Protein-protein interaction (PPI is essential to most biological processes. Abnormal interactions may have implications in a number of neurological syndromes. Given that the association and dissociation of protein molecules is crucial, computational tools capable of effectively identifying PPI are desirable. In this paper, we propose a simple yet effective method to detect PPI based on pairwise similarity and using only the primary structure of the protein. The PPI based on Pairwise Similarity (PPI-PS method consists of a representation of each protein sequence by a vector of pairwise similarities against large subsequences of amino acids created by a shifting window which passes over concatenated protein training sequences. Each coordinate of this vector is typically the E-value of the Smith-Waterman score. These vectors are then used to compute the kernel matrix which will be exploited in conjunction with support vector machines. Results To assess the ability of the proposed method to recognize the difference between "interacted" and "non-interacted" proteins pairs, we applied it on different datasets from the available yeast saccharomyces cerevisiae protein interaction. The proposed method achieved reasonable improvement over the existing state-of-the-art methods for PPI prediction. Conclusion Pairwise similarity score provides a relevant measure of similarity between protein sequences. This similarity incorporates biological knowledge about proteins and it is extremely powerful when combined with support vector machine to predict PPI.

  19. Bioinformatic Prediction of WSSV-Host Protein-Protein Interaction

    Directory of Open Access Journals (Sweden)

    Zheng Sun


    Full Text Available WSSV is one of the most dangerous pathogens in shrimp aquaculture. However, the molecular mechanism of how WSSV interacts with shrimp is still not very clear. In the present study, bioinformatic approaches were used to predict interactions between proteins from WSSV and shrimp. The genome data of WSSV (NC_003225.1 and the constructed transcriptome data of F. chinensis were used to screen potentially interacting proteins by searching in protein interaction databases, including STRING, Reactome, and DIP. Forty-four pairs of proteins were suggested to have interactions between WSSV and the shrimp. Gene ontology analysis revealed that 6 pairs of these interacting proteins were classified into “extracellular region” or “receptor complex” GO-terms. KEGG pathway analysis showed that they were involved in the “ECM-receptor interaction pathway.” In the 6 pairs of interacting proteins, an envelope protein called “collagen-like protein” (WSSV-CLP encoded by an early virus gene “wsv001” in WSSV interacted with 6 deduced proteins from the shrimp, including three integrin alpha (ITGA, two integrin beta (ITGB, and one syndecan (SDC. Sequence analysis on WSSV-CLP, ITGA, ITGB, and SDC revealed that they possessed the sequence features for protein-protein interactions. This study might provide new insights into the interaction mechanisms between WSSV and shrimp.

  20. A new protein structure representation for efficient protein function prediction. (United States)

    Maghawry, Huda A; Mostafa, Mostafa G M; Gharib, Tarek F


    One of the challenging problems in bioinformatics is the prediction of protein function. Protein function is the main key that can be used to classify different proteins. Protein function can be inferred experimentally with very small throughput or computationally with very high throughput. Computational methods are sequence based or structure based. Structure-based methods produce more accurate protein function prediction. In this article, we propose a new protein structure representation for efficient protein function prediction. The representation is based on three-dimensional patterns of protein residues. In the analysis, we used protein function based on enzyme activity through six mechanistically diverse enzyme superfamilies: amidohydrolase, crotonase, haloacid dehalogenase, isoprenoid synthase type I, and vicinal oxygen chelate. We applied three different classification methods, naïve Bayes, k-nearest neighbors, and random forest, to predict the enzyme superfamily of a given protein. The prediction accuracy using the proposed representation outperforms a recently introduced representation method that is based only on the distance patterns. The results show that the proposed representation achieved prediction accuracy up to 98%, with improvement of about 10% on average.

  1. Role for protein-protein interaction databases in human genetics. (United States)

    Pattin, Kristine A; Moore, Jason H


    Proteomics and the study of protein-protein interactions are becoming increasingly important in our effort to understand human diseases on a system-wide level. Thanks to the development and curation of protein-interaction databases, up-to-date information on these interaction networks is accessible and publicly available to the scientific community. As our knowledge of protein-protein interactions increases, it is important to give thought to the different ways that these resources can impact biomedical research. In this article, we highlight the importance of protein-protein interactions in human genetics and genetic epidemiology. Since protein-protein interactions demonstrate one of the strongest functional relationships between genes, combining genomic data with available proteomic data may provide us with a more in-depth understanding of common human diseases. In this review, we will discuss some of the fundamentals of protein interactions, the databases that are publicly available and how information from these databases can be used to facilitate genome-wide genetic studies.

  2. Mapping Protein-Protein Interactions by Quantitative Proteomics

    DEFF Research Database (Denmark)

    Dengjel, Joern; Kratchmarova, Irina; Blagoev, Blagoy


    Proteins exert their function inside a cell generally in multiprotein complexes. These complexes are highly dynamic structures changing their composition over time and cell state. The same protein may thereby fulfill different functions depending on its binding partners. Quantitative mass...... spectrometry (MS)-based proteomics in combination with affinity purification protocols has become the method of choice to map and track the dynamic changes in protein-protein interactions, including the ones occurring during cellular signaling events. Different quantitative MS strategies have been used...... to characterize protein interaction networks. In this chapter we describe in detail the use of stable isotope labeling by amino acids in cell culture (SILAC) for the quantitative analysis of stimulus-dependent dynamic protein interactions....

  3. Revisiting the Voronoi description of protein-protein interfaces. (United States)

    Cazals, Frédéric; Proust, Flavien; Bahadur, Ranjit P; Janin, Joël


    We developed a model of macromolecular interfaces based on the Voronoi diagram and the related alpha-complex, and we tested its properties on a set of 96 protein-protein complexes taken from the Protein Data Bank. The Voronoi model provides a natural definition of the interfaces, and it yields values of the number of interface atoms and of the interface area that have excellent correlation coefficients with those of the classical model based on solvent accessibility. Nevertheless, some atoms that do not lose solvent accessibility are part of the interface defined by the Voronoi model. The Voronoi model provides robust definitions of the curvature and of the connectivity of the interfaces, and leads to estimates of these features that generally agree with other approaches. Our implementation of the model allows an analysis of protein-water contacts that highlights the role of structural water molecules at protein-protein interfaces.

  4. Duchenne Muscular Dystrophy (DMD) Protein-Protein Interaction Mapping. (United States)

    Rezaei Tavirani, Mostafa; OkHOVATIAN, Farshad; Zamanian Azodi, Mona; Rezaei Tavirani, Majid


    Duchenne muscular dystrophy (DMD) is one of the mortal diseases, subjected to study in terms of molecular investigation. In this study, the protein interaction map of this muscle-wasting condition was generated to gain a better knowledge of interactome profile of DMD. Applying Cytoscape and String Database, the protein-protein interaction network was constructed and the gene ontology of the constructed network was analyzed for biological process, molecular function, and cellular component annotations. Among 100 proteins related to DMD, dystrophin, utrophin, caveolin 3, and myogenic differentiation 1 play key roles in DMD network. In addition, the gene ontology analysis showed that regulation processes, kinase activity, and sarcoplasmic reticulum were the highlighted biological processes, molecular function, and cell component enrichments respectively for the proteins related to DMD. The central proteins and the enriched ontologies can be suggested as possible prominent agents in DMD; however, the validation studies may be required.

  5. On the role of electrostatics on protein-protein interactions (United States)

    Zhang, Zhe; Witham, Shawn; Alexov, Emil


    The role of electrostatics on protein-protein interactions and binding is reviewed in this article. A brief outline of the computational modeling, in the framework of continuum electrostatics, is presented and basic electrostatic effects occurring upon the formation of the complex are discussed. The role of the salt concentration and pH of the water phase on protein-protein binding free energy is demonstrated and indicates that the increase of the salt concentration tends to weaken the binding, an observation that is attributed to the optimization of the charge-charge interactions across the interface. It is pointed out that the pH-optimum (pH of optimal binding affinity) varies among the protein-protein complexes, and perhaps is a result of their adaptation to particular subcellular compartment. At the end, the similarities and differences between hetero- and homo-complexes are outlined and discussed with respect to the binding mode and charge complementarity. PMID:21572182

  6. Methods for detection of protein-protein and protein-DNA interactions using HaloTag. (United States)

    Urh, Marjeta; Hartzell, Danette; Mendez, Jacqui; Klaubert, Dieter H; Wood, Keith


    HaloTag is a protein fusion tag which was genetically engineered to covalently bind a series of specific synthetic ligands. All ligands carry two groups, the reactive group and the functional/reporter group. The reactive group, the choloroalkane, is the same in all the ligands and is involved in binding to the HaloTag. The functional reporter group is variable and can carry many different moieties including fluorescent dyes, affinity handles like biotin or solid surfaces such as agarose beads. Thus, HaloTag can serve either as a labeling tag or as a protein immobilization tag depending on which ligand is bound to it. Here, we describe a procedure for immobilization of HaloTag fusion proteins and how immobilized proteins can be used to study protein-protein and protein-DNA interactions in vivo and in vitro.

  7. Manipulating protein adsorption using a patchy protein-resistant brush. (United States)

    Gon, Saugata; Bendersky, Marina; Ross, Jennifer L; Santore, Maria M


    Toward the development of surfaces for the precise manipulation of proteins, this study explores the fabrication and protein-interactive behavior of a new type of surface containing extremely small (on the order of 10 nm or less) flat adhesive "patches" or islands embedded in and partially concealed by a protein-repellant PEG (poly(ethylene glycol)) brush. The adsorption of fibrinogen, the model protein chosen to probe the biomaterial interactions of these surfaces, is very sensitive to the surface density of the adhesive patches, occurring only above a threshold. This suggests that two or more adhesive patches are needed to capture each protein. When the average spacing of the adhesive patches exceeds the fibrinogen length, no adsorption occurs because individual patches are too weakly binding for protein capture, as a result of being at least partially obstructed by the brush. The small size of the adhesive patches relative to the 47 nm fibrinogen length thus defines a limiting regime of surface design, distinct from surfaces where larger features can adhere single isolated proteins or multiple proteins together. The restricted protein-surface contact may comprise a means of preserving protein structure and function in the adsorbed state. This article demonstrates several additional interesting features of PEG brushes relevant to biomaterial design. First a moderate amount of adhesive material can be buried at the base of a brush without a measurable impact on the corona density. Second, a different amount of material at the base of a brush can be rendered ineffective to capturing adhesive proteins, despite a modest compromise of the brush corona. From this will follow insight into the design of patterned biomaterial surfaces, the bioactivity of the edges of patterned features, and an understanding of how flaws in brushes compromise protein resistance or allow access to small adhesive sites.

  8. Concentration dependent model of protein-protein interaction networks

    CERN Document Server

    Zhang, Jingshan


    The scale free structure p(k)~k^{-gamma} of protein-protein interaction networks can be produced by a static physical model. We find the earlier study of deterministic threshold models with exponential fitness distributions can be generalized to explain the apparent scale free degree distribution of the physical model, and this explanation provides a generic mechanism of "scale free" networks. We predict the dependence of gamma on experimental protein concentrations. The clustering coefficient distribution of the model is also studied.

  9. Proteins aggregation and human diseases (United States)

    Hu, Chin-Kun


    Many human diseases and the death of most supercentenarians are related to protein aggregation. Neurodegenerative diseases include Alzheimer's disease (AD), Huntington's disease (HD), Parkinson's disease (PD), frontotemporallobar degeneration, etc. Such diseases are due to progressive loss of structure or function of neurons caused by protein aggregation. For example, AD is considered to be related to aggregation of Aβ40 (peptide with 40 amino acids) and Aβ42 (peptide with 42 amino acids) and HD is considered to be related to aggregation of polyQ (polyglutamine) peptides. In this paper, we briefly review our recent discovery of key factors for protein aggregation. We used a lattice model to study the aggregation rates of proteins and found that the probability for a protein sequence to appear in the conformation of the aggregated state can be used to determine the temperature at which proteins can aggregate most quickly. We used molecular dynamics and simple models of polymer chains to study relaxation and aggregation of proteins under various conditions and found that when the bending-angle dependent and torsion-angle dependent interactions are zero or very small, then protein chains tend to aggregate at lower temperatures. All atom models were used to identify a key peptide chain for the aggregation of insulin chains and to find that two polyQ chains prefer anti-parallel conformation. It is pointed out that in many cases, protein aggregation does not result from protein mis-folding. A potential drug from Chinese medicine was found for Alzheimer's disease.

  10. Viral organization of human proteins.

    Directory of Open Access Journals (Sweden)

    Stefan Wuchty


    Full Text Available Although maps of intracellular interactions are increasingly well characterized, little is known about large-scale maps of host-pathogen protein interactions. The investigation of host-pathogen interactions can reveal features of pathogenesis and provide a foundation for the development of drugs and disease prevention strategies. A compilation of experimentally verified interactions between HIV-1 and human proteins and a set of HIV-dependency factors (HDF allowed insights into the topology and intricate interplay between viral and host proteins on a large scale. We found that targeted and HDF proteins appear predominantly in rich-clubs, groups of human proteins that are strongly intertwined among each other. These assemblies of proteins may serve as an infection gateway, allowing the virus to take control of the human host by reaching protein pathways and diversified cellular functions in a pronounced and focused way. Particular transcription factors and protein kinases facilitate indirect interactions between HDFs and viral proteins. Discerning the entanglement of directly targeted and indirectly interacting proteins may uncover molecular and functional sites that can provide novel perspectives on the progression of HIV infection and highlight new avenues to fight this virus.

  11. Protein Adaptations in Archaeal Extremophiles

    Directory of Open Access Journals (Sweden)

    Christopher J. Reed


    Full Text Available Extremophiles, especially those in Archaea, have a myriad of adaptations that keep their cellular proteins stable and active under the extreme conditions in which they live. Rather than having one basic set of adaptations that works for all environments, Archaea have evolved separate protein features that are customized for each environment. We categorized the Archaea into three general groups to describe what is known about their protein adaptations: thermophilic, psychrophilic, and halophilic. Thermophilic proteins tend to have a prominent hydrophobic core and increased electrostatic interactions to maintain activity at high temperatures. Psychrophilic proteins have a reduced hydrophobic core and a less charged protein surface to maintain flexibility and activity under cold temperatures. Halophilic proteins are characterized by increased negative surface charge due to increased acidic amino acid content and peptide insertions, which compensates for the extreme ionic conditions. While acidophiles, alkaliphiles, and piezophiles are their own class of Archaea, their protein adaptations toward pH and pressure are less discernible. By understanding the protein adaptations used by archaeal extremophiles, we hope to be able to engineer and utilize proteins for industrial, environmental, and biotechnological applications where function in extreme conditions is required for activity.

  12. Protein detection system (United States)

    Fruetel, Julie A [Livermore, CA; Fiechtner, Gregory J [Bethesda, MD; Kliner, Dahv A. V. [San Ramon, CA; McIlroy, Andrew [Livermore, CA


    The present embodiment describes a miniature, microfluidic, absorption-based sensor to detect proteins at sensitivities comparable to LIF but without the need for tagging. This instrument utilizes fiber-based evanescent-field cavity-ringdown spectroscopy, in combination with faceted prism microchannels. The combination of these techniques will increase the effective absorption path length by a factor of 10.sup.3 to 10.sup.4 (to .about.1-m), thereby providing unprecedented sensitivity using direct absorption. The coupling of high-sensitivity absorption with high-performance microfluidic separation will enable real-time sensing of biological agents in aqueous samples (including aerosol collector fluids) and will provide a general method with spectral fingerprint capability for detecting specific bio-agents.

  13. JAK protein kinase inhibitors. (United States)

    Thompson, James E


    In humans, the Janus protein tyrosine kinase family (JAKs) contains four members: JAK1, JAK2, JAK3 and TYK2. JAKs phosphorylate signal transducers and activators of transcription (STATs) simultaneously with other phosphorylations required for activation, and there are several cellular mechanisms in place to inhibit JAK/STAT signaling. That one might be able to modulate selected JAK/STAT-mediated cellular signals by inhibiting JAK kinase activity to effect a positive therapeutic outcome is a tantalizing prospect, as yet incompletely realized. While current data suggest no therapeutic use for JAK1 and TYK2 inhibition, JAK2 inhibition seems a promising but not definitively tested mechanism for treatment of leukemia. More promising, however, are data indicating a possible therapeutic use of JAK3 inhibition. The restriction of the JAK3-deficient phenotype to the hematopoietic system and the resulting profound immune suppression suggest that JAK3 could be a target for immunosuppressive therapies used to prevent organ transplant rejection.

  14. Protein Polymers and Amyloids

    DEFF Research Database (Denmark)

    Risør, Michael Wulff


    that inhibits its target protease through a large conformational change but mutations compromise this function and cause premature structural collapse into hyperstable polymers. Understanding the conformational disorders at a molecular level is not only important for our general knowledge on protein folding...... of this mechanism were investigated through a series of interaction experiments. Despite a very buried location in the native structure, evidence here suggest that the C-terminal tail is labile under slightly destabilizing conditions, providing new detail to this matter. A small infectious polymer unit was also...... constructed and used to show how polymerogenic seeding and polymer propagation might happen inside the body. The locking of central structural elements during α1AT folding or in the native state represents a therapeutic strategy to prevent polymerization. Using Molecular Dynamics simulations, we identified...

  15. Protein Hormones and Immunity‡ (United States)

    Kelley, Keith W.; Weigent, Douglas A.; Kooijman, Ron


    A number of observations and discoveries over the past 20 years support the concept of important physiological interactions between the endocrine and immune systems. The best known pathway for transmission of information from the immune system to the neuroendocrine system is humoral in the form of cytokines, although neural transmission via the afferent vagus is well documented also. In the other direction, efferent signals from the nervous system to the immune system are conveyed by both the neuroendocrine and autonomic nervous systems. Communication is possible because the nervous and immune systems share a common biochemical language involving shared ligands and receptors, including neurotransmitters, neuropeptides, growth factors, neuroendocrine hormones and cytokines. This means that the brain functions as an immune-regulating organ participating in immune responses. A great deal of evidence has accumulated and confirmed that hormones secreted by the neuroendocrine system play an important role in communication and regulation of the cells of the immune system. Among protein hormones, this has been most clearly documented for prolactin (PRL), growth hormone (GH), and insulin-like growth factor-1 (IGF-I), but significant influences on immunity by thyroid stimulating hormone (TSH) have also been demonstrated. Here we review evidence obtained during the past 20 years to clearly demonstrate that neuroendocrine protein hormones influence immunity and that immune processes affect the neuroendocrine system. New findings highlight a previously undiscovered route of communication between the immune and endocrine systems that is now known to occur at the cellular level. This communication system is activated when inflammatory processes induced by proinflammatory cytokines antagonize the function of a variety of hormones, which then causes endocrine resistance in both the periphery and brain. Homeostasis during inflammation is achieved by a balance between cytokines and

  16. Novel protein-protein interactions inferred from literature context.

    Directory of Open Access Journals (Sweden)

    Herman H H B M van Haagen

    Full Text Available We have developed a method that predicts Protein-Protein Interactions (PPIs based on the similarity of the context in which proteins appear in literature. This method outperforms previously developed PPI prediction algorithms that rely on the conjunction of two protein names in MEDLINE abstracts. We show significant increases in coverage (76% versus 32% and sensitivity (66% versus 41% at a specificity of 95% for the prediction of PPIs currently archived in 6 PPI databases. A retrospective analysis shows that PPIs can efficiently be predicted before they enter PPI databases and before their interaction is explicitly described in the literature. The practical value of the method for discovery of novel PPIs is illustrated by the experimental confirmation of the inferred physical interaction between CAPN3 and PARVB, which was based on frequent co-occurrence of both proteins with concepts like Z-disc, dysferlin, and alpha-actinin. The relationships between proteins predicted by our method are broader than PPIs, and include proteins in the same complex or pathway. Dependent on the type of relationships deemed useful, the precision of our method can be as high as 90%. The full set of predicted interactions is available in a downloadable matrix and through the webtool Nermal, which lists the most likely interaction partners for a given protein. Our framework can be used for prioritizing potential interaction partners, hitherto undiscovered, for follow-up studies and to aid the generation of accurate protein interaction maps.

  17. Protein complexes predictions within protein interaction networks using genetic algorithms. (United States)

    Ramadan, Emad; Naef, Ahmed; Ahmed, Moataz


    Protein-protein interaction networks are receiving increased attention due to their importance in understanding life at the cellular level. A major challenge in systems biology is to understand the modular structure of such biological networks. Although clustering techniques have been proposed for clustering protein-protein interaction networks, those techniques suffer from some drawbacks. The application of earlier clustering techniques to protein-protein interaction networks in order to predict protein complexes within the networks does not yield good results due to the small-world and power-law properties of these networks. In this paper, we construct a new clustering algorithm for predicting protein complexes through the use of genetic algorithms. We design an objective function for exclusive clustering and overlapping clustering. We assess the quality of our proposed clustering algorithm using two gold-standard data sets. Our algorithm can identify protein complexes that are significantly enriched in the gold-standard data sets. Furthermore, our method surpasses three competing methods: MCL, ClusterOne, and MCODE in terms of the quality of the predicted complexes. The source code and accompanying examples are freely available at .

  18. Water-Protein Interactions: The Secret of Protein Dynamics

    Directory of Open Access Journals (Sweden)

    Silvia Martini


    Full Text Available Water-protein interactions help to maintain flexible conformation conditions which are required for multifunctional protein recognition processes. The intimate relationship between the protein surface and hydration water can be analyzed by studying experimental water properties measured in protein systems in solution. In particular, proteins in solution modify the structure and the dynamics of the bulk water at the solute-solvent interface. The ordering effects of proteins on hydration water are extended for several angstroms. In this paper we propose a method for analyzing the dynamical properties of the water molecules present in the hydration shells of proteins. The approach is based on the analysis of the effects of protein-solvent interactions on water protons NMR relaxation parameters. NMR relaxation parameters, especially the nonselective (R1NS and selective (R1SE spin-lattice relaxation rates of water protons, are useful for investigating the solvent dynamics at the macromolecule-solvent interfaces as well as the perturbation effects caused by the water-macromolecule interactions on the solvent dynamical properties. In this paper we demonstrate that Nuclear Magnetic Resonance Spectroscopy can be used to determine the dynamical contributions of proteins to the water molecules belonging to their hydration shells.

  19. Protein intake, body composition, and protein status following bariatric surgery. (United States)

    Andreu, Alba; Moizé, Violeta; Rodríguez, Lucía; Flores, Lilliam; Vidal, Josep


    Daily protein intake recommendations have recently been proposed for the bariatric patient. We aimed to evaluate the accomplishment of these recommendations, and the influence of protein intake (PI) on fat free mass (FFM) and protein status changes following bariatric surgery. We examined 101 consecutive patients undergoing laparoscopic Roux-in-Y gastric gypass (LGBP) or laparoscopic sleeve gastrectomy (LSG). Based on 3-day food records, PI from food and supplements were quantified at 4, 8, and 12 months after surgery. The association between PI and body composition (bioelectrical impedance), plasma albumin and pre-albumin was evaluated at all study time points. A PI protein supplementation, supplements were taken only by 63.4, 50.5, and 33.7% of the participants at 4, 8, and 12 months. However, protein supplementation was effective in helping patients to achieve the daily protein intake goal. In linear regression analysis, male gender and weight loss, but not PI, were significantly associated with loss of FFM (p protein supplementation for the achievement of the recommended daily protein intake in the bariatric patient. However, our data does not help to define a PI goal as critical in determining the FFM and protein status changes following LGBP or LSG.

  20. Protein-Protein Interaction Detection: Methods and Analysis

    Directory of Open Access Journals (Sweden)

    V. Srinivasa Rao


    Full Text Available Protein-protein interaction plays key role in predicting the protein function of target protein and drug ability of molecules. The majority of genes and proteins realize resulting phenotype functions as a set of interactions. The in vitro and in vivo methods like affinity purification, Y2H (yeast 2 hybrid, TAP (tandem affinity purification, and so forth have their own limitations like cost, time, and so forth, and the resultant data sets are noisy and have more false positives to annotate the function of drug molecules. Thus, in silico methods which include sequence-based approaches, structure-based approaches, chromosome proximity, gene fusion, in silico 2 hybrid, phylogenetic tree, phylogenetic profile, and gene expression-based approaches were developed. Elucidation of protein interaction networks also contributes greatly to the analysis of signal transduction pathways. Recent developments have also led to the construction of networks having all the protein-protein interactions using computational methods for signaling pathways and protein complex identification in specific diseases.

  1. Modular protein switches derived from antibody mimetic proteins. (United States)

    Nicholes, N; Date, A; Beaujean, P; Hauk, P; Kanwar, M; Ostermeier, M


    Protein switches have potential applications as biosensors and selective protein therapeutics. Protein switches built by fusion of proteins with the prerequisite input and output functions are currently developed using an ad hoc process. A modular switch platform in which existing switches could be readily adapted to respond to any ligand would be advantageous. We investigated the feasibility of a modular protein switch platform based on fusions of the enzyme TEM-1 β-lactamase (BLA) with two different antibody mimetic proteins: designed ankyrin repeat proteins (DARPins) and monobodies. We created libraries of random insertions of the gene encoding BLA into genes encoding a DARPin or a monobody designed to bind maltose-binding protein (MBP). From these libraries, we used a genetic selection system for β-lactamase activity to identify genes that conferred MBP-dependent ampicillin resistance to Escherichia coli. Some of these selected genes encoded switch proteins whose enzymatic activity increased up to 14-fold in the presence of MBP. We next introduced mutations into the antibody mimetic domain of these switches that were known to cause binding to different ligands. To different degrees, introduction of the mutations resulted in switches with the desired specificity, illustrating the potential modularity of these platforms. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail:

  2. Noninvasive imaging of protein-protein interactions in living animals (United States)

    Luker, Gary D.; Sharma, Vijay; Pica, Christina M.; Dahlheimer, Julie L.; Li, Wei; Ochesky, Joseph; Ryan, Christine E.; Piwnica-Worms, Helen; Piwnica-Worms, David


    Protein-protein interactions control transcription, cell division, and cell proliferation as well as mediate signal transduction, oncogenic transformation, and regulation of cell death. Although a variety of methods have been used to investigate protein interactions in vitro and in cultured cells, none can analyze these interactions in intact, living animals. To enable noninvasive molecular imaging of protein-protein interactions in vivo by positron-emission tomography and fluorescence imaging, we engineered a fusion reporter gene comprising a mutant herpes simplex virus 1 thymidine kinase and green fluorescent protein for readout of a tetracycline-inducible, two-hybrid system in vivo. By using micro-positron-emission tomography, interactions between p53 tumor suppressor and the large T antigen of simian virus 40 were visualized in tumor xenografts of HeLa cells stably transfected with the imaging constructs. Imaging protein-binding partners in vivo will enable functional proteomics in whole animals and provide a tool for screening compounds targeted to specific protein-protein interactions in living animals.

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  20. Protein (Viridiplantae): 159470305 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  1. Protein (Cyanobacteria): 493210752 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  2. Protein (Cyanobacteria): 295749 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  3. Protein (Cyanobacteria): 497073171 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  4. Protein (Cyanobacteria): 518320325 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  5. Protein (Cyanobacteria): 424444 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  6. Protein (Cyanobacteria): 515881707 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  7. Protein (Cyanobacteria): 424446 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  8. Protein (Cyanobacteria): 76081 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  9. Protein (Cyanobacteria): 497312480 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  10. Protein (Cyanobacteria): 499683197 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  11. Protein (Cyanobacteria): 499682832 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  12. Protein (Cyanobacteria): 499440544 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  13. Protein (Cyanobacteria): 24305 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  14. Protein (Cyanobacteria): 653152304 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  15. Protein (Cyanobacteria): 500464022 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  16. Protein (Cyanobacteria): 499305066 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  17. Protein (Cyanobacteria): 515856463 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  18. Protein (Cyanobacteria): 504938346 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  19. Protein (Cyanobacteria): 515858423 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)


  20. Protein (Viridiplantae): 224125616 [PGDBj - Ortholog DB

    Lifescience Database Archive (English)