CLMSVault: A Software Suite for Protein Cross-Linking Mass-Spectrometry Data Analysis and Visualization.

Science.gov (United States)

Courcelles, Mathieu; Coulombe-Huntington, Jasmin; Cossette, Émilie; Gingras, Anne-Claude; Thibault, Pierre; Tyers, Mike

2017-07-07

Protein cross-linking mass spectrometry (CL-MS) enables the sensitive detection of protein interactions and the inference of protein complex topology. The detection of chemical cross-links between protein residues can identify intra- and interprotein contact sites or provide physical constraints for molecular modeling of protein structure. Recent innovations in cross-linker design, sample preparation, mass spectrometry, and software tools have significantly improved CL-MS approaches. Although a number of algorithms now exist for the identification of cross-linked peptides from mass spectral data, a dearth of user-friendly analysis tools represent a practical bottleneck to the broad adoption of the approach. To facilitate the analysis of CL-MS data, we developed CLMSVault, a software suite designed to leverage existing CL-MS algorithms and provide intuitive and flexible tools for cross-platform data interpretation. CLMSVault stores and combines complementary information obtained from different cross-linkers and search algorithms. CLMSVault provides filtering, comparison, and visualization tools to support CL-MS analyses and includes a workflow for label-free quantification of cross-linked peptides. An embedded 3D viewer enables the visualization of quantitative data and the mapping of cross-linked sites onto PDB structural models. We demonstrate the application of CLMSVault for the analysis of a noncovalent Cdc34-ubiquitin protein complex cross-linked under different conditions. CLMSVault is open-source software (available at https://gitlab.com/courcelm/clmsvault.git ), and a live demo is available at http://democlmsvault.tyerslab.com/ .

Chemical cross-linking with thiol-cleavable reagents combined with differential mass spectrometric peptide mapping--a novel approach to assess intermolecular protein contacts

DEFF Research Database (Denmark)

Bennett, K L; Kussmann, M; Björk, P

2000-01-01

The intermolecular contact regions between monomers of the homodimeric DNA binding protein ParR and the interaction between the glycoproteins CD28 and CD80 were investigated using a strategy that combined chemical cross-linking with differential MALDI-MS analyses. ParR dimers were modified in vit...

Characterization of a bombesin receptor on Swiss mouse 3T3 cells by affinity cross-linking

International Nuclear Information System (INIS)

Sinnett-Smith, J.; Zachary, I.; Rozengurt, E.

1988-01-01

We have previously identified by chemical cross-linking a cell surface protein in Swiss 3T3 cells of apparent Mr 75,000-85,000, which may represent a major component of the receptor for peptides of the bombesin family in these cells. Because bombesin-like peptides may interact with other cell surface molecules, it was important to establish the correlation between receptor binding and functions of this complex and further characterize the Mr 75,000-85,000 cross-linked protein. Detailed time courses carried out at different temperatures demonstrated that the Mr 75,000-85,000 affinity-labelled band was the earliest cross-linked complex detected in Swiss 3T3 cells incubated with 125I-labelled gastrin-releasing peptide (125I-GRP). Furthermore, the ability of various nonradioactive bombesin agonists and antagonists to block the formation of the Mr 75,000-85,000 cross-linked complex correlated extremely well (r = 0.994) with the relative capacity of these peptides to inhibit 125I-GRP specific binding. Pretreatment with unlabelled GRP for up to 6 h caused only a slight decrease in both specific 125I-GRP binding and the affinity labelling of the Mr 75,000-85,000 protein. We also show that the cross-linked complex is a glycoprotein. First, solubilized affinity labelled Mr 75,000-85,000 complex applied to wheat germ lectin-sepharose columns was eluted by addition of 0.3 M N-acetyl-D-glucosamine. Second, treatment with endo-beta-N-acetylglucosaminidase F reduced the apparent molecular weight of the affinity-labelled band from 75,000-85,000 to 43,000, indicating the presence of N-linked oligosaccharide groups

Cross-link guided molecular modeling with ROSETTA.

Directory of Open Access Journals (Sweden)

Abdullah Kahraman

Full Text Available Chemical cross-links identified by mass spectrometry generate distance restraints that reveal low-resolution structural information on proteins and protein complexes. The technology to reliably generate such data has become mature and robust enough to shift the focus to the question of how these distance restraints can be best integrated into molecular modeling calculations. Here, we introduce three workflows for incorporating distance restraints generated by chemical cross-linking and mass spectrometry into ROSETTA protocols for comparative and de novo modeling and protein-protein docking. We demonstrate that the cross-link validation and visualization software Xwalk facilitates successful cross-link data integration. Besides the protocols we introduce XLdb, a database of chemical cross-links from 14 different publications with 506 intra-protein and 62 inter-protein cross-links, where each cross-link can be mapped on an experimental structure from the Protein Data Bank. Finally, we demonstrate on a protein-protein docking reference data set the impact of virtual cross-links on protein docking calculations and show that an inter-protein cross-link can reduce on average the RMSD of a docking prediction by 5.0 Å. The methods and results presented here provide guidelines for the effective integration of chemical cross-link data in molecular modeling calculations and should advance the structural analysis of particularly large and transient protein complexes via hybrid structural biology methods.

Supermacroporous chemically cross-linked poly(aspartic acid) hydrogels.

Science.gov (United States)

Gyarmati, Benjámin; Mészár, E Zsuzsanna; Kiss, Lóránd; Deli, Mária A; László, Krisztina; Szilágyi, András

2015-08-01

Chemically cross-linked poly(aspartic acid) (PASP) gels were prepared by a solid-liquid phase separation technique, cryogelation, to achieve a supermacroporous interconnected pore structure. The precursor polymer of PASP, polysuccinimide (PSI) was cross-linked below the freezing point of the solvent and the forming crystals acted as templates for the pores. Dimethyl sulfoxide was chosen as solvent instead of the more commonly used water. Thus larger temperatures could be utilized for the preparation and the drawback of increase in specific volume of water upon freezing could be eliminated. The morphology of the hydrogels was characterized by scanning electron microscopy and interconnectivity of the pores was proven by the small flow resistance of the gels. Compression tests also confirmed the interconnected porous structure and the complete re-swelling and shape recovery of the supermacroporous PASP hydrogels. The prepared hydrogels are of interest for several biomedical applications as scaffolding materials because of their cytocompatibility, controllable morphology and pH-responsive character. Copyright © 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Novel chemically cross-linked solid state electrolyte for dye-sensitized solar cells

International Nuclear Information System (INIS)

Yin Xiong; Tan Weiwei; Xiang Wangchun; Lin Yuan; Zhang Jingbo; Xiao Xurui; Li Xueping; Zhou Xiaowen; Fang Shibi

2010-01-01

Poly(vinylpyridine-co-ethylene glycol methyl ether methacrylate) (P(VP-co-MEOMA)) and α,ω-diiodo poly(ethylene oxide-co-propylene oxide) (I[(EO) 0.8 -co-(PO) 0.2 ] y I) were synthesized and used as chemically cross-linked precursors of the electrolyte for dye-sensitized solar cells. Meanwhile, α-iodo poly(ethylene oxide-co-propylene oxide) methyl ether (CH 3 O[(EO) 0.8 -co-(PO) 0.2 ] x I) was synthesized and added into the electrolyte as an internal plasticizer. Novel polymer electrolyte resulting from chemically cross-linked precursors was obtained by the quaterisation at 90 o C for 30 min. The characteristics for this kind of electrolyte were investigated by means of ionic conductivity, thermogravimetric and photocurrent-voltage. The ambient ionic conductivity was significantly enhanced to 2.3 x 10 -4 S cm -1 after introducing plasticizer, modified-ionic liquid. The weight loss of the solid state electrolyte at 200 o C was 1.8%, and its decomposition temperature was 287 o C. Solid state dye-sensitized solar cell based on chemically cross-linked electrolyte presented an overall conversion efficiency of 2.35% under AM1.5 irradiation (100 mW cm -2 ). The as-fabricated device maintained 88% of its initial performance at room temperature even without sealing for 30 days, showing a good stability.

A Redox-Active, Compact Molecule for Cross-Linking Amyloidogenic Peptides into Nontoxic, Off-Pathway Aggregates: In Vitro and In Vivo Efficacy and Molecular Mechanisms

Energy Technology Data Exchange (ETDEWEB)

Derrick, Jeffrey S.; Kerr, Richard A.; Nam, Younwoo; Oh, Shin Bi; Lee, Hyuck Jin; Earnest, Kaylin G.; Suh, Nayoung; Peck, Kristy L.; Ozbil, Mehmet; Korshavn, Kyle J.; Ramamoorthy, Ayyalusamy; Prabhakar, Rajeev; Merino, Edward J.; Shearer, Jason; Lee, Joo-Yong; Ruotolo, Brandon T.; Lim, Mi Hee

2015-11-25

Chemical reagents targeting and controlling amyloidogenic peptides have received much attention for helping identify their roles in the pathogenesis of protein-misfolding disorders. Herein, we report a novel strategy for redirecting amyloidogenic peptides into nontoxic, off-pathway aggregates, which utilizes redox properties of a small molecule (DMPD, N,N-dimethyl-p-phenylenediamine) to trigger covalent adduct formation with the peptide. In addition, for the first time, biochemical, biophysical, and molecular dynamics simulation studies have been performed to demonstrate a mechanistic understanding for such an interaction between a small molecule (DMPD) and amyloid-β (Aβ) and its subsequent anti-amyloidogenic activity, which, upon its transformation, generates ligand–peptide adducts via primary amine-dependent intramolecular cross-linking correlated with structural compaction. Furthermore, in vivo efficacy of DMPD toward amyloid pathology and cognitive impairment was evaluated employing 5xFAD mice of Alzheimer’s disease (AD). Such a small molecule (DMPD) is indicated to noticeably reduce the overall cerebral amyloid load of soluble Aβ forms and amyloid deposits as well as significantly improve cognitive defects in the AD mouse model. Overall, our in vitro and in vivo studies of DMPD toward Aβ with the first molecular-level mechanistic investigations present the feasibility of developing new, innovative approaches that employ redox-active compounds without the structural complexity as next-generation chemical tools for amyloid management.

Dissociation Behavior of a TEMPO-Active Ester Cross-Linker for Peptide Structure Analysis by Free Radical Initiated Peptide Sequencing (FRIPS) in Negative ESI-MS.

Science.gov (United States)

Hage, Christoph; Ihling, Christian H; Götze, Michael; Schäfer, Mathias; Sinz, Andrea

2017-01-01

We have synthesized a homobifunctional amine-reactive cross-linking reagent, containing a TEMPO (2,2,6,6-tetramethylpiperidine-1-oxy) and a benzyl group (Bz), termed TEMPO-Bz-linker, to derive three-dimensional structural information of proteins. The aim for designing this novel cross-linker was to facilitate the mass spectrometric analysis of cross-linked products by free radical initiated peptide sequencing (FRIPS). In an initial study, we had investigated the fragmentation behavior of TEMPO-Bz-derivatized peptides upon collision activation in (+)-electrospray ionization collision-induced dissociation tandem mass spectrometry (ESI-CID-MS/MS) experiments. In addition to the homolytic NO-C bond cleavage FRIPS pathway delivering the desired odd-electron product ions, an alternative heterolytic NO-C bond cleavage, resulting in even-electron product ions mechanism was found to be relevant. The latter fragmentation route clearly depends on the protonation of the TEMPO-Bz-moiety itself, which motivated us to conduct (-)-ESI-MS, CID-MS/MS, and MS 3 experiments of TEMPO-Bz-cross-linked peptides to further clarify the fragmentation behavior of TEMPO-Bz-peptide molecular ions. We show that the TEMPO-Bz-linker is highly beneficial for conducting FRIPS in negative ionization mode as the desired homolytic cleavage of the NO-C bond is the major fragmentation pathway. Based on characteristic fragments, the isomeric amino acids leucine and isoleucine could be discriminated. Interestingly, we observed pronounced amino acid side chain losses in cross-linked peptides if the cross-linked peptides contain a high number of acidic amino acids. Graphical Abstract ᅟ.

The cross linking of EPDM and NBR rubber

Directory of Open Access Journals (Sweden)

Samardžija-Jovanović Suzana

2005-01-01

Full Text Available In the process of macromolecule cross linking, the choice of type and quantity of the components and the experimental conditions are important to obtain the new cross linked materials with better mechanical and chemical characteristics. The cross linking method depends on the rubber type and structure. Intermolecular cross linking results in the formation elastomer network. The basis of the cross linking process, between ethylene propylene diene rubber (EPDM and acrylonitrile butadiene rubber (NBR, is a chemical reaction. Fillers and other additives are present in different mass ratios in the material. The exploitation properties of the cross linked materials depend on the quantity of additive in the cross linked systems.

Analytical characterisation of glutardialdehyde cross-linking products in gelatine-gum arabic complex coacervates

Energy Technology Data Exchange (ETDEWEB)

Fuguet, Elisabet [Advanced Measurement and Imaging, Unilever Food and Health Research Institute, Olivier van Noortlaan 120, 3133 AT Vlaardingen (Netherlands)], E-mail: eli.fuguet@gmail.com; Platerink, Chris van [Advanced Measurement and Imaging, Unilever Food and Health Research Institute, Olivier van Noortlaan 120, 3133 AT Vlaardingen (Netherlands); Department of Biomolecular Mass Spectrometry, Bijvoet Center for Biomolecular Research, Utrecht University, Sorbonnelaan 16, 3584 CA Utrecht (Netherlands); Janssen, Hans-Gerd [Advanced Measurement and Imaging, Unilever Food and Health Research Institute, Olivier van Noortlaan 120, 3133 AT Vlaardingen (Netherlands); van' t Hoff Institute for Molecular Sciences, University of Amsterdam, Nieuwe Achtergracht 166, 1018 WV Amsterdam (Netherlands)

2007-11-26

Encapsulates having shells of cross-linked mixtures of proteins and polysaccharides are widely used in the food and pharmaceutical industry for controlled release of actives and flavour compounds. In order to be able to predict the behaviour and the release characteristics of the microcapsules, a better understanding of the nature and extent of the cross-linking reaction is needed. Several analytical techniques were applied for the characterisation of glutardialdehyde (GDA) cross-linked encapsulates made of gelatine and gum arabic. To allow the use of sensitive, high-resolution methods such as chromatography and mass spectrometry, the sample first had to be hydrolysed. In this way, a mixture of amino acids, small peptides and the cross-link moieties was obtained. High-resolution liquid chromatography coupled to high-resolution mass spectrometry (HPLC-MS) was applied to detect possible cross-link markers through a comparison of HPLC-MS mass-chromatograms obtained for cross-linked and non-cross-linked coacervates. HPLC-MS/MS was used to identify the species responsible for the differences. Cross-linking occurred between GDA molecules and lysine and hydroxylysine {epsilon}-amino groups, and up to eight cross-link products of different nature could be identified. They included pyridinium ions and Schiff bases, and also unreacted GDA condensation products. Next, based on the insight gained in the possible chemical structures present in the cross-link markers, methods for selective labelling of these functionalities were employed to allow easier detection of related reaction products. Both liquid chromatography (LC) and gas chromatography (GC) were used in these experiments. Unfortunately, these approaches failed to detect new cross-link markers, most likely as a result of the low levels at which these are present.

Analytical characterisation of glutardialdehyde cross-linking products in gelatine-gum arabic complex coacervates

International Nuclear Information System (INIS)

Fuguet, Elisabet; Platerink, Chris van; Janssen, Hans-Gerd

2007-01-01

Encapsulates having shells of cross-linked mixtures of proteins and polysaccharides are widely used in the food and pharmaceutical industry for controlled release of actives and flavour compounds. In order to be able to predict the behaviour and the release characteristics of the microcapsules, a better understanding of the nature and extent of the cross-linking reaction is needed. Several analytical techniques were applied for the characterisation of glutardialdehyde (GDA) cross-linked encapsulates made of gelatine and gum arabic. To allow the use of sensitive, high-resolution methods such as chromatography and mass spectrometry, the sample first had to be hydrolysed. In this way, a mixture of amino acids, small peptides and the cross-link moieties was obtained. High-resolution liquid chromatography coupled to high-resolution mass spectrometry (HPLC-MS) was applied to detect possible cross-link markers through a comparison of HPLC-MS mass-chromatograms obtained for cross-linked and non-cross-linked coacervates. HPLC-MS/MS was used to identify the species responsible for the differences. Cross-linking occurred between GDA molecules and lysine and hydroxylysine ε-amino groups, and up to eight cross-link products of different nature could be identified. They included pyridinium ions and Schiff bases, and also unreacted GDA condensation products. Next, based on the insight gained in the possible chemical structures present in the cross-link markers, methods for selective labelling of these functionalities were employed to allow easier detection of related reaction products. Both liquid chromatography (LC) and gas chromatography (GC) were used in these experiments. Unfortunately, these approaches failed to detect new cross-link markers, most likely as a result of the low levels at which these are present

In-line formation of chemically cross-linked P84® co-polyimide hollow fibre membranes for H2/CO2 separation

KAUST Repository

Choi, Seung Hak

2010-12-13

In this study, chemically cross-linked asymmetric P84® co-polyimide hollow fibre membranes with enhanced separation performance were fabricated, using a dry-wet spinning process with an innovative in-line cross-linking step. The chemical modification was conducted by controlled immersion of the coagulated fibre in an aqueous 1,5-diamino-2-methylpentane (DAMP) cross-linker solution before the take-up. The effect of the cross-linker concentration on the thermal, mechanical, chemical and gas transport properties of the membranes was investigated. FT-IR/ATR analysis was used to identify the chemical changes in the polymer, while DSC analysis confirmed the changes in the Tg and the specific heat of the polymer upon cross-linking. Chemical cross-linking with a 10 wt.% aqueous DAMP solution strongly enhanced the H2/CO2 ideal selectivity from 5.3 to 16.1, while the H2 permeance of the membranes decreased from 7.06 × 10−3 to 1.01 × 10−3 m3(STP) m−2 h−1 bar−1 for a feed pressure of 1 bar at 25 °C. The increase of selectivity with decreasing permeance is somewhat higher than the slope in the Robeson upper bound, evidencing the positive effect of the cross-linking on the separation performance of the fibres. Simultaneously, the cross-linking leads to improved mechanical resistance of the membranes, which could be further enhanced by an additional thermal treatment. The produced membranes are therefore more suitable for use under harsh conditions and have a better overall performance than the uncross-linked ones.

In-line formation of chemically cross-linked P84® co-polyimide hollow fibre membranes for H2/CO2 separation

KAUST Repository

Choi, Seung Hak; Jansen, Johannes C.; Tasselli, Franco; Barbieri, Giuseppe; Drioli, Enrico

2010-01-01

In this study, chemically cross-linked asymmetric P84® co-polyimide hollow fibre membranes with enhanced separation performance were fabricated, using a dry-wet spinning process with an innovative in-line cross-linking step. The chemical

A colourimetric method for the determination of the degree of chemical cross-linking in aspartic acid-based polymer gels

Directory of Open Access Journals (Sweden)

B. Gyarmati

2015-02-01

Full Text Available A 2,4,6-trinitrobenzenesulphonic acid (TNBS-based assay is developed to determine the degree of chemical cross-linking in aspartic acid-based polymer gels. The conventional colourimetric method for the quantitative determination of amine groups is difficult to use in polymer networks; thus, an improved method is developed to analyse polymer gels swollen in dimethyl sulfoxide (DMSO. Reaction products of the derivatizing reaction are examined by NMR. The chemical stability of the reagent is increased in DMSO, and the method shows satisfactory linearity and accuracy. The degree of chemical cross-linking in the investigated gels is close to its theoretical maximum, but the conversion of the pendant amine groups to cross-linking points is strongly dependent on the feed composition of the gels.

Manufacture of polyethylene foam by electron beam cross-linking

International Nuclear Information System (INIS)

Tamai, Isamu

1976-01-01

The manufacturing process of polyethylene foam, comparison between electron beam cross-linking process and chemical cross-linking process, the electron beam irradiation technique for continuous sheets, the characteristics and uses of polyethylene foam are reviewed. The pore diameter can be controlled by selecting the dose rate, because there is strong relationship between the pore diameter and the dose rate. As the dose if higher, the foam becomes finer. The electron accelerators having large capacity show the lowest cost as the radiation source, and are applicable industrially. If the production capacity exceeds about 200 tons per month, the costs of electron beam irradiation process may be more advantageous than that of chemical process according to the circumstances. It is difficult to obtain the uniform distribution of absorption dose in the direction of thickness. General characteristics of cross-linked polyethylene foam are listed. The special feature of electron beam process is that the degree of cross-linking can be controlled arbitrarily before foaming. The products obtained by the electron beam cross-linking process have finer foams and smoother surfaces than those obtained by the chemical process, because the separation of the decomposition of foaming agents from that of cross-linking agents in the chemical cross-linking is difficult. (Iwakiri, K.)

In vivo covalent cross-linking of photon-converted rare-earth nanostructures for tumour localization and theranostics

Science.gov (United States)

Ai, Xiangzhao; Ho, Chris Jun Hui; Aw, Junxin; Attia, Amalina Binte Ebrahim; Mu, Jing; Wang, Yu; Wang, Xiaoyong; Wang, Yong; Liu, Xiaogang; Chen, Huabing; Gao, Mingyuan; Chen, Xiaoyuan; Yeow, Edwin K. L.; Liu, Gang; Olivo, Malini; Xing, Bengang

2016-01-01

The development of precision nanomedicines to direct nanostructure-based reagents into tumour-targeted areas remains a critical challenge in clinics. Chemical reaction-mediated localization in response to tumour environmental perturbations offers promising opportunities for rational design of effective nano-theranostics. Here, we present a unique microenvironment-sensitive strategy for localization of peptide-premodified upconversion nanocrystals (UCNs) within tumour areas. Upon tumour-specific cathepsin protease reactions, the cleavage of peptides induces covalent cross-linking between the exposed cysteine and 2-cyanobenzothiazole on neighbouring particles, thus triggering the accumulation of UCNs into tumour site. Such enzyme-triggered cross-linking of UCNs leads to enhanced upconversion emission upon 808 nm laser irradiation, and in turn amplifies the singlet oxygen generation from the photosensitizers attached on UCNs. Importantly, this design enables remarkable tumour inhibition through either intratumoral UCNs injection or intravenous injection of nanoparticles modified with the targeting ligand. Our strategy may provide a multimodality solution for effective molecular sensing and site-specific tumour treatment.

Acetone-Linked Peptides: A Convergent Approach for Peptide Macrocyclization and Labeling.

Science.gov (United States)

Assem, Naila; Ferreira, David J; Wolan, Dennis W; Dawson, Philip E

2015-07-20

Macrocyclization is a broadly applied approach for overcoming the intrinsically disordered nature of linear peptides. Herein, it is shown that dichloroacetone (DCA) enhances helical secondary structures when introduced between peptide nucleophiles, such as thiols, to yield an acetone-linked bridge (ACE). Aside from stabilizing helical structures, the ketone moiety embedded in the linker can be modified with diverse molecular tags by oxime ligation. Insights into the structure of the tether were obtained through co-crystallization of a constrained S-peptide in complex with RNAse S. The scope of the acetone-linked peptides was further explored through the generation of N-terminus to side chain macrocycles and a new approach for generating fused macrocycles (bicycles). Together, these studies suggest that acetone linking is generally applicable to peptide macrocycles with a specific utility in the synthesis of stabilized helices that incorporate functional tags. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Dinosaur peptides suggest mechanisms of protein survival.

Science.gov (United States)

San Antonio, James D; Schweitzer, Mary H; Jensen, Shane T; Kalluri, Raghu; Buckley, Michael; Orgel, Joseph P R O

2011-01-01

Eleven collagen peptide sequences recovered from chemical extracts of dinosaur bones were mapped onto molecular models of the vertebrate collagen fibril derived from extant taxa. The dinosaur peptides localized to fibril regions protected by the close packing of collagen molecules, and contained few acidic amino acids. Four peptides mapped to collagen regions crucial for cell-collagen interactions and tissue development. Dinosaur peptides were not represented in more exposed parts of the collagen fibril or regions mediating intermolecular cross-linking. Thus functionally significant regions of collagen fibrils that are physically shielded within the fibril may be preferentially preserved in fossils. These results show empirically that structure-function relationships at the molecular level could contribute to selective preservation in fossilized vertebrate remains across geological time, suggest a 'preservation motif', and bolster current concepts linking collagen structure to biological function. This non-random distribution supports the hypothesis that the peptides are produced by the extinct organisms and suggests a chemical mechanism for survival.

Dinosaur Peptides Suggest Mechanisms of Protein Survival

Energy Technology Data Exchange (ETDEWEB)

San Antonio, James D.; Schweitzer, Mary H.; Jensen, Shane T.; Kalluri, Raghu; Buckley, Michael; Orgel, Joseph P.R.O. (Harvard-Med); (IIT); (NCSU); (UPENN); (Manchester); (Orthovita)

2011-09-16

Eleven collagen peptide sequences recovered from chemical extracts of dinosaur bones were mapped onto molecular models of the vertebrate collagen fibril derived from extant taxa. The dinosaur peptides localized to fibril regions protected by the close packing of collagen molecules, and contained few acidic amino acids. Four peptides mapped to collagen regions crucial for cell-collagen interactions and tissue development. Dinosaur peptides were not represented in more exposed parts of the collagen fibril or regions mediating intermolecular cross-linking. Thus functionally significant regions of collagen fibrils that are physically shielded within the fibril may be preferentially preserved in fossils. These results show empirically that structure-function relationships at the molecular level could contribute to selective preservation in fossilized vertebrate remains across geological time, suggest a 'preservation motif', and bolster current concepts linking collagen structure to biological function. This non-random distribution supports the hypothesis that the peptides are produced by the extinct organisms and suggests a chemical mechanism for survival.

Preparation and properties of silk sericin/cellulose cross-linking films

Directory of Open Access Journals (Sweden)

Wang Kunyan

2017-01-01

Full Text Available Silk sericin/cellulose cross-linked films were successfully prepared using glutaraldehyde as cross-linkinger. FTIR was applied to characterize the chemical structure of films. Cross-linked silk sericin film was found the peak intensity of FTIR for cross-linked film decreased markedly compared to pure silk sericin, which indicating cross-linking reaction has been occurred. The increasing value of swelling ratio also indicated the cross-linking has been happened. The cross-linking reaction increased the thermal decomposition temperature.

The Colibactin Genotoxin Generates DNA Interstrand Cross-Links in Infected Cells

Directory of Open Access Journals (Sweden)

Nadège Bossuet-Greif

2018-03-01

Full Text Available Colibactins are hybrid polyketide-nonribosomal peptides produced by Escherichia coli, Klebsiella pneumoniae, and other Enterobacteriaceae harboring the pks genomic island. These genotoxic metabolites are produced by pks-encoded peptide-polyketide synthases as inactive prodrugs called precolibactins, which are then converted to colibactins by deacylation for DNA-damaging effects. Colibactins are bona fide virulence factors and are suspected of promoting colorectal carcinogenesis when produced by intestinal E. coli. Natural active colibactins have not been isolated, and how they induce DNA damage in the eukaryotic host cell is poorly characterized. Here, we show that DNA strands are cross-linked covalently when exposed to enterobacteria producing colibactins. DNA cross-linking is abrogated in a clbP mutant unable to deacetylate precolibactins or by adding the colibactin self-resistance protein ClbS, confirming the involvement of the mature forms of colibactins. A similar DNA-damaging mechanism is observed in cellulo, where interstrand cross-links are detected in the genomic DNA of cultured human cells exposed to colibactin-producing bacteria. The intoxicated cells exhibit replication stress, activation of ataxia-telangiectasia and Rad3-related kinase (ATR, and recruitment of the DNA cross-link repair Fanconi anemia protein D2 (FANCD2 protein. In contrast, inhibition of ATR or knockdown of FANCD2 reduces the survival of cells exposed to colibactin-producing bacteria. These findings demonstrate that DNA interstrand cross-linking is the critical mechanism of colibactin-induced DNA damage in infected cells.

Radiation cross-linked polymers: Recent developments and new applications

International Nuclear Information System (INIS)

Rouif, Sophie

2005-01-01

The purpose of the present paper is to review the innovative and recent applications of radiation cross-linking of polymers that reinforces their dimensional stability in chemically aggressive and high temperature conditions. Radiation cross-linking can be applied to a great number of plastics: thermoplastics, elastomers and thermoplastic elastomers (TPE). Some of them can cross-link on their own, some others need to be formulated with a cross-linking agent (promoter) or to be modified during their polymerization. Some results of chemical and thermomechanical characterizations of radiation cross-linked plastics based on engineering polymers will be described, and their advantages will be emphasized in relation with their applications in various sectors: pipes and cables, packaging, automotive, electrical engineering and electronics, including connectors, surface mounted devices, integrated circuits, 3D-MID technology, etc. The paper will conclude with a short review of the industrial irradiation facilities (EB facilities and gamma plants) adapted to the treatment of such various products

Supramolecular domains in mixed peptide self-assembled monolayers on gold nanoparticles.

Science.gov (United States)

Duchesne, Laurence; Wells, Geoff; Fernig, David G; Harris, Sarah A; Lévy, Raphaël

2008-09-01

Self-organization in mixed self-assembled monolayers of small molecules provides a route towards nanoparticles with complex molecular structures. Inspired by structural biology, a strategy based on chemical cross-linking is introduced to probe proximity between functional peptides embedded in a mixed self-assembled monolayer at the surface of a nanoparticle. The physical basis of the proximity measurement is a transition from intramolecular to intermolecular cross-linking as the functional peptides get closer. Experimental investigations of a binary peptide self-assembled monolayer show that this transition happens at an extremely low molar ratio of the functional versus matrix peptide. Molecular dynamics simulations of the peptide self-assembled monolayer are used to calculate the volume explored by the reactive groups. Comparison of the experimental results with a probabilistic model demonstrates that the peptides are not randomly distributed at the surface of the nanoparticle, but rather self-organize into supramolecular domains.

Dual Cross-Linked Biofunctional and Self-Healing Networks to Generate User-Defined Modular Gradient Hydrogel Constructs.

Science.gov (United States)

Wei, Zhao; Lewis, Daniel M; Xu, Yu; Gerecht, Sharon

2017-08-01

Gradient hydrogels have been developed to mimic the spatiotemporal differences of multiple gradient cues in tissues. Current approaches used to generate such hydrogels are restricted to a single gradient shape and distribution. Here, a hydrogel is designed that includes two chemical cross-linking networks, biofunctional, and self-healing networks, enabling the customizable formation of modular gradient hydrogel construct with various gradient distributions and flexible shapes. The biofunctional networks are formed via Michael addition between the acrylates of oxidized acrylated hyaluronic acid (OAHA) and the dithiol of matrix metalloproteinase (MMP)-sensitive cross-linker and RGD peptides. The self-healing networks are formed via dynamic Schiff base reaction between N-carboxyethyl chitosan (CEC) and OAHA, which drives the modular gradient units to self-heal into an integral modular gradient hydrogel. The CEC-OAHA-MMP hydrogel exhibits excellent flowability at 37 °C under shear stress, enabling its injection to generate gradient distributions and shapes. Furthermore, encapsulated sarcoma cells respond to the gradient cues of RGD peptides and MMP-sensitive cross-linkers in the hydrogel. With these superior properties, the dual cross-linked CEC-OAHA-MMP hydrogel holds significant potential for generating customizable gradient hydrogel constructs, to study and guide cellular responses to their microenvironment such as in tumor mimicking, tissue engineering, and stem cell differentiation and morphogenesis. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Chemical cross-linked polyvinyl alcohol/cellulose nanocrystal composite films with high structural stability by spraying Fenton reagent as initiator.

Science.gov (United States)

Song, Meili; Yu, Houyong; Gu, Jiping; Ye, Shounuan; Zhou, Yuwei

2018-07-01

Cross-linked polyvinyl alcohol (PVA) composite films with high structural stability were prepared by free radical copolymerization between cellulose nanocrystal (CNC) and maleic anhydride (MAH) modified PVA through spraying Fenton free radical as initiator. The influence of chemical cross-linked and physical network structure on mechanical, thermal and water absorption properties of the composite films were investigated. Compared to PVA and PVA/CNC composite film, significant improvements in the mechanical, thermal and water uptake properties of the cross-linked composite film were found. The tensile strength of the cross-linked composite film was enhanced from 23.1MPa (neat PVA film) and 32.6MPa (PVA/CNC-10%) to 42.5MPa, and the maximum thermal degradation temperature was increased from 266.8°C and 281.2°C to 366.7°C (cross-linked composite film). Besides, the water absorption was reduced from 385.9% and 220.6% to 175.7% for cross-linked composite film. It indicates that compared with physical network structure in PVA/CNC composite film, the multiple cross-linked networks showed excellent thermal stability, resistance of water swelling and structural stability at the same CNC loading level. Thus, the PVA/CNC composite film with the multiple cross-linked network shows greater property reinforcements. Copyright © 2018 Elsevier B.V. All rights reserved.

Hydrogen peroxide and ferulic acid-mediated oxidative cross-linking ...

African Journals Online (AJOL)

STORAGESEVER

2009-12-15

Dec 15, 2009 ... G250 in a 4.5:4.5:1 (v/v) mixture of deionized water, methanol and glacial acetic ... mixture of 1:1:8 (v/v) methanol, glacial acetic acid and deionized water until the ..... Cross-linking of tyrosine-containing peptides by hydrogen.

Porous Cross-Linked Polyimide-Urea Networks

Science.gov (United States)

Meador, Mary Ann B. (Inventor); Nguyen, Baochau N. (Inventor)

2015-01-01

Porous cross-linked polyimide-urea networks are provided. The networks comprise a subunit comprising two anhydride end-capped polyamic acid oligomers in direct connection via a urea linkage. The oligomers (a) each comprise a repeating unit of a dianhydride and a diamine and a terminal anhydride group and (b) are formulated with 2 to 15 of the repeating units. The subunit was formed by reaction of the diamine and a diisocyanate to form a diamine-urea linkage-diamine group, followed by reaction of the diamine-urea linkage-diamine group with the dianhydride and the diamine to form the subunit. The subunit has been cross-linked via a cross-linking agent, comprising three or more amine groups, at a balanced stoichiometry of the amine groups to the terminal anhydride groups. The subunit has been chemically imidized to yield the porous cross-linked polyimide-urea network. Also provided are wet gels, aerogels, and thin films comprising the networks, and methods of making the networks.

Mass spectrometric analysis of a UV-cross-linked protein-DNA complex: tryptophans 54 and 88 of E. coli SSB cross-link to DNA

DEFF Research Database (Denmark)

Steen, Hanno; Petersen, Jørgen; Mann, Matthias

2001-01-01

acid and peptide entities present in such heteroconjugates. Sample preparation of the peptide-nucleic acid heteroconjugates is, therefore, a crucial step in any mass spectrometry-based analytical procedure. This study demonstrates the performance of four different MS-based strategies to characterize E....... coli single-stranded DNA binding protein (SSB) that was UV-cross-linked to a 5-iodouracil containing DNA oligomer. Two methods were optimized to circumvent the need for standard liquid chromatography and gel electrophoresis, thereby dramatically increasing the overall sensitivity of the analysis...

Encoded libraries of chemically modified peptides.

Science.gov (United States)

Heinis, Christian; Winter, Greg

2015-06-01

The use of powerful technologies for generating and screening DNA-encoded protein libraries has helped drive the development of proteins as pharmaceutical ligands. However the development of peptides as pharmaceutical ligands has been more limited. Although encoded peptide libraries are typically several orders of magnitude larger than classical chemical libraries, can be more readily screened, and can give rise to higher affinity ligands, their use as pharmaceutical ligands is limited by their intrinsic properties. Two of the intrinsic limitations include the rotational flexibility of the peptide backbone and the limited number (20) of natural amino acids. However these limitations can be overcome by use of chemical modification. For example, the libraries can be modified to introduce topological constraints such as cyclization linkers, or to introduce new chemical entities such as small molecule ligands, fluorophores and photo-switchable compounds. This article reviews the chemistry involved, the properties of the peptide ligands, and the new opportunities offered by chemical modification of DNA-encoded peptide libraries. Copyright © 2015. Published by Elsevier Ltd.

Grafted Cross-Linked Polyolefin Substrates for Peptide Synthesis and Assays

DEFF Research Database (Denmark)

1999-01-01

suited for use in solid-phase biosystems, notably bioassays, such as immunoassays, DNA hybridization assays or PCR amplification. The grafted chains may bear substituents which are such that the polymer-grafted cross-linked polyolefin substrate is swellable by water or aqueous media, in other words...

Chemical reactions directed Peptide self-assembly.

Science.gov (United States)

Rasale, Dnyaneshwar B; Das, Apurba K

2015-05-13

Fabrication of self-assembled nanostructures is one of the important aspects in nanoscience and nanotechnology. The study of self-assembled soft materials remains an area of interest due to their potential applications in biomedicine. The versatile properties of soft materials can be tuned using a bottom up approach of small molecules. Peptide based self-assembly has significant impact in biology because of its unique features such as biocompatibility, straight peptide chain and the presence of different side chain functionality. These unique features explore peptides in various self-assembly process. In this review, we briefly introduce chemical reaction-mediated peptide self-assembly. Herein, we have emphasised enzymes, native chemical ligation and photochemical reactions in the exploration of peptide self-assembly.

Quaternary structure of the ATPase complex of human 26S proteasomes determined by chemical cross-linking

DEFF Research Database (Denmark)

Hartmann-Petersen, R; Tanaka, K; Hendil, K B

2001-01-01

and substrate specificity. Among the approximately 18 subunits of PA700 regulator, six are ATPases. The ATPases presumably recognize, unfold, and translocate substrates into the interior of the 26S proteasome. It is generally believed that the ATPases form a hexameric ring. By means of chemical cross......-linking, immunoprecipitation, and blotting, we have determined that the ATPases are organized in the order S6-S6'-S10b-S8-S4-S7. Additionally, we found cross-links between the ATPase S10b and the 20S proteasome subunit alpha6. Together with the previously known interaction between S8 and alpha1 and between S4 and alpha7......, these data establish the relative orientations of ATPases with respect to the 20S proteasome....

Mussel-Inspired Self-Healing Double-Cross-Linked Hydrogels by Controlled Combination of Metal Coordination and Covalent Cross-Linking

DEFF Research Database (Denmark)

Andersen, Amanda; Krogsgaard, Marie; Birkedal, Henrik

2018-01-01

a catechol-based hydrogel design that allows for the degree of oxidative covalent cross-linking to be controlled. Double cross-linked hydrogels with tunable stiffness are constructed by adding the oxidizable catechol analogue, tannic acid, to an oxidation-resistant hydrogel construct held together...... by coordination of the dihydroxy functionality of 1-(2'-carboxyethyl)-2-methyl-3-hydroxy-4-pyridinone to trivalent metal ions. By varying the amount of tannic acid, the hydrogel stiffness can be customized to a given application while retaining the self-healing capabilities of the hydrogel's coordination chemical...

The Suzuki–Miyaura Cross-Coupling as a Versatile Tool for Peptide Diversification and Cyclization

Directory of Open Access Journals (Sweden)

Tom Willemse

2017-02-01

Full Text Available The (site-selective derivatization of amino acids and peptides represents an attractive field with potential applications in the establishment of structure–activity relationships and labeling of bioactive compounds. In this respect, bioorthogonal cross-coupling reactions provide valuable means for ready access to peptide analogues with diversified structure and function. Due to the complex and chiral nature of peptides, mild reaction conditions are preferred; hence, a suitable cross-coupling reaction is required for the chemical modification of these challenging substrates. The Suzuki reaction, involving organoboron species, is appropriate given the stability and environmentally benign nature of these reactants and their amenability to be applied in (partial aqueous reaction conditions, an expected requirement upon the derivatization of peptides. Concerning the halogenated reaction partner, residues bearing halogen moieties can either be introduced directly as halogenated amino acids during solid-phase peptide synthesis (SPPS or genetically encoded into larger proteins. A reversed approach building in boron in the peptidic backbone is also possible. Furthermore, based on this complementarity, cyclic peptides can be prepared by halogenation, and borylation of two amino acid side chains present within the same peptidic substrate. Here, the Suzuki–Miyaura reaction is a tool to induce the desired cyclization. In this review, we discuss diverse amino acid and peptide-based applications explored by means of this extremely versatile cross-coupling reaction. With the advent of peptide-based drugs, versatile bioorthogonal conversions on these substrates have become highly valuable.

Investigation and characterization of receptors for pituitary adenylate cyclase-activating polypeptide in human brain by radioligand binding and chemical cross-linking

International Nuclear Information System (INIS)

Suda, K.; Smith, D.M.; Ghatei, M.A.; Murphy, J.K.; Bloom, S.R.

1991-01-01

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a novel peptide of hypothalamic origin which increases adenylate cyclase activity in rat anterior pituitary cell cultures. The 38-amino acid peptide shows a close sequence homology to vasoactive intestinal peptide (VIP). Binding sites for PACAP in membranes from postmortem human brain tissue were studied using [ 125 I]PACAP27 as the radioligand. High specific binding sites (amount of specific binding measured at 0.25 nM [ 125 I]PACAP27 in femtomoles per mg protein +/- SEM; n = 4) were present in hypothalamus (344.5 +/- 13.0), brain stem (343.0 +/- 29.3), cerebellum (292.0 +/- 21.1), cortex (259.6 +/- 19.8), and basal ganglia (259.2 +/- 50.3). Specific binding sites in pituitary, although present, were less abundant (35.0 +/- 8.9). Binding of [ 125 I]PACAP27 was reversible and time, pH, and temperature dependent. Despite the homology with VIP, VIP was a poor inhibitor of [ 125 I]PACAP27 binding (IC50, greater than 1 microM) compared with PACAP27 (IC50, 0.5-1.3 nM) and PACAP38 (IC50, 0.2-1.3 nM). Scatchard plots of [ 125 I]PACAP27 binding showed the presence of both high and lower affinity sites. Chemical cross-linking of PACAP-binding sites revealed that [ 125 I]PACAP27 was bound to polypeptide chains of 67,000 and 48,000 mol wt. Thus, we have demonstrated the presence of PACAP-specific receptors in human brain which are not VIP receptors. This opens the possibility of PACAP functioning as a novel neurotransmitter/neuromodulator in human brain

Light-induced cross-linking and post-cross-linking modification of polyglycidol.

Science.gov (United States)

Marquardt, F; Bruns, M; Keul, H; Yagci, Y; Möller, M

2018-02-08

The photoinduced radical generation process has received renewed interest due to its economic and ecological appeal. Herein the light-induced cross-linking of functional polyglycidol and its post-cross-linking modification are presented. Linear polyglycidol was first functionalized with a tertiary amine in a two-step reaction. Dimethylaminopropyl functional polyglycidol was cross-linked in a UV-light mediated reaction with camphorquinone as a type II photoinitiator. The cross-linked polyglycidol was further functionalized by quaternization with various organoiodine compounds. Aqueous dispersions of the cross-linked polymers were investigated by means of DLS and zeta potential measurements. Polymer films were evaluated by DSC and XPS.

Angiotensin I-Converting Enzyme Inhibitor Derived from Cross-Linked Oyster Protein

Directory of Open Access Journals (Sweden)

Cheng-Liang Xie

2014-01-01

Full Text Available Following cross-linking by microbial transglutaminase, modified oyster proteins were hydrolyzed to improve inhibitory activity against angiotensin-converting enzyme (ACE inhibitory activity with the use of a single protease, or a combination of six proteases. The oyster hydrolysate with the lowest 50% ACE inhibitory concentration (IC50 of 0.40 mg/mL was obtained by two-step hydrolysis of the cross-linked oyster protein using Protamex and Neutrase. Five ACE inhibitory peptides were purified from the oyster hydrolysate using a multistep chromatographic procedure comprised of ion-exchange, size exclusion, and reversed-phase liquid chromatography. Their sequences were identified as TAY, VK, KY, FYN, and YA, using automated Edman degradation and mass spectrometry. These peptides were synthesized, and their IC50 values were measured to be 16.7, 29.0, 51.5, 68.2, and 93.9 μM, respectively. Toxicity of the peptides on the HepG2 cell line was not detected. The oyster hydrolysate also significantly decreased the systolic blood pressure of spontaneously hypertensive rats (SHR. The antihypertensive effect of the oyster hydrolysate on SHR was rapid and long-lasting, compared to commercially obtained sardine hydrolysate. These results suggest that the oyster hydrolysate could be a source of effective nutraceuticals against hypertension.

Oligomerisation of Synaptobrevin-2 Studied by Native Mass Spectrometry and Chemical Cross-Linking

Science.gov (United States)

Wittig, Sabine; Haupt, Caroline; Hoffmann, Waldemar; Kostmann, Susann; Pagel, Kevin; Schmidt, Carla

2018-06-01

Synaptobrevin-2 is a key player in signal transmission in neurons. It forms, together with SNAP25 and Syntaxin-1A, the neuronal soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex and mediates exocytosis of synaptic vesicles with the pre-synaptic membrane. While Synaptobrevin-2 is part of a four-helix bundle in this SNARE complex, it is natively unstructured in the absence of lipids or other SNARE proteins. Partially folded segments, presumably SNARE complex formation intermediates, as well as formation of Synaptobrevin-2 dimers and oligomers, were identified in previous studies. Here, we employ three Synaptobrevin-2 variants—the full-length protein Syb(1-116), the soluble, cytosolic variant Syb(1-96) as well as a shorter version Syb(49-96) containing structured segments but omitting a trigger site for SNARE complex formation—to study oligomerisation in the absence of interaction partners or when incorporated into the lipid bilayer of liposomes. Combining native mass spectrometry with chemical cross-linking, we find that the truncated versions show increased oligomerisation. Our findings from both techniques agree well and confirm the presence of oligomers in solution while membrane-bound Synaptobrevin-2 is mostly monomeric. Using ion mobility mass spectrometry, we could further show that lower charge states of Syb(49-96) oligomers, which most likely represent solution structures, follow an isotropic growth curve suggesting that they are intrinsically disordered. From a technical point of view, we show that the combination of native ion mobility mass spectrometry with chemical cross-linking is well-suited for the analysis of protein homo-oligomers. [Figure not available: see fulltext.

Bone induction by biomimetic PLGA copolymer loaded with a novel synthetic RADA16-P24 peptide in vivo

International Nuclear Information System (INIS)

Pan, Haitao; Hao, Shaofei; Zheng, Qixin; Li, Jingfeng; Zheng, Jin; Hu, Zhilei; Yang, Shuhua; Guo, Xiaodong; Yang, Qin

2013-01-01

Bone morphogenetic protein-2 (BMP-2) is a key bone morphogenetic protein, and poly(lactic-co-glycolic acid) (PLGA) has been widely used as scaffold for clinical use to carry treatment protein. In the previous studies, we have synthesized BMP-2-related peptide (P24) and found its capacity of inducing bone regeneration. In this research, we have synthesized a new amphiphilic peptide Ac-RADA RADA RADA RADA S[PO4]KIPKASSVPTELSAISTLYLDDD-CONH2 (RADA16-P24) with an assembly peptide RADA16-Ion the P24 item of BMP2 to form divalent ion-induced gelatin. Two methods of physisorption and chemical cross-linking were used to bind RADA16-P24 onto the surface of the copolymer PLGA to synthesize RADA16-P24–PLGA, and its capacity of attaching bone marrow stromal cells (BMSCs) was evaluated in vitro and inducing ectopic bone formation was examined in vivo. In vitro our results demonstrated that RADA16-P24–PLGA copolymer prepared by physisorbing or prepared by chemical cross-linking had a peptide binding rate of (2.0180 ± 0.5296)% or (10.0820 ± 0.8405)% respectively (P < 0.05). In addition the BMSCs proliferated vigorously in the RADA16-P24–PLGA biomaterials. Significantly the percentage of BMSCs attached to RADA16-P24–PLGA composite prepared by chemical cross-linking and physisorbing were (71.4 ± 7.5) % or (46.7 ± 5.8) % (P < 0.05). The in vivo study showed that RADA16-P24–PLGA chemical cross-linking could better induce ectopic bone formation compared with RADA16-P24–PLGA physisorbing and PLGA. It is concluded that the PLGA copolymer is a good RADA16-P24 carrier. This novel RADA16-P24–PLGA composite has strong osteogenic capability. - Highlights: • We have synthesized a new RADA16-P24 amphiphilic peptide. • It is an assembly peptide RADA16-Ion the P24 to form divalent ion-induced gelatin. • RADA16-P24/PLGA could better induce etopia osteogenesis compared with PLGA. • RADA16-P24–PLGA has strong osteogenic capability

Bone induction by biomimetic PLGA copolymer loaded with a novel synthetic RADA16-P24 peptide in vivo

Energy Technology Data Exchange (ETDEWEB)

Pan, Haitao; Hao, Shaofei [Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022 (China); Zheng, Qixin, E-mail: zheng-qx@163.com [Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022 (China); Li, Jingfeng [Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022 (China); Department of Orthopedics, Zhongnan Hospital of Wuhan University, Wuhan 430071 (China); Zheng, Jin; Hu, Zhilei; Yang, Shuhua; Guo, Xiaodong [Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022 (China); Yang, Qin [Advanced Biomaterials and Tissue Engineering Center, Huazhong University of Science and Technology, Wuhan 430074 (China)

2013-08-01

Bone morphogenetic protein-2 (BMP-2) is a key bone morphogenetic protein, and poly(lactic-co-glycolic acid) (PLGA) has been widely used as scaffold for clinical use to carry treatment protein. In the previous studies, we have synthesized BMP-2-related peptide (P24) and found its capacity of inducing bone regeneration. In this research, we have synthesized a new amphiphilic peptide Ac-RADA RADA RADA RADA S[PO4]KIPKASSVPTELSAISTLYLDDD-CONH2 (RADA16-P24) with an assembly peptide RADA16-Ion the P24 item of BMP2 to form divalent ion-induced gelatin. Two methods of physisorption and chemical cross-linking were used to bind RADA16-P24 onto the surface of the copolymer PLGA to synthesize RADA16-P24–PLGA, and its capacity of attaching bone marrow stromal cells (BMSCs) was evaluated in vitro and inducing ectopic bone formation was examined in vivo. In vitro our results demonstrated that RADA16-P24–PLGA copolymer prepared by physisorbing or prepared by chemical cross-linking had a peptide binding rate of (2.0180 ± 0.5296)% or (10.0820 ± 0.8405)% respectively (P < 0.05). In addition the BMSCs proliferated vigorously in the RADA16-P24–PLGA biomaterials. Significantly the percentage of BMSCs attached to RADA16-P24–PLGA composite prepared by chemical cross-linking and physisorbing were (71.4 ± 7.5) % or (46.7 ± 5.8) % (P < 0.05). The in vivo study showed that RADA16-P24–PLGA chemical cross-linking could better induce ectopic bone formation compared with RADA16-P24–PLGA physisorbing and PLGA. It is concluded that the PLGA copolymer is a good RADA16-P24 carrier. This novel RADA16-P24–PLGA composite has strong osteogenic capability. - Highlights: • We have synthesized a new RADA16-P24 amphiphilic peptide. • It is an assembly peptide RADA16-Ion the P24 to form divalent ion-induced gelatin. • RADA16-P24/PLGA could better induce etopia osteogenesis compared with PLGA. • RADA16-P24–PLGA has strong osteogenic capability.

An Investigation on Rheology of Peroxide Cross-linking of Low Density Polyethylene

DEFF Research Database (Denmark)

Ghasemi, Ismaeil; Rasmussen, Henrik K.; Szabo, Peter

2005-01-01

One of the most important post-reactor modifications of polyethylene is cross-linking. It improves some properties of polyethylene such as environmental stress cracking resistance, chemical and abrasion resistance, and service temperature. In this study, the effect of peroxide cross-linking on th......One of the most important post-reactor modifications of polyethylene is cross-linking. It improves some properties of polyethylene such as environmental stress cracking resistance, chemical and abrasion resistance, and service temperature. In this study, the effect of peroxide cross......-linking on the rheological behaviour of low density polyethylene was investigated by using a combination of creep test and differential scanning calorimeter (DSC) in isotherm condition. The used peroxide was di-cumyl peroxide and its concentration was 2 wt%. The experiments were carried out at 150,160, and 170 degrees C...

LFQProfiler and RNP(xl): Open-Source Tools for Label-Free Quantification and Protein-RNA Cross-Linking Integrated into Proteome Discoverer.

Science.gov (United States)

Veit, Johannes; Sachsenberg, Timo; Chernev, Aleksandar; Aicheler, Fabian; Urlaub, Henning; Kohlbacher, Oliver

2016-09-02

Modern mass spectrometry setups used in today's proteomics studies generate vast amounts of raw data, calling for highly efficient data processing and analysis tools. Software for analyzing these data is either monolithic (easy to use, but sometimes too rigid) or workflow-driven (easy to customize, but sometimes complex). Thermo Proteome Discoverer (PD) is a powerful software for workflow-driven data analysis in proteomics which, in our eyes, achieves a good trade-off between flexibility and usability. Here, we present two open-source plugins for PD providing additional functionality: LFQProfiler for label-free quantification of peptides and proteins, and RNP(xl) for UV-induced peptide-RNA cross-linking data analysis. LFQProfiler interacts with existing PD nodes for peptide identification and validation and takes care of the entire quantitative part of the workflow. We show that it performs at least on par with other state-of-the-art software solutions for label-free quantification in a recently published benchmark ( Ramus, C.; J. Proteomics 2016 , 132 , 51 - 62 ). The second workflow, RNP(xl), represents the first software solution to date for identification of peptide-RNA cross-links including automatic localization of the cross-links at amino acid resolution and localization scoring. It comes with a customized integrated cross-link fragment spectrum viewer for convenient manual inspection and validation of the results.

Chemical cross-linking of polypropylenes towards new shape memory polymers.

Science.gov (United States)

Raidt, Thomas; Hoeher, Robin; Katzenberg, Frank; Tiller, Joerg C

2015-04-01

In this work, syndiotactic polypropylene (sPP) as well as isotactic polypropylene (iPP) are cross-linked to gain a shape memory effect. Both prepared PP networks exhibit maximum strains of 700%, stored strains of up to 680%, and recoveries of nearly 100%. While x-iPP is stable for many cycles, x-sPP ruptures after the first shape-memory cycle. It is shown by wide-angle X-ray scattering (WAXS) experiments that cross-linked iPP exhibits homoepitaxy in the temporary, stretched shape but in contrast to previous reports it contains a higher amount of daughter than mother crystals. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

[Biophysical principles of collagen cross-linking].

Science.gov (United States)

Spörl, E; Raiskup-Wolf, F; Pillunat, L E

2008-02-01

The reduced mechanical stability of the cornea in keratoconus or in keratectasia after Lasik may be increased by photooxidative cross-linking of corneal collagen. The biophysical principles are compiled for the safe and effective application of this new treatment method. The setting of the therapy parameters should be elucidated from the absorption behaviour of the cornea. The safety of the method for the endothelium cells and the lens will be discussed. The induced cross-links are shown to be the result of changes in the physico-chemical properties of the cornea. To reach a high absorption of the irradiation energy in the cornea, riboflavin of a concentration of 0.1% and UV light of a wavelength of 370 nm, corresponding to the relative maximum of absorption of riboflavin, were used. An irradiance of 3 mW/cm(2) and an irradiation time of 30 min lead to an increase of the mechanical stiffness. The endothelium cells will be protected due to the high absorption within the cornea, that means the damaging threshold of the endothelium cells will not be reached in a 400 microm thick stroma. As evidence for cross-links we can consider the increase of the biomechanical stiffness, the increased resistance against enzymatic degradation, a higher shrinkage temperature, a lower swelling rate and an increased diameter of collagen fibres. The therapy parameters were tested experimentally and have been proven clinically in the corneal collagen cross-linking. These parameters should be respected to reach a safe cross-linking effect without damage of the adjacent tissues.

Reactive electrospinning and biodegradation of cross-linked methacrylated polycarbonate nanofibers

Energy Technology Data Exchange (ETDEWEB)

Wu Ruizhi; Zhang Jianfeng; Fan Yuwei; Xu Xiaoming [Department of Comprehensive Dentistry and Biomaterials, Louisiana State University Health Sciences Center, 1100 Florida Avenue, Box 137, New Orleans, LA 70119 (United States); Stoute, Diana; Lallier, Thomas, E-mail: xxu@lsuhsc.edu [Department of Cell Biology and Anatomy, Louisiana State University Health Science Center, 1100 Florida Avenue, Box 137, New Orleans, LA 70119 (United States)

2011-06-15

The objectives of this study were to fabricate cross-linked biodegradable polycarbonate nanofibers and to investigate their biodegradability by different enzymes. Poly(2,3-dihydroxycarbonate) was synthesized from naturally occurring l-tartaric acid. The hydroxyl groups on the functional polycarbonate were converted to methacrylate groups to enable the polymer to cross-link under UV irradiation. Smooth cross-linked methacrylated polycarbonate nanofibers (300-1800 nm) were fabricated by a reactive electrospinning process with in situ UV radiation from a mixed solution of linear methacrylated polycarbonate (MPC) and poly(ethylene oxide) (PEO) (MPC:PEO = 9:1) in methanol/chloroform (50/50). These cross-linked nanofibers have shown excellent solvent resistance and their solubility decreases with increasing degree of cross-linking. The thermal properties of linear and cross-linked polycarbonate nanofibers were investigated by differential scanning calorimetry and thermogravimetric analysis. The cross-linked polycarbonate nanofibers show no melting point below 200 {sup 0}C and their decomposition temperature increases with increasing cross-linking degree. Their biodegradation products by five different enzymes were analyzed using liquid chromatography-mass spectrometry (LC-MS). The biodegradability of the polycarbonate nanofibers decreases with increasing cross-linking degree. These nanofibers were found to support human fibroblast survival and to promote cell attachment. This study demonstrates that cross-linked biodegradable polycarbonate nanofibers with different chemical properties and biodegradability can be fabricated using the novel reactive electrospinning technology to meet the needs of different biomedical applications.

Advancements in mass spectrometry for biological samples: Protein chemical cross-linking and metabolite analysis of plant tissues

Energy Technology Data Exchange (ETDEWEB)

Klein, Adam [Iowa State Univ., Ames, IA (United States)

2015-01-01

This thesis presents work on advancements and applications of methodology for the analysis of biological samples using mass spectrometry. Included in this work are improvements to chemical cross-linking mass spectrometry (CXMS) for the study of protein structures and mass spectrometry imaging and quantitative analysis to study plant metabolites. Applications include using matrix-assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI) to further explore metabolic heterogeneity in plant tissues and chemical interactions at the interface between plants and pests. Additional work was focused on developing liquid chromatography-mass spectrometry (LC-MS) methods to investigate metabolites associated with plant-pest interactions.

A structural and kinetic study on myofibrils prevented from shortening by chemical cross-linking.

Science.gov (United States)

Herrmann, C; Sleep, J; Chaussepied, P; Travers, F; Barman, T

1993-07-20

In previous work, we studied the early steps of the Mg(2+)-ATPase activity of Ca(2+)-activated myofibrils [Houadjeto, M., Travers, F., & Barman, T. (1992) Biochemistry 31, 1564-1569]. The myofibrils were free to contract, and the results obtained refer to the ATPase cycle of myofibrils contracting with no external load. Here we studied the ATPase of myofibrils contracting isometrically. To prevent shortening, we cross-linked them with 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide (EDC). SDS-PAGE and Western blot analyses showed that the myosin rods were extensively cross-linked and that 8% of the myosin heads were cross-linked to the thin filament. The transient kinetics of the cross-linked myofibrils were studied in 0.1 M potassium acetate, pH 7.4 and 4 degrees C, by the rapid-flow quench method. The ATP binding steps were studied by the cold ATP chase and the cleavage and release of products steps by the Pi burst method. In Pi burst experiments, the sizes of the bursts were equal within experimental error to the ATPase site concentrations (as determined by the cold ATP chase methods) for both cross-linked (isometric) and un-cross-linked (isotonic) myofibrils. This shows that in both cases the rate-limiting step is after the cleavage of ATP. When cross-linked, the kcat of Ca(2+)-activated myofibrils was reduced from 1.7 to 0.8 s-1. This is consistent with the observation that fibers shortening at moderate velocity have a higher ATPase activity than isometric fibers.(ABSTRACT TRUNCATED AT 250 WORDS)

Computational investigation of kinetics of cross-linking reactions in proteins: importance in structure prediction.

Science.gov (United States)

Bandyopadhyay, Pradipta; Kuntz, Irwin D

2009-01-01

The determination of protein structure using distance constraints is a new and promising field of study. One implementation involves attaching residues of a protein using a cross-linking agent, followed by protease digestion, analysis of the resulting peptides by mass spectroscopy, and finally sequence threading to detect the protein folds. In the present work, we carry out computational modeling of the kinetics of cross-linking reactions in proteins using the master equation approach. The rate constants of the cross-linking reactions are estimated using the pKas and the solvent-accessible surface areas of the residues involved. This model is tested with fibroblast growth factor (FGF) and cytochrome C. It is consistent with the initial experimental rate data for individual lysine residues for cytochrome C. Our model captures all observed cross-links for FGF and almost 90% of the observed cross-links for cytochrome C, although it also predicts cross-links that were not observed experimentally (false positives). However, the analysis of the false positive results is complicated by the fact that experimental detection of cross-links can be difficult and may depend on specific experimental conditions such as pH, ionic strength. Receiver operator characteristic plots showed that our model does a good job in predicting the observed cross-links. Molecular dynamics simulations showed that for cytochrome C, in general, the two lysines come closer for the observed cross-links as compared to the false positive ones. For FGF, no such clear pattern exists. The kinetic model and MD simulation can be used to study proposed cross-linking protocols.

A general method for targeted quantitative cross-linking mass spectrometry

Science.gov (United States)

Chemical cross-linking mass spectrometry (XL-MS) provides protein structural information by identifying covalently linked proximal amino acid residues on protein surfaces. The information gained by this technique is complementary to other structural biology methods such as x-ray crystallography, NM...

Molecular Model for HNBR with Tunable Cross-Link Density.

Science.gov (United States)

Molinari, N; Khawaja, M; Sutton, A P; Mostofi, A A

2016-12-15

We introduce a chemically inspired, all-atom model of hydrogenated nitrile butadiene rubber (HNBR) and assess its performance by computing the mass density and glass-transition temperature as a function of cross-link density in the structure. Our HNBR structures are created by a procedure that mimics the real process used to produce HNBR, that is, saturation of the carbon-carbon double bonds in NBR, either by hydrogenation or by cross-linking. The atomic interactions are described by the all-atom "Optimized Potentials for Liquid Simulations" (OPLS-AA). In this paper, first, we assess the use of OPLS-AA in our models, especially using NBR bulk properties, and second, we evaluate the validity of the proposed model for HNBR by investigating mass density and glass transition as a function of the tunable cross-link density. Experimental densities are reproduced within 3% for both elastomers, and qualitatively correct trends in the glass-transition temperature as a function of monomer composition and cross-link density are obtained.

Preparation of Nanocellulose Reinforced Chitosan Films, Cross-Linked by Adipic Acid.

Science.gov (United States)

Falamarzpour, Pouria; Behzad, Tayebeh; Zamani, Akram

2017-02-13

Adipic acid, an abundant and nontoxic compound, was used to dissolve and cross-link chitosan. After the preparation of chitosan films through casting technique, the in situ amidation reaction was performed at 80-100 °C as verified by Fourier transform infrared (FT-IR). The reaction was accompanied by the release of water which was employed to investigate the reaction kinetics. Accordingly, the reaction rate followed the first-order model and Arrhenius equation, and the activation energy was calculated to be 18 kJ/mol. Furthermore, the mechanical properties of the chitosan films were comprehensively studied. First, optimal curing conditions (84 °C, 93 min) were introduced through a central composite design. In order to evaluate the effects of adipic acid, the mechanical properties of physically cross-linked (uncured), chemically cross-linked (cured), and uncross-linked (prepared by acetic acid) films were compared. The use of adipic acid improved the tensile strength of uncured and chemically cross-linked films more than 60% and 113%, respectively. Finally, the effect of cellulose nanofibrils (CNFs) on the mechanical performance of cured films, in the presence of glycerol as a plasticizer, was investigated. The plasticized chitosan films reinforced by 5 wt % CNFs showed superior properties as a promising material for the development of chitosan-based biomaterials.

Cross-linked self-assembled micelle based nanosensor for intracellular pH measurements

DEFF Research Database (Denmark)

Ek, Pramod Kumar; Søndergaard, Rikke Vicki; Windschiegl, Barbara

2014-01-01

A micelle based nanosensor was synthesized and investigated as a ratiometric pH sensor for use in measurements in living cells by fluorescent microscopy. The nanosensor synthesis was based on self-assembly of an amphiphilic triblock copolymer, which was chemically cross-linked after micelle......-linked by an amidation reaction using 3,6,9-trioxaundecandioic acid cross-linker. The cross-linked micelle was functionalized with two pH sensitive fluorophores and one reference fluorophore, which resulted in a highly uniform ratiometric pH nanosensor with a diameter of 29 nm. The use of two sensor fluorophores...... provided a sensor with a very broad measurement range that seems to be influenced by the chemical design of the sensor. Cell experiments show that the sensor is capable of monitoring the pH distributions in HeLa cells....

Effects of Thermal Cross-Linking on the Structure and Property of Asymmetric Membrane Prepared from the Polyacrylonitrile

Directory of Open Access Journals (Sweden)

Xin Jin

2018-05-01

Full Text Available Improving the thermal and chemical stabilities of classical polymer membranes will be beneficial to extend their applications in the high temperature or aggressive environment. In this work, the asymmetric ultrafiltration membranes prepared from the polyacrylonitrile (PAN were used to fabricate the cross-linking asymmetric (CLA PAN membranes via thermal cross-linking in air to improve their thermal and chemical stabilities. The effects of thermal cross-linking parameters such as temperature and holding time on the structure, gas separation performance, thermal and chemical stabilities of PAN membranes were investigated by Fourier transform infrared spectroscopy (FTIR, X-ray photoelectron spectroscopy (XPS, positron annihilation lifetime spectroscopy (PALS, scanning electron microscopy (SEM, thermogravimetic analysis (TGA and gas permeation test. The thermal cross-linking significantly influences the chemical structure, microstructure and pore structure of PAN membrane. During the thermal cross-linking, the shrinkage of membrane and coalescence or collapse of pore and microstructure make large pores diminish, small pores disappear and pore volumes reduce. The gas permeances of CLA-PAN membranes increase as the increasing of cross-linking temperature and holding time due to the volatilization of small molecules. The CLA-PAN membranes demonstrate excellent thermal and chemical stabilities and present good prospects for application in ultrafiltration for water treatment and for use as a substrate for nanofiltration or gas separation with an aggressive and demanding environment.

Synthesis and characterization of N-hydroxysuccinimide ester chemical affinity derivatives of asialoorosomucoid that covalently cross-link to galactosyl receptors on isolated rat hepatocytes

International Nuclear Information System (INIS)

Herzig, M.C.S.; Weigel, P.H.

1989-01-01

The authors have developed chemical affinity reagents for the hepatic galactosyl receptor. Asialoorosomucoid (ASOR) was derivatized with five homobifunctional N-hydroxysuccinimide (NHS) ester cross-linkers. NHS/ASOR derivatives were synthesized, purified, and applied within 10 min to isolated rat hepatocytes at 4 degree C. Specific binding of these 125 I-labeled derivatives was ∼90% in the presence of either EGTA or excess ASOR. Specific cross-linking assessed by the resistance of specifically bound NHS/ 125 I-ASOR to release by EGTA, was 50-75% of the specifically bound ligand. The extent of specific cross-linking correlated with the average number of NHS groups per ASOR and was controlled by varying the molar ratio of cross-linker to ASOR during the synthesis. After being cross-linked with any of the NHS/ 125 I-ASOR derivatives, cells were washed with EGTA, solubilized in Triton X-100, and analyzed by SDA-PAGE and autoradiography. They conclude that all three receptor subunits can cross-link to ligand. They propose a model in which the native receptor is a heterohexamer composed of four subunits of RHL 1 and two subunits of RHL 2 and/or RHL 3

Effects of Supercritical CO 2 Conditioning on Cross-Linked Polyimide Membranes

KAUST Repository

Kratochvil, Adam M.

2010-05-25

The effects of supercritical CO2 (scCO2) conditioning on high-performance cross-linked polyimide membranes is examined through gas permeation and sorption experiments. Under supercritical conditions, the cross-linked polymers do not exhibit a structural reorganization of the polymer matrix that was observed in the non-cross-linkable, free acid polymer. Pure gas permeation isotherms and mixed gas permeabilities and selectivities show the cross-linked polymers to be much more stable to scCO2 conditioning than the free acid polymer. In fact, following scCO2 conditioning, the mixed gas CO2 permeabilities of the cross-linked polymers increased while the CO2/CH4 separation factors remained relatively unchanged. This response highlights the stability and high performance of these cross-linked membranes in aggressive environments. In addition, this response reveals the potential for the preconditioning of cross-linked polymer membranes to enhance productivity without sacrificing efficiency in practical applications which, in effect, provides another tool to \\'tune\\' membrane properties for a given separation. Finally, the dual mode model accurately describes the sorption and dilation characteristics of the cross-linked polymers. The changes in the dual mode sorption model parameters before and after the scCO2 exposure also provide insights into the alterations in the different glassy samples due to the cross-linking and scCO2 exposure. © 2010 American Chemical Society.

Cα chemical shift tensors in helical peptides by dipolar-modulated chemical shift recoupling NMR

International Nuclear Information System (INIS)

Yao Xiaolan; Yamaguchi, Satoru; Hong Mei

2002-01-01

The Cα chemical shift tensors of proteins contain information on the backbone conformation. We have determined the magnitude and orientation of the Cα chemical shift tensors of two peptides with α-helical torsion angles: the Ala residue in G*AL (φ=-65.7 deg., ψ=-40 deg.), and the Val residue in GG*V (φ=-81.5 deg., ψ=-50.7 deg.). The magnitude of the tensors was determined from quasi-static powder patterns recoupled under magic-angle spinning, while the orientation of the tensors was extracted from Cα-Hα and Cα-N dipolar modulated powder patterns. The helical Ala Cα chemical shift tensor has a span of 36 ppm and an asymmetry parameter of 0.89. Its σ 11 axis is 116 deg. ± 5 deg. from the Cα-Hα bond while the σ 22 axis is 40 deg. ± 5 deg. from the Cα-N bond. The Val tensor has an anisotropic span of 25 ppm and an asymmetry parameter of 0.33, both much smaller than the values for β-sheet Val found recently (Yao and Hong, 2002). The Val σ 33 axis is tilted by 115 deg. ± 5 deg. from the Cα-Hα bond and 98 deg. ± 5 deg. from the Cα-N bond. These represent the first completely experimentally determined Cα chemical shift tensors of helical peptides. Using an icosahedral representation, we compared the experimental chemical shift tensors with quantum chemical calculations and found overall good agreement. These solid-state chemical shift tensors confirm the observation from cross-correlated relaxation experiments that the projection of the Cα chemical shift tensor onto the Cα-Hα bond is much smaller in α-helices than in β-sheets

Mechanical Strength Improvements of Carbon Nanotube Threads through Epoxy Cross-Linking

Directory of Open Access Journals (Sweden)

Qingyue Yu

2016-01-01

Full Text Available Individual Carbon Nanotubes (CNTs have a great mechanical strength that needs to be transferred into macroscopic fiber assemblies. One approach to improve the mechanical strength of the CNT assemblies is by creating covalent bonding among their individual CNT building blocks. Chemical cross-linking of multiwall CNTs (MWCNTs within the fiber has significantly improved the strength of MWCNT thread. Results reported in this work show that the cross-linked thread had a tensile strength six times greater than the strength of its control counterpart, a pristine MWCNT thread (1192 MPa and 194 MPa, respectively. Additionally, electrical conductivity changes were observed, revealing 2123.40 S·cm−1 for cross-linked thread, and 3984.26 S·cm−1 for pristine CNT thread. Characterization suggests that the obtained high tensile strength is due to the cross-linking reaction of amine groups from ethylenediamine plasma-functionalized CNT with the epoxy groups of the cross-linking agent, 4,4-methylenebis(N,N-diglycidylaniline.

PH dependent adhesive peptides

Science.gov (United States)

Tomich, John; Iwamoto, Takeo; Shen, Xinchun; Sun, Xiuzhi Susan

2010-06-29

A novel peptide adhesive motif is described that requires no receptor or cross-links to achieve maximal adhesive strength. Several peptides with different degrees of adhesive strength have been designed and synthesized using solid phase chemistries. All peptides contain a common hydrophobic core sequence flanked by positively or negatively charged amino acids sequences.

3.3. Sorption activity of cross-linked polymers of ethynyl-piperidol

International Nuclear Information System (INIS)

Khalikov, D.Kh.

2012-01-01

The sorption activity of cross-linked polymers of ethynyl-piperidol was studied. The bilirubin sorption was studied as well. The kinetic of bilirubin sorption and human serum albumin at their joint presence in hydrogel solutions was defined. Bilirubin sorption and change of albumin composition was considered. The sorption of middle molecular peptides was considered as well. The sorption of endogenous toxin by means of ethynyl-piperidol polymers was done.

Covalently Cross-Linked Sulfone Polybenzimidazole Membranes with Poly(Vinylbenzyl Chloride) for Fuel Cell Applications

DEFF Research Database (Denmark)

Yang, Jingshuai; Aili, David; Li, Qingfeng

2013-01-01

Covalently cross-linked polymer membranes were fabricated from poly(aryl sulfone benzimidazole) (SO(2) PBI) and poly(vinylbenzyl chloride) (PVBCl) as electrolytes for high-temperature proton-exchange-membrane fuel cells. The cross-linking imparted organo insolubility and chemical stability against...

Preparation of Nanocellulose Reinforced Chitosan Films, Cross-Linked by Adipic Acid

Directory of Open Access Journals (Sweden)

Pouria Falamarzpour

2017-02-01

Full Text Available Adipic acid, an abundant and nontoxic compound, was used to dissolve and cross-link chitosan. After the preparation of chitosan films through casting technique, the in situ amidation reaction was performed at 80–100 °C as verified by Fourier transform infrared (FT-IR. The reaction was accompanied by the release of water which was employed to investigate the reaction kinetics. Accordingly, the reaction rate followed the first-order model and Arrhenius equation, and the activation energy was calculated to be 18 kJ/mol. Furthermore, the mechanical properties of the chitosan films were comprehensively studied. First, optimal curing conditions (84 °C, 93 min were introduced through a central composite design. In order to evaluate the effects of adipic acid, the mechanical properties of physically cross-linked (uncured, chemically cross-linked (cured, and uncross-linked (prepared by acetic acid films were compared. The use of adipic acid improved the tensile strength of uncured and chemically cross-linked films more than 60% and 113%, respectively. Finally, the effect of cellulose nanofibrils (CNFs on the mechanical performance of cured films, in the presence of glycerol as a plasticizer, was investigated. The plasticized chitosan films reinforced by 5 wt % CNFs showed superior properties as a promising material for the development of chitosan-based biomaterials.

Radiation cross-linked collagen/dextran dermal scaffolds: effects of dextran on cross-linking and degradation.

Science.gov (United States)

Zhang, Yaqing; Zhang, Xiangmei; Xu, Ling; Wei, Shicheng; Zhai, Maolin

2015-01-01

Ionizing radiation effectively cross-links collagen into network with enhanced anti-degradability and biocompatibility, while radiation-cross-linked collagen scaffold lacks flexibility, satisfactory surface appearance, and performs poor in cell penetration and ingrowth. To make the radiation-cross-linked collagen scaffold to serve as an ideal artificial dermis, dextran was incorporated into collagen. Scaffolds with the collagen/dextran (Col/Dex) ratios of 10/0, 7/3, and 5/5 were fabricated via (60)Co γ-irradiation cross-linking, followed by lyophilization. The morphology, microstructure, physicochemical, and biological properties were investigated. Compared with pure collagen, scaffolds with dextran demonstrated more porous appearance, enhanced hydrophilicity while the cross-linking density was lower with the consequence of larger pore size, higher water uptake, as well as reduced stiffness. Accelerated degradation was observed when dextran was incorporated in both the in vitro and in vivo assays, which led to earlier integration with cell and host tissue. The effect of dextran on degradation was ascribed to the decreased cross-linking density, looser microstructure, more porous and hydrophilic surface. Considering the better appearance, softness, moderate degradation rate due to controllable cross-linking degree and good biocompatibility as well, radiation-cross-linked collagen/dextran scaffolds are expected to serve as promising artificial dermal substitutes.

Fingerprinting Desmosine-Containing Elastin Peptides

Science.gov (United States)

Schräder, Christoph U.; Heinz, Andrea; Majovsky, Petra; Schmelzer, Christian E. H.

2015-05-01

Elastin is a vital protein of the extracellular matrix of jawed vertebrates and provides elasticity to numerous tissues. It is secreted in the form of its soluble precursor tropoelastin, which is subsequently cross-linked in the course of the elastic fiber assembly. The process involves the formation of the two tetrafunctional amino acids desmosine (DES) and isodesmosine (IDES), which are unique to elastin. The resulting high degree of cross-linking confers remarkable properties, including mechanical integrity, insolubility, and long-term stability to the protein. These characteristics hinder the structural elucidation of mature elastin. However, MS2 data of linear and cross-linked peptides released by proteolysis can provide indirect insights into the structure of elastin. In this study, we performed energy-resolved collision-induced dissociation experiments of DES, IDES, their derivatives, and DES-/IDES-containing peptides to determine characteristic product ions. It was found that all investigated compounds yielded the same product ion clusters at elevated collision energies. Elemental composition determination using the exact masses of these ions revealed molecular formulas of the type CxHyN, suggesting that the pyridinium core of DES/IDES remains intact even at relatively high collision energies. The finding of these specific product ions enabled the development of a similarity-based scoring algorithm that was successfully applied on LC-MS/MS data of bovine elastin digests for the identification of DES-/IDES-cross-linked peptides. This approach facilitates the straightforward investigation of native cross-links in elastin.

Cross-linking of polymeric materials

International Nuclear Information System (INIS)

Bloom, L.I.; Du Plessis, T.A.; Meij, G.O.

1991-01-01

The invention provides a method of producing a cured polymeric artifact from a polymeric thermoplastic starting material, the material of the artifact having reduced thermoplasticity relative to the starting material and exhibiting an enhanced degree of cross-linking relative to the starting material. The method includes subjecting a polymeric thermoplastic starting material, which is capable of being cross-linked by irradiation, to sufficient irradiation partially to cross-linked the starting material to produce a thermoplastic partially cross-linked intermediate material. The thermoplasticity of the intermediate material is then reduced by heating it to raise its melting point. The invention also provides a method of making a partially cross-linked feedstocks and a master batch for use in making such artifacts

Evaluation of cross-linked gelatin membranes as delivery carriers for retinal sheets

Energy Technology Data Exchange (ETDEWEB)

Lai, Jui-Yang, E-mail: jylai@mail.cgu.edu.tw [Institute of Biochemical and Biomedical Engineering, Chang Gung University, Taoyuan, 33302 Taiwan (China); Biomedical Engineering Research Center, Chang Gung University, Taoyuan, 33302 Taiwan (China); Molecular Medicine Research Center, Chang Gung University, Taoyuan, 33302 Taiwan (China); Li, Ya-Ting [Institute of Biochemical and Biomedical Engineering, Chang Gung University, Taoyuan, 33302 Taiwan (China)

2010-06-15

The delivery of intact sheet transplants to the subretinal space can prevent cell loss that is generally associated with the injection of cell suspensions or cell aggregates. The aim of this study was to develop chemically modified gelatin matrices that enhance the delivery efficiency and analyze whether the gelatin membranes cross-linked with 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide (EDC) can be considered as potential carriers for retinal sheets. The characteristics of EDC cross-linked gelatin membranes were determined by mechanical and in vitro degradation tests, melting point measurements, cell proliferation assays, cytokine expression analyses, and tissue delivery studies. Gelatin membranes without cross-linking and glutaraldehyde cross-linked gelatin samples were used for comparison. Results of this study indicated that introduction of cross-links is capable of rendering the gelatin network more stable against mechanical stresses and deformations as well as rapid hydrolysis during intraocular delivery of delicate tissue sheets. In comparison with the glutaraldehyde treated samples, the EDC cross-linked gelatin membranes showed a better degradation profile and a relatively higher cytocompatibility. In addition, after EDC cross-linking, the gelatin matrices having an acceptable melting point could be used for the fabrication of a sandwich-like carrier with a high transfer and encapsulation efficiency. These findings suggest that the cross-linking agent type gives an influence on delivery functionality of gelatin membranes. In summary, the EDC cross-linked gelatin is an ideal candidate for use as a carrier material in retinal sheet delivery applications.

Mapping protein-RNA interactions by RCAP, RNA-cross-linking and peptide fingerprinting.

Science.gov (United States)

Vaughan, Robert C; Kao, C Cheng

2015-01-01

RNA nanotechnology often feature protein RNA complexes. The interaction between proteins and large RNAs are difficult to study using traditional structure-based methods like NMR or X-ray crystallography. RCAP, an approach that uses reversible-cross-linking affinity purification method coupled with mass spectrometry, has been developed to map regions within proteins that contact RNA. This chapter details how RCAP is applied to map protein-RNA contacts within virions.

Chemical methods for peptide and protein production.

Science.gov (United States)

Chandrudu, Saranya; Simerska, Pavla; Toth, Istvan

2013-04-12

Since the invention of solid phase synthetic methods by Merrifield in 1963, the number of research groups focusing on peptide synthesis has grown exponentially. However, the original step-by-step synthesis had limitations: the purity of the final product decreased with the number of coupling steps. After the development of Boc and Fmoc protecting groups, novel amino acid protecting groups and new techniques were introduced to provide high quality and quantity peptide products. Fragment condensation was a popular method for peptide production in the 1980s, but unfortunately the rate of racemization and reaction difficulties proved less than ideal. Kent and co-workers revolutionized peptide coupling by introducing the chemoselective reaction of unprotected peptides, called native chemical ligation. Subsequently, research has focused on the development of novel ligating techniques including the famous click reaction, ligation of peptide hydrazides, and the recently reported α-ketoacid-hydroxylamine ligations with 5-oxaproline. Several companies have been formed all over the world to prepare high quality Good Manufacturing Practice peptide products on a multi-kilogram scale. This review describes the advances in peptide chemistry including the variety of synthetic peptide methods currently available and the broad application of peptides in medicinal chemistry.

Chemical cross-linking and mass spectrometry as structure determination tools

Czech Academy of Sciences Publication Activity Database

Novák, Petr; Giannakopulos, A.

2007-01-01

Roč. 13, - (2007), s. 105-113 ISSN 1469-0667 R&D Projects: GA AV ČR KJB400200501; GA MŠk LC545 Grant - others:SE(XE) Marie Curie Actions TOK, Contract No. MTKD-CT-2004-014407 Institutional research plan: CEZ:AV0Z50200510 Keywords : cross-linking * protein * high order structure Subject RIV: EE - Microbiology, Virology Impact factor: 1.198, year: 2007

Comparative study of PBI Cross Linked Utilizing Agents of Varying Steric Configurations

DEFF Research Database (Denmark)

Kirkebcek, Andreas; Aili, David; Li, Qingfeng

2016-01-01

ionic or covalent cross linking. When considering such, little attention is devoted to explore the effect of the sterical configuration of the cross linking agent. In this contribution three different cross linking agents are utilized to evaluate how these affects final membrane properties.......The high thermal and chemical stability of poly[2,2'-(m-phenylene)-5,5' bibenzimidazole] (PBI) accounts for its wise spread use in high temperature polymer electrolyte membrane fuel cells (HT- PEMFC). By doping the membrane with phosphoric acid (PA) ionic conductivity is obtained. Thus conductivity...... is dependent on the amount of PA present within the membrane. However mechanical properties are reduced are significantly reduced due to the plasticizing effect shown by PA [1]. This effect is due to PBI chain displacement. This effect can be lessened by use of cross linking [2-4]. This can be obtained using...

Gamma-radiation induced cross-links in ethylene-propylene rubber studied by CP-MAS NMR

International Nuclear Information System (INIS)

Sohma, J.; Shiotani, M.; Murakami, S.

1983-01-01

A new technique of 13 C-NMR, the CP-MAS method, was applied to study a chemistry of cross-links induced by #betta#-irradiation of ethylene-propylene rubber. The chemical species of cross-linking points were specified with their relative concentrations by the analysis of the CP-MAS spectra obtained before and after the irradiation. It was found that the short branches were also formed by the irradiation. A comparison was made between the cross-links detected by the CP-MAS method and those obtained by the Charlesby-Pinner analysis of the gelation caused by the #betta#-irradiation. The conventional 13 C-NMR of the cross-linked and swollen EPR provided us an information on the sol parts of the sample but little information on the cross-links in the gel parts. (author)

Designer Self-Assembling Peptide Nanofiber Scaffolds Containing Link Protein N-Terminal Peptide Induce Chondrogenesis of Rabbit Bone Marrow Stem Cells

Directory of Open Access Journals (Sweden)

Baichuan Wang

2014-01-01

Full Text Available Designer self-assembling peptide nanofiber hydrogel scaffolds have been considered as promising biomaterials for tissue engineering because of their excellent biocompatibility and biofunctionality. Our previous studies have shown that a novel designer functionalized self-assembling peptide nanofiber hydrogel scaffold (RLN/RADA16, LN-NS containing N-terminal peptide sequence of link protein (link N can promote nucleus pulposus cells (NPCs adhesion and three-dimensional (3D migration and stimulate biosynthesis of type II collagen and aggrecan by NPCs in vitro. The present study has extended these investigations to determine the effects of this functionalized LN-NS on bone marrow stem cells (BMSCs, a potential cell source for NP regeneration. Although the functionalized LN-NS cannot promote BMSCs proliferation, it significantly promotes BMSCs adhesion compared with that of the pure RADA16 hydrogel scaffold. Moreover, the functionalized LN-NS remarkably stimulates biosynthesis and deposition of type II collagen and aggrecan. These data demonstrate that the functionalized peptide nanofiber hydrogel scaffold containing link N peptide as a potential matrix substrate will be very useful in the NP tissue regeneration.

Photo-cross-linked small-molecule microarrays as chemical genomic tools for dissecting protein-ligand interactions.

Science.gov (United States)

Kanoh, Naoki; Asami, Aya; Kawatani, Makoto; Honda, Kaori; Kumashiro, Saori; Takayama, Hiroshi; Simizu, Siro; Amemiya, Tomoyuki; Kondoh, Yasumitsu; Hatakeyama, Satoru; Tsuganezawa, Keiko; Utata, Rei; Tanaka, Akiko; Yokoyama, Shigeyuki; Tashiro, Hideo; Osada, Hiroyuki

2006-12-18

We have developed a unique photo-cross-linking approach for immobilizing a variety of small molecules in a functional-group-independent manner. Our approach depends on the reactivity of the carbene species generated from trifluoromethylaryldiazirine upon UV irradiation. It was demonstrated in model experiments that the photogenerated carbenes were able to react with every small molecule tested, and they produced multiple conjugates in most cases. It was also found in on-array immobilization experiments that various small molecules were immobilized, and the immobilized small molecules retained their ability to interact with their binding proteins. With this approach, photo-cross-linked microarrays of about 2000 natural products and drugs were constructed. This photo-cross-linked microarray format was found to be useful not merely for ligand screening but also to study the structure-activity relationship, that is, the relationship between the structural motif (or pharmacophore) found in small molecules and its binding affinity toward a protein, by taking advantage of the nonselective nature of the photo-cross-linking process.

A novel nanoemulsion-based method to produce ultrasmall, water-dispersible nanoparticles from chitosan, surface modified with cell-penetrating peptide for oral delivery of proteins and peptides

Directory of Open Access Journals (Sweden)

Barbari GR

2017-05-01

Full Text Available Ghullam Reza Barbari,1 Farid Abedin Dorkoosh,1 Mohsen Amini,2 Mohammad Sharifzadeh,3 Fateme Atyabi,1 Saeed Balalaie,4 Niyousha Rafiee Tehrani,5 Morteza Rafiee Tehrani1 1Department of Pharmaceutics, 2Department of Medicinal Chemistry, 3Department of Pharmacology, School of Pharmacy, Tehran University of Medical Sciences, 4Department of Chemistry, Khaje Nasiroddin University, 5Department of Biochemistry, School of Medicine, Tehran University of Medical Sciences, Tehran, Iran Abstract: A simple and reproducible water-in-oil (W/O nanoemulsion technique for making ultrasmall (<15 nm, monodispersed and water-dispersible nanoparticles (NPs from chitosan (CS is reported. The nano-sized (50 nm water pools of the W/O nanoemulsion serve as “nano-containers and nano-reactors”. The entrapped polymer chains of CS inside these “nano-reactors” are covalently cross-linked with the chains of polyethylene glycol (PEG, leading to rigidification and formation of NPs. These NPs possess excessive swelling properties in aqueous medium and preserve integrity in all pH ranges due to chemical cross-linking with PEG. A potent and newly developed cell-penetrating peptide (CPP is further chemically conjugated to the surface of the NPs, leading to development of a novel peptide-conjugated derivative of CS with profound tight-junction opening properties. The CPP-conjugated NPs can easily be loaded with almost all kinds of proteins, peptides and nucleotides for oral delivery applications. Feasibility of this nanoparticulate system for oral delivery of a model peptide (insulin is investigated in Caco-2 cell line. The cell culture results for translocation of insulin across the cell monolayer are very promising (15%–19% increase, and animal studies are actively under progress and will be published separately. Keywords: ultrasmall, cell-penetrating peptide, chitosan, oral insulin, nanoemulsion, Caco-2 cell

Physicochemical characterization of chitosan/nylon6/polyurethane foam chemically cross-linked ternary blends.

Science.gov (United States)

Jayakumar, S; Sudha, P N

2013-03-15

Chitosan/nylon6/polyurethane foam (CS/Ny6/PUF) ternary blend was prepared and chemically cross-linked with glutaraldehyde. Structural, thermal and morphological studies were performed for the prepared ternary blends. Characterizations of the ternary blends were investigated by Fourier transform infrared spectroscopy (FTIR), thermo gravimetric analysis (TGA), differential scanning calorimetry (DSC), X-ray diffraction (XRD) and scanning electron microscope (SEM). The FTIR results showed that the strong intermolecular hydrogen bonds took place between CS, Ny6 and PUF. TGA and DSC studies reveal that the thermal stability of the blend is enhanced by glutaraldehyde as crosslinking agent. Results of XRD indicated that the relative crystalline of pure CS film was reduced when the polymeric network was reticulated by glutaraldehyde. Finally, the results of scanning electron microscopy (SEM) indicated that the morphology of the blend is rough and heterogeneous, further it confirms the interaction between the functional groups of the blend components. Copyright © 2012 Elsevier B.V. All rights reserved.

Chemical Methods for Peptide and Protein Production

Directory of Open Access Journals (Sweden)

Istvan Toth

2013-04-01

Full Text Available Since the invention of solid phase synthetic methods by Merrifield in 1963, the number of research groups focusing on peptide synthesis has grown exponentially. However, the original step-by-step synthesis had limitations: the purity of the final product decreased with the number of coupling steps. After the development of Boc and Fmoc protecting groups, novel amino acid protecting groups and new techniques were introduced to provide high quality and quantity peptide products. Fragment condensation was a popular method for peptide production in the 1980s, but unfortunately the rate of racemization and reaction difficulties proved less than ideal. Kent and co-workers revolutionized peptide coupling by introducing the chemoselective reaction of unprotected peptides, called native chemical ligation. Subsequently, research has focused on the development of novel ligating techniques including the famous click reaction, ligation of peptide hydrazides, and the recently reported a-ketoacid-hydroxylamine ligations with 5-oxaproline. Several companies have been formed all over the world to prepare high quality Good Manufacturing Practice peptide products on a multi-kilogram scale. This review describes the advances in peptide chemistry including the variety of synthetic peptide methods currently available and the broad application of peptides in medicinal chemistry.

Chemical structure, biosynthesis and synthesis of free and glycosylated pyridinolines formed by cross-link of bone and synovium collagen.

Science.gov (United States)

Anastasia, Luigi; Rota, Paola; Anastasia, Mario; Allevi, Pietro

2013-09-21

This review focuses on the chemical structure, biosynthesis and synthesis of free and glycosylated pyridinolines (Pyds), fluorescent collagen cross-links, with a pyridinium salt structure. Pyds derive from the degradation of bone collagen and have attracted attention for their use as biochemical markers of bone resorption and to assess fracture risk prediction in persons suffering from osteoporosis, bone cancer and other bone or collagen diseases. We consider and critically discuss all reported syntheses of free and glycosylated Pyds evidencing an unrevised chemistry, original and of general utility, analysis of which allows us to also support a previously suggested non-enzymatic formation of Pyds in collagen better rationalizing and justifying the chemical events.

Characterization of the degree of cross-linking in radiation cross-linked low and high density polyethylenes

International Nuclear Information System (INIS)

Posselt, K.; Haedrich, W.

1986-01-01

In practice the cross-linking of irradiated polyethylene is mostly characterized by solubility and thermomechanical data. The irradiation of samples of a LDPE and a HDPE yields very different gel-dose curves. But for a quantitative comparison the complicated connection between the gel values and the corresponding densities of cross-links, especially the dependence on the initial molecular size distribution, has to take into consideration. The analysis of the solubility data according to the statistical theory of cross-linking developed by Inokuti and Saito shows that at equal doses in both investigated PE types in spite of the different gel values nearly the same densities of cross-links are present. That result is confirmed by the densities of cross-links determined from stress-strain measurements at 423 K. (author)

Comparative Study of One-Step Cross-Linked Electrospun Chitosan-Based Membranes

Directory of Open Access Journals (Sweden)

Yanet E. Aguirre-Chagala

2017-01-01

Full Text Available Chitosan membranes are widely applied for tissue engineering; however, a major drawback is their low resistance in aqueous phases and therefore the structure collapses impeding their long-term use. Although there is extensive research, because of chitosan’s importance as a biomaterial, studies involving chitosan-based membranes are still needed. Herein, a detailed investigation of diverse chemical routes to cross-link fibers in situ by electrospinning process is described. In case of using genipin as cross-linker, a close relationship with the content and the mean diameter values is reported, suggesting a crucial effect over the design of nanostructures. Also, the physical resistance is enhanced for the combination of two types of methods, such as chemical and physical methods. Cross-linked fibers upon exposure to long wave ultraviolet A (UVA light change their morphology, but not their chemical composition. When they are incubated in aqueous phase for 70 days, they show an extensive improvement of their macrostructural integrity which makes them attractive candidates for tissue engineering application. As a result, the thermal properties of these materials reveal less crystallinity and higher temperature of degradation.

Fast and Accurate Identification of Cross-Linked Peptides for the Structural Analysis of Large Protein Complexes and Elucidation of Interaction Networks. / Tahir, Salman; Bukowski-Wills, Jimi-Carlo; Rasmussen, Morten; Rappsilber, Juri

DEFF Research Database (Denmark)

Rasmussen, Morten

to investigate protein structure and protein-protein interactions. When applied to single proteins or small purified protein complexes, this methodology works well. However certain challenges arise when applied to more complex samples. One of the main problems is the combinatorial increase in the search space...... simplify a spectrum because we remove all peaks that are accounted for by the fragmentation of peptide one. This approach is highly sensitive and scales well as revealed by searching our data of synthetic cross-links against a large sequence database. Currently, against a protein database of >1300 proteins...... a spectrum is searched in 0.35 seconds - a vast improvement when compared to the exhaustive search method of combining every potential cross-link for each spectrum(60 hours). In fact the search time is comparable, if not better, than existing linear search engines. Furthermore, we auto-validate the results...

Isolation, chemical and functional characterization of several new K(+)-channel blocking peptides from the venom of the scorpion Centruroides tecomanus.

Science.gov (United States)

Olamendi-Portugal, Timoteo; Bartok, Adam; Zamudio-Zuñiga, Fernando; Balajthy, Andras; Becerril, Baltazar; Panyi, Gyorgy; Possani, Lourival D

2016-06-01

Six new peptides were isolated from the venom of the Mexican scorpion Centruroides tecomanus; their primary structures were determined and the effects on ion channels were verified by patch-clamp experiments. Four are K(+)-channel blockers of the α-KTx family, containing 32 to 39 amino acid residues, cross-linked by three disulfide bonds. They all block Kv1.2 in nanomolar concentrations and show various degree of selectivity over Kv1.1, Kv1.3, Shaker and KCa3.1 channels. One peptide has 42 amino acids cross-linked by four disulfides; it blocks ERG-channels and belongs to the γ-KTx family. The sixth peptide has only 32 amino acid residues, three disulfide bonds and has no effect on the ion-channels assayed. It also does not have antimicrobial activity. Systematic numbers were assigned (time of elution on HPLC): α-KTx 10.4 (time 24.1); α-KTx 2.15 (time 26.2); α-KTx 2.16 (time 23.8); α-KTx 2.17 (time 26.7) and γ-KTx 1.9 (elution time 29.6). A partial proteomic analysis of the short chain basic peptides of this venom, which elutes on carboxy-methyl-cellulose column fractionation, is included. The pharmacological properties of the peptides described in this study may provide valuable tools for understanding the structure-function relationship of K(+) channel blocking scorpion toxins. Copyright © 2016 Elsevier Ltd. All rights reserved.

Characterization of Disulfide-Linked Peptides Using Tandem Mass Spectrometry Coupled with Automated Data Analysis Software

Science.gov (United States)

Liang, Zhidan; McGuinness, Kenneth N.; Crespo, Alejandro; Zhong, Wendy

2018-05-01

Disulfide bond formation is critical for maintaining structure stability and function of many peptides and proteins. Mass spectrometry has become an important tool for the elucidation of molecular connectivity. However, the interpretation of the tandem mass spectral data of disulfide-linked peptides has been a major challenge due to the lack of appropriate tools. Developing proper data analysis software is essential to quickly characterize disulfide-linked peptides. A thorough and in-depth understanding of how disulfide-linked peptides fragment in mass spectrometer is a key in developing software to interpret the tandem mass spectra of these peptides. Two model peptides with inter- and intra-chain disulfide linkages were used to study fragmentation behavior in both collisional-activated dissociation (CAD) and electron-based dissociation (ExD) experiments. Fragments generated from CAD and ExD can be categorized into three major types, which result from different S-S and C-S bond cleavage patterns. DiSulFinder is a computer algorithm that was newly developed based on the fragmentation observed in these peptides. The software is vendor neutral and capable of quickly and accurately identifying a variety of fragments generated from disulfide-linked peptides. DiSulFinder identifies peptide backbone fragments with S-S and C-S bond cleavages and, more importantly, can also identify fragments with the S-S bond still intact to aid disulfide linkage determination. With the assistance of this software, more comprehensive disulfide connectivity characterization can be achieved. [Figure not available: see fulltext.

Characterization of Disulfide-Linked Peptides Using Tandem Mass Spectrometry Coupled with Automated Data Analysis Software.

Science.gov (United States)

Liang, Zhidan; McGuinness, Kenneth N; Crespo, Alejandro; Zhong, Wendy

2018-01-25

Disulfide bond formation is critical for maintaining structure stability and function of many peptides and proteins. Mass spectrometry has become an important tool for the elucidation of molecular connectivity. However, the interpretation of the tandem mass spectral data of disulfide-linked peptides has been a major challenge due to the lack of appropriate tools. Developing proper data analysis software is essential to quickly characterize disulfide-linked peptides. A thorough and in-depth understanding of how disulfide-linked peptides fragment in mass spectrometer is a key in developing software to interpret the tandem mass spectra of these peptides. Two model peptides with inter- and intra-chain disulfide linkages were used to study fragmentation behavior in both collisional-activated dissociation (CAD) and electron-based dissociation (ExD) experiments. Fragments generated from CAD and ExD can be categorized into three major types, which result from different S-S and C-S bond cleavage patterns. DiSulFinder is a computer algorithm that was newly developed based on the fragmentation observed in these peptides. The software is vendor neutral and capable of quickly and accurately identifying a variety of fragments generated from disulfide-linked peptides. DiSulFinder identifies peptide backbone fragments with S-S and C-S bond cleavages and, more importantly, can also identify fragments with the S-S bond still intact to aid disulfide linkage determination. With the assistance of this software, more comprehensive disulfide connectivity characterization can be achieved. Graphical Abstract ᅟ.

Effect of nordihydroguaiaretic acid cross-linking on fibrillar collagen: in vitro evaluation of fibroblast adhesion strength and migration

Directory of Open Access Journals (Sweden)

Ana Y. Rioja

2017-04-01

Full Text Available Fixation is required to reinforce reconstituted collagen for orthopedic bioprostheses such as tendon or ligament replacements. Previous studies have demonstrated that collagen fibers cross-linked by the biocompatible dicatechol nordihydroguaiaretic acid (NDGA have mechanical strength comparable to native tendons. This work focuses on investigating fibroblast behavior on fibrillar and NDGA cross-linked type I collagen to determine if NDGA modulates cell adhesion, morphology, and migration. A spinning disk device that applies a range of hydrodynamic forces under uniform chemical conditions was employed to sensitively quantify cell adhesion strength, and a radial barrier removal assay was used to measure cell migration on films suitable for these quantitative in vitro assays. The compaction of collagen films, mediated by the drying and cross-linking fabrication process, suggests a less open organization compared to native fibrillar collagen that likely allowed the collagen to form more inter-chain bonds and chemical links with NDGA polymers. Fibroblasts strongly adhered to and migrated on native and NDGA cross-linked fibrillar collagen; however, NDGA modestly reduced cell spreading, adhesion strength and migration rate. Thus, it is hypothesized that NDGA cross-linking masked some adhesion receptor binding sites either physically, chemically, or both, thereby modulating adhesion and migration. This alteration in the cell-material interface is considered a minimal trade-off for the superior mechanical and compatibility properties of NDGA cross-linked collagen compared to other fixation approaches.

Role of solvent-mediated carbodiimide cross-linking in fabrication of electrospun gelatin nanofibrous membranes as ophthalmic biomaterials

International Nuclear Information System (INIS)

Chou, Shih-Feng; Luo, Li-Jyuan; Lai, Jui-Yang; Ma, David Hui-Kang

2017-01-01

Due to their ability to mimic the structure of extracellular matrix, electrospun gelatin nanofibers are promising cell scaffolding materials for tissue engineering applications. However, the hydrophilic gelatin molecules usually need stabilization before use in aqueous physiological environment. Considering that biomaterials cross-linked via film immersion technique may have a more homogeneous cross-linked structure than vapor phase cross-linking, this work aims to investigate the chemical modification of electrospun gelatin nanofibrous membranes by liquid phase carbodiimide in the presence of ethanol/water co-solvents with varying ethanol concentrations ranging from 80 to 99.5 vol%. The results of characterization showed that increasing water content in the binary reaction solvent system increases the extent of cross-linking of gelatin nanofibers, but simultaneously promotes the effect of biopolymer swelling and distortion in fiber mat structure. As compared to non-cross-linked counterparts, carbodiimide treated gelatin nanofibrous mats exhibited better thermal and biological stability where the shrinkage temperature and resistance to enzymatic degradation varied in response to ethanol/water solvent composition-mediated generation of cross-links. Irrespective of their cross-linking density, all studied membrane samples did not induce any responses in ocular epithelial cell cultures derived from cornea, lens, and retina. Unlike many other cross-linking agents and/or methods (e.g., excessive vapor phase cross-linking) that may pose a risk of toxicity, our study demonstrated that these nanofibrous materials are well tolerated by anterior segment tissues. These findings also indicate the safety of using ethanol/water co-solvents for chemical cross-linking of gelatin to engineer nanofibrous materials with negligible biological effects. In summary, the present results suggest the importance of solvent-mediated carbodiimide cross-linking in modulating structure�

Role of solvent-mediated carbodiimide cross-linking in fabrication of electrospun gelatin nanofibrous membranes as ophthalmic biomaterials

Energy Technology Data Exchange (ETDEWEB)

Chou, Shih-Feng [Department of Mechanical Engineering, University of Texas at Tyler, Tyler, TX 75799 (United States); Luo, Li-Jyuan [Department of Chemical and Materials Engineering, Chang Gung University, Taoyuan 33302, Taiwan, ROC (China); Lai, Jui-Yang, E-mail: jylai@mail.cgu.edu.tw [Institute of Biochemical and Biomedical Engineering, Chang Gung University, Taoyuan 33302, Taiwan, ROC (China); Biomedical Engineering Research Center, Chang Gung University, Taoyuan 33302, Taiwan, ROC (China); Center for Tissue Engineering, Chang Gung Memorial Hospital, Taoyuan 33305, Taiwan, ROC (China); Department of Ophthalmology, Chang Gung Memorial Hospital, Taoyuan 33305, Taiwan, ROC (China); Department of Materials Engineering, Ming Chi University of Technology, New Taipei City 24301, Taiwan, ROC (China); Ma, David Hui-Kang [Center for Tissue Engineering, Chang Gung Memorial Hospital, Taoyuan 33305, Taiwan, ROC (China); Department of Ophthalmology, Chang Gung Memorial Hospital, Taoyuan 33305, Taiwan, ROC (China); Department of Chinese Medicine, Chang Gung University, Taoyuan 33302, Taiwan, ROC (China)

2017-02-01

Due to their ability to mimic the structure of extracellular matrix, electrospun gelatin nanofibers are promising cell scaffolding materials for tissue engineering applications. However, the hydrophilic gelatin molecules usually need stabilization before use in aqueous physiological environment. Considering that biomaterials cross-linked via film immersion technique may have a more homogeneous cross-linked structure than vapor phase cross-linking, this work aims to investigate the chemical modification of electrospun gelatin nanofibrous membranes by liquid phase carbodiimide in the presence of ethanol/water co-solvents with varying ethanol concentrations ranging from 80 to 99.5 vol%. The results of characterization showed that increasing water content in the binary reaction solvent system increases the extent of cross-linking of gelatin nanofibers, but simultaneously promotes the effect of biopolymer swelling and distortion in fiber mat structure. As compared to non-cross-linked counterparts, carbodiimide treated gelatin nanofibrous mats exhibited better thermal and biological stability where the shrinkage temperature and resistance to enzymatic degradation varied in response to ethanol/water solvent composition-mediated generation of cross-links. Irrespective of their cross-linking density, all studied membrane samples did not induce any responses in ocular epithelial cell cultures derived from cornea, lens, and retina. Unlike many other cross-linking agents and/or methods (e.g., excessive vapor phase cross-linking) that may pose a risk of toxicity, our study demonstrated that these nanofibrous materials are well tolerated by anterior segment tissues. These findings also indicate the safety of using ethanol/water co-solvents for chemical cross-linking of gelatin to engineer nanofibrous materials with negligible biological effects. In summary, the present results suggest the importance of solvent-mediated carbodiimide cross-linking in modulating structure�

Collagen cross-linking in sun-exposed and unexposed sites of aged human skin

Science.gov (United States)

Yamauchi, M.; Prisayanh, P.; Haque, Z.; Woodley, D. T.

1991-01-01

A recently described nonreducible, acid-heat stable compound, histidinohydroxylysinonorleucine (HHL), is a collagen cross-link isolated from mature skin tissue. Its abundance is related to chronologic aging of skin. The present communication describes the quantity of HHL from aged human skin of the same individuals in sun-exposed (wrist) and unexposed (buttock) sites. Punch biopsies were obtained from these sites from nine people of age 60 or older. HHL contents (moles/mole of collagen) at these sites were for wrist 0.13 +/- 0.07 and for buttock 0.69 +/- 0.17 (mean +/- SD, p less than 0.001). In addition, it was found that acute irradiation of the cross-linked peptides with UVA (up to 250 J/cm2) and UVB (up to 1 J/cm2) had no effect on HHL structure. The same treatment significantly degraded another nonreducible, stable collagen cross-link, pyridinoline. The results suggest that chronic sunlight exposure may be associated with an impediment to normal maturation of human dermal collagen resulting in tenuous amount of HHL. Thus, the process of photoaging in dermal collagen is different from that of chronologic aging in human skin.

The synthesis and coupling of photoreactive collagen-based peptides to restore integrin reactivity to an inert substrate, chemically-crosslinked collagen

Science.gov (United States)

Malcor, Jean-Daniel; Bax, Daniel; Hamaia, Samir W.; Davidenko, Natalia; Best, Serena M.; Cameron, Ruth E.; Farndale, Richard W.; Bihan, Dominique

2016-01-01

Collagen is frequently advocated as a scaffold for use in regenerative medicine. Increasing the mechanical stability of a collagen scaffold is widely achieved by cross-linking using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS). However, this treatment consumes the carboxylate-containing amino acid sidechains that are crucial for recognition by the cell-surface integrins, abolishing cell adhesion. Here, we restore cell reactivity to a cross-linked type I collagen film by covalently linking synthetic triple-helical peptides (THPs), mimicking the structure of collagen. These THPs are ligands containing an active cell-recognition motif, GFOGER, a high-affinity binding site for the collagen-binding integrins. We end-stapled peptide strands containing GFOGER by coupling a short diglutamate-containing peptide to their N-terminus, improving the thermal stability of the resulting THP. A photoreactive Diazirine group was grafted onto the end-stapled THP to allow covalent linkage to the collagen film upon UV activation. Such GFOGER-derivatized collagen films showed restored affinity for the ligand-binding I domain of integrin α2β1, and increased integrin-dependent cell attachment and spreading of HT1080 and Rugli cell lines, expressing integrins α2β1 and α1β1, respectively. The method we describe has wide application, beyond collagen films or scaffolds, since the photoreactive diazirine will react with many organic carbon skeletons. PMID:26854392

Towards a fully synthetic substitute of alginate: optimization of a thermal gelation/chemical cross-linking scheme ("tandem" gelation) for the production of beads and liquid-core capsules.

Science.gov (United States)

Cellesi, F; Weber, W; Fussenegger, M; Hubbell, J A; Tirelli, N

2004-12-20

Fully synthetic polymers were used for the preparation of hydrogel beads and capsules, in a processing scheme that, originally designed for calcium alginate, was adapted to a "tandem" process, that is the combination a physical gelation with a chemical cross-linking. The polymers feature a Tetronic backbone (tetra armed Pluronics), which exhibits a reverse thermal gelation in water solutions within a physiological range of temperatures and pHs. The polymers bear terminal reactive groups that allow for a mild, but effective chemical cross-linking. Given an appropriate temperature jump, the thermal gelation provides a hardening kinetics similar to that of alginate. With slower kinetics, the chemical cross-linking then develops an irreversible and elastic gel structure, and determines its transport properties. In the present article this process has been optimized for the production of monodisperse, high elastic, hydrogel microbeads, and liquid-core microcapsules. We also show the feasibility of the use of liquid-core microcapsules in cell encapsulation. In preliminary experiments, CHO cells have been successfully encapsulated preserving their viability during the process and after incubation. The advantages of this process are mainly in the use of synthetic polymers, which provide great flexibility in the molecular design. This, in principle, allows for a precise tailoring of mechanical and transport properties and of bioactivity of the hydrogels, and also for a precise control in material purification.

Cross-linking of cell surface amyloid precursor protein leads to increased β-amyloid peptide production in hippocampal neurons: implications for Alzheimer's disease.

Science.gov (United States)

Lefort, Roger; Pozueta, Julio; Shelanski, Michael

2012-08-01

The accumulation of the β-amyloid peptide (Aβ) in Alzheimer's disease (AD) is thought to play a causative role in triggering synaptic dysfunction in neurons, leading to their eventual demise through apoptosis. Aβ is produced and secreted upon sequential cleavage of the amyloid precursor protein (APP) by β-secretases and γ-secretases. However, while Aβ levels have been shown to be increased in the brains of AD patients, little is known about how the cleavage of APP and the subsequent generation of Aβ is influenced, or whether the cleavage process changes over time. It has been proposed that Aβ can bind APP and promote amyloidogenic processing of APP, further enhancing Aβ production. Proof of this idea has remained elusive because a clear mechanism has not been identified, and the promiscuous nature of Aβ binding complicates the task of demonstrating the idea. To work around these problems, we used an antibody-mediated approach to bind and cross-link cell-surface APP in cultured rat primary hippocampal neurons. Here we show that cross-linking of APP is sufficient to raise the levels of Aβ in viable neurons with a concomitant increase in the levels of the β-secretase BACE1. This appears to occur as a result of a sorting defect that stems from the caspase-3-mediated inactivation of a key sorting adaptor protein, namely GGA3, which prevents the lysosomal degradation of BACE1. Together, our data suggest the occurrence of a positive pathogenic feedback loop involving Aβ and APP in affected neurons possibly allowing Aβ to spread to nearby healthy neurons.

Fabrication and characterization of hydrothermal cross-linked chitosan porous scaffolds for cartilage tissue engineering applications.

Science.gov (United States)

Shamekhi, Mohammad Amin; Rabiee, Ahmad; Mirzadeh, Hamid; Mahdavi, Hamid; Mohebbi-Kalhori, Davod; Baghaban Eslaminejad, Mohamadreza

2017-11-01

The use of various chemical cross-linking agents for the improvement of scaffolds physical and mechanical properties is a common practical method, which is limited by cytotoxicity effects. Due to exerting contract type forces, chondrocytes are known to implement shrinkage on the tissue engineered constructs, which can be avoided by the scaffold cross-linking. In the this research, chitosan scaffolds are cross-linked with hydrothermal treatment with autoclave sterilization time of 0, 10, 20 and 30min, to avoid the application of the traditional chemical toxic materials. The optimization studies with gel content and crosslink density measurements indicate that for 20min sterilization time, the gel content approaches to ~80%. The scaffolds are fully characterized by the conventional techniques such as SEM, porosity and permeability, XRD, compression, thermal analysis and dynamic mechanical thermal analysis (DMTA). FT-IR studies shows that autoclave inter-chain cross-linking reduces the amine group absorption at 1560cm -1 and increase the absorption of N-acetylated groups at 1629cm -1 . It is anticipated, that this observation evidenced by chitosan scaffold browning upon autoclave cross-linking is an indication of the familiar maillard reaction between amine moieties and carbonyl groups. The biodegradation rate analysis shows that chitosan scaffolds with lower concentrations, possess suitable degradation rate for cartilage tissue engineering applications. In addition, cytotoxicity analysis shows that fabricated scaffolds are biocompatible. The human articular chondrocytes seeding into 3D cross-linked scaffolds shows a higher viability and proliferation in comparison with the uncross-linked samples and 2D controls. Investigation of cell morphology on the scaffolds by SEM, shows a more spherical morphology of chondrocytes on the cross-linked scaffolds for 21days of in vitro culture. Copyright © 2017. Published by Elsevier B.V.

Key aromatic/hydrophobic amino acids controlling a cross-amyloid peptide interaction versus amyloid self-assembly.

Science.gov (United States)

Bakou, Maria; Hille, Kathleen; Kracklauer, Michael; Spanopoulou, Anna; Frost, Christina V; Malideli, Eleni; Yan, Li-Mei; Caporale, Andrea; Zacharias, Martin; Kapurniotu, Aphrodite

2017-09-01

The interaction of the intrinsically disordered polypeptide islet amyloid polypeptide (IAPP), which is associated with type 2 diabetes (T2D), with the Alzheimer's disease amyloid-β (Aβ) peptide modulates their self-assembly into amyloid fibrils and may link the pathogeneses of these two cell-degenerative diseases. However, the molecular determinants of this interaction remain elusive. Using a systematic alanine scan approach, fluorescence spectroscopy, and other biophysical methods, including heterocomplex pulldown assays, far-UV CD spectroscopy, the thioflavin T binding assay, transmission EM, and molecular dynamics simulations, here we identified single aromatic/hydrophobic residues within the amyloid core IAPP region as hot spots or key residues of its cross-interaction with Aβ40(42) peptide. Importantly, we also find that none of these residues in isolation plays a key role in IAPP self-assembly, whereas simultaneous substitution of four aromatic/hydrophobic residues with Ala dramatically impairs both IAPP self-assembly and hetero-assembly with Aβ40(42). Furthermore, our experiments yielded several novel IAPP analogs, whose sequences are highly similar to that of IAPP but have distinct amyloid self- or cross-interaction potentials. The identified similarities and major differences controlling IAPP cross-peptide interaction with Aβ40(42) versus its amyloid self-assembly offer a molecular basis for understanding the underlying mechanisms. We propose that these insights will aid in designing intervention strategies and novel IAPP analogs for the management of type 2 diabetes, Alzheimer's disease, or other diseases related to IAPP dysfunction or cross-amyloid interactions. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Mapping protein structural changes by quantitative cross-linking

Czech Academy of Sciences Publication Activity Database

Kukačka, Zdeněk; Strohalm, Martin; Kavan, Daniel; Novák, Petr

2015-01-01

Roč. 89, NOV 2015 (2015), s. 112-120 ISSN 1046-2023 R&D Projects: GA MŠk(CZ) EE2.3.20.0055; GA MŠk(CZ) EE2.3.30.0003; GA MŠk(CZ) ED1.1.00/02.0109 Grant - others:OPPC(XE) CZ.2.16/3.1.00/24023 Institutional support: RVO:61388971 Keywords : Chemical cross-linking * Proteolysis * Mass spectrometry Subject RIV: CE - Biochemistry Impact factor: 3.503, year: 2015

Enzyme-Linked Immunosorbent Assay Using a Virus Type-Specific Peptide Based on a Subdomain of Envelope Protein Erns for Serologic Diagnosis of Pestivirus Infections in Swine

Science.gov (United States)

Langedijk, J. P. M.; Middel, W. G. J.; Meloen, R. H.; Kramps, J. A.; de Smit, J. A.

2001-01-01

Peptides deduced from the C-terminal end (residues 191 to 227) of pestivirus envelope protein Erns were used to develop enzyme-linked immunosorbent assays (ELISAs) to measure specifically antibodies against different types of pestiviruses. The choice of the peptide was based on the modular structure of the Erns protein, and the peptide was selected for its probable independent folding and good exposure, which would make it a good candidate for an antigenic peptide to be used in a diagnostic test. A solid-phase peptide ELISA which was cross-reactive for several types of pestivirus antibodies and which can be used for the general detection of pestivirus antibodies was developed. To identify type-specific pestivirus antibodies, a liquid-phase peptide ELISA, with a labeled, specific classical swine fever virus (CSFV) peptide and an unlabeled bovine viral diarrhea virus peptide to block cross-reactivity, was developed. Specificity and sensitivity of the liquid-phase peptide ELISA for CSFV were 98 and 100%, respectively. Because the peptide is a fragment of the Erns protein, it can be used to differentiate between infected and vaccinated animals when a vaccine based on the E2 protein, which is another pestivirus envelope protein, is used. PMID:11230402

Solvent Composition is Critical for Carbodiimide Cross-Linking of Hyaluronic Acid as an Ophthalmic Biomaterial

Directory of Open Access Journals (Sweden)

Jui-Yang Lai

2012-10-01

Full Text Available Hyaluronic acid (HA is one of the most important ophthalmic biomaterials, while also being used for tissue engineering and drug delivery. Although chemical cross-linking is an effective way to improve the material performance, it may as a consequence be detrimental to the living cells/tissues. Given that the cross-linking efficiency is mediated by the solvent composition during the chemical modification, this study aims to explore the stability and biocompatibility of carbodiimide cross-linked HA in relation to material processing conditions by varying the acetone/water volume ratio (from 70:30 to 95:5 at a constant 1-ethyl-3-(3-dimethyl aminopropyl carbodiimide (EDC concentration of 100 mM. Our results indicated that after the EDC treatment in the presence of an acetone/water mixture (85:15, v/v, the HA hydrogel membranes have the lowest equilibrium water content, the highest stress at break and the greatest resistance to hyaluronidase digestion. Live/Dead assays and pro-inflammatory cytokine expression analyses showed that the cross-linked HA hydrogel membranes, irrespective of the solvent composition, are compatible with human RPE cell lines without causing toxicity and inflammation. However, it should be noted that the test samples prepared by the cross-linking in the presence of acetone/water mixtures containing 70, 75, and 95 vol % of acetone slightly inhibit the metabolic activity of viable ARPE-19 cultures, probably due to the alteration in the ionic interaction between the medium nutrients and polysaccharide biomaterials. In summary, the water content, mechanical strength and RPE cell proliferative capacity strongly depends on the solvent composition for carbodiimide cross-linking of HA materials.

Competition between dewetting and cross-linking in poly(N-vinylpyrrolidone)/polystyrene bilayer films.

Science.gov (United States)

Telford, Andrew M; Thickett, Stuart C; James, Michael; Neto, Chiara

2011-12-06

We investigated the dewetting of metastable poly(N-vinylpyrrolidone) (PNVP) thin films (45 nm) on top of polystyrene (PS) thin films (58 nm) as a function of annealing temperature and molecular weight of PS (96 and 6850 kg/mol). We focused on the competition between dewetting, occurring as a result of unfavorable intermolecular interactions at the PNVP/PS interface, and spontaneous cross-linking of PNVP, occurring during thermal annealing, as we recently reported (Telford, A. M.; James, M.; Meagher, L.; Neto, C. ACS Appl. Mater. Interfaces 2010, 2, 2399-2408). Using optical microscopy, we studied how the dewetting morphology and dynamics at different temperatures depended on the relative viscosity of the top PNVP film, which increased with cross-linking time, and of the bottom PS film. In the PNVP/PS96K system, cross-linking dominated over dewetting at temperatures below 180 °C, reducing drastically nucleated hole density and their maximum size, while above 180 °C the two processes reversed, with complete dewetting occurring at 200 °C. On the other hand, the PNVP/PS6850K system never achieved advanced dewetting stages as the dewetting was slower than cross-linking in the investigated temperature range. In both systems, dewetting of the PNVP films could be avoided altogether by thermally annealing the bilayers at temperatures where cross-linking dominated. The cross-linking was characterized quantitatively using neutron reflectometry, which indicated shrinkage and densification of the PNVP film, and qualitatively through selective removal of the bottom PS film. A simple model accounting for progressive cross-linking during the dewetting process predicted well the observed hole growth profiles and produced estimates of the PNVP cross-linking rate coefficients and of the activation energy of the process, in good agreement with literature values for similar systems. © 2011 American Chemical Society

Laccase-Based CLEAs: Chitosan as a Novel Cross-Linking Agent

Directory of Open Access Journals (Sweden)

Alexandre Arsenault

2011-01-01

Full Text Available Laccase from Coriolopsis Polyzona was insolubilized as cross-linked enzyme aggregates (CLEAs for the first time with chitosan as the cross-linking agent. Concentrations between 0.01 and 1.867 g/L of chitosan were used and between 0.05 and 600 mM of 1-ethyl-3-(3-dimethylaminopropylcarbodiimide hydrochloride. The laccase was precipitated using ammonium sulphate and cross-linked simultaneously. Specific activity and thermal stability of these biocatalysts were measured. Activities of up to 737 U/g were obtained when 2,2-azino-bis-(3-ethylbenzthiazoline-6-sulfonic acid (ABTS was used as a substrate. Moreover, the stability of these biocatalysts was improved with regards to thermal degradation compared to free laccase when exposed to denaturing conditions of high temperature and low pH. The CLEAs stability against chemical denaturants was also tested but no significant improvement was detected. The total amount of ABTS to be oxidized during thermal degradation by CLEAs and free laccase was calculated and the insolubilized enzymes were reported to oxidize more substrate than free laccase. The formation conditions were analyzed by response surface methodology in order to determine an optimal environment for the production of efficient laccase-based CLEAs using chitosan as the cross-linking agent. After 24 hours of formation at pH 3 and at 4°C without agitation, the CLEAs exhibit the best specific activity.

Concentration-dependent oligomerization of cross-linked complexes between ferredoxin and ferredoxin–NADP+ reductase

International Nuclear Information System (INIS)

Kimata-Ariga, Yoko; Kubota-Kawai, Hisako; Lee, Young-Ho; Muraki, Norifumi; Ikegami, Takahisa; Kurisu, Genji; Hase, Toshiharu

2013-01-01

Highlights: •Cross-linked complexes of ferredoxin (Fd) and Fd–NADP + reductase form oligomers. •In the crystal structures, Fd- and FNR moieties swap across the molecules. •The complexes exhibit concentration-dependent oligomerization at sub-milimolar order. -- Abstract: Ferredoxin–NADP + reductase (FNR) forms a 1:1 complex with ferredoxin (Fd), and catalyzes the electron transfer between Fd and NADP + . In our previous study, we prepared a series of site-specifically cross-linked complexes of Fd and FNR, which showed diverse electron transfer properties. Here, we show that X-ray crystal structures of the two different Fd–FNR cross-linked complexes form oligomers by swapping Fd and FNR moieties across the molecules; one complex is a dimer from, and the other is a successive multimeric form. In order to verify whether these oligomeric structures are formed only in crystal, we investigated the possibility of the oligomerization of these complexes in solution. The mean values of the particle size of these cross-linked complexes were shown to increase with the rise of protein concentration at sub-milimolar order, whereas the size of dissociable wild-type Fd:FNR complex was unchanged as analyzed by dynamic light scattering measurement. The oligomerization products were detected by SDS–PAGE after chemical cross-linking of these complexes at the sub-milimolar concentrations. The extent and concentration-dependent profile of the oligomerizaion were differentiated between the two cross-linked complexes. These results show that these Fd–FNR cross-linked complexes exhibit concentration-dependent oligomerization, possibly through swapping of Fd and FNR moieties also in solution. These findings lead to the possibility that some native multi-domain proteins may present similar phenomenon in vivo

Recent advances in corneal collagen cross-linking

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Gitansha Shreyas Sachdev

2017-01-01

Full Text Available Corneal collagen cross-linking has become the preferred modality of treatment for corneal ectasia since its inception in late 1990s. Numerous studies have demonstrated the safety and efficacy of the conventional protocol. Our understanding of the cross-linking process is ever evolving, with its wide implications in the form of accelerated and pulsed protocols. Newer advancements in technology include various riboflavin formulations and the ability to deliver higher fluence protocols with customised irradiation patterns. A greater degree of customisation is likely the path forward, which will aim at achieving refractive improvements along with disease stability. The use of cross-linking for myopic correction is another avenue under exploration. Combination of half fluence cross-linking with refractive correction for high errors to prevent post LASIK regression is gaining interest. This review aims to highlight the various advancements in the cross-linking technology and its clinical applications.

Prediction of Xaa-Pro peptide bond conformation from sequence and chemical shifts

Energy Technology Data Exchange (ETDEWEB)

Shen Yang; Bax, Ad, E-mail: bax@nih.go [National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Laboratory of Chemical Physics (United States)

2010-03-15

We present a program, named Promega, to predict the Xaa-Pro peptide bond conformation on the basis of backbone chemical shifts and the amino acid sequence. Using a chemical shift database of proteins of known structure together with the PDB-extracted amino acid preference of cis Xaa-Pro peptide bonds, a cis/trans probability score is calculated from the backbone and {sup 13}C{sup {beta}} chemical shifts of the proline and its neighboring residues. For an arbitrary number of input chemical shifts, which may include Pro-{sup 13}C{sup {gamma}}, Promega calculates the statistical probability that a Xaa-Pro peptide bond is cis. Besides its potential as a validation tool, Promega is particularly useful for studies of larger proteins where Pro-{sup 13}C{sup {gamma}} assignments can be challenging, and for on-going efforts to determine protein structures exclusively on the basis of backbone and {sup 13}C{sup {beta}} chemical shifts.

Nitric oxide-induced interstrand cross-links in DNA.

Science.gov (United States)

Caulfield, Jennifer L; Wishnok, John S; Tannenbaum, Steven R

2003-05-01

The DNA damaging effects of nitrous acid have been extensively studied, and the formation of interstrand cross-links have been observed. The potential for this cross-linking to occur through a common nitrosating intermediate derived from nitric oxide is investigated here. Using a HPLC laser-induced fluorescence (LIF) system, the amount of interstrand cross-link formed on nitric oxide treatment of the 5'-fluorescein-labeled oligomer ATATCGATCGATAT was determined. This self-complimentary sequence contains two 5'-CG sequences, which is the preferred site for nitrous acid-induced cross-linking. Nitric oxide was delivered to an 0.5 mM oligomer solution at 15 nmol/mL/min to give a final nitrite concentration of 652 microM. The resulting concentration of the deamination product, xanthine, in this sample was found to be 211 +/- 39 nM, using GC/MS, and the amount of interstrand cross-link was determined to be 13 +/- 2.5 nM. Therefore, upon nitric oxide treatment, the cross-link is found at approximately 6% of the amount of the deamination product. Using this system, detection of the cross-link is also possible for significantly lower doses of nitric oxide, as demonstrated by treatment of the same oligomer with NO at a rate of 18 nmol/mL/min resulting in a final nitrite concentration of 126 microM. The concentration of interstrand cross-link was determined to be 3.6 +/- 0.1 nM in this sample. Therefore, using the same dose rate, when the total nitric oxide concentration delivered drops by a factor of approximately 5, the concentration of cross-link drops by a factor of about 4-indicating a qausi-linear response. It may now be possible to predict the number of cross-links in a small genome based on the number of CpG sequences and the yield of xanthine derived from nitrosative deamination.

Polymers and Cross-Linking: A CORE Experiment to Help Students Think on the Submicroscopic Level

Science.gov (United States)

Bruce, Mitchell R. M.; Bruce, Alice E.; Avargil, Shirly; Amar, Francois G.; Wemyss, Thomas M.; Flood, Virginia J.

2016-01-01

The Polymers and Cross-Linking experiment is presented via a new three phase learning cycle: CORE (Chemical Observations, Representations, Experimentation), which is designed to model productive chemical inquiry and to promote a deeper understanding about the chemistry operating at the submicroscopic level. The experiment is built on two familiar…

Cross-reactive microbial peptides can modulate HIV-specific CD8+ T cell responses.

Directory of Open Access Journals (Sweden)

Christopher W Pohlmeyer

Full Text Available Heterologous immunity is an important aspect of the adaptive immune response. We hypothesized that this process could modulate the HIV-1-specific CD8+ T cell response, which has been shown to play an important role in HIV-1 immunity and control. We found that stimulation of peripheral blood mononuclear cells (PBMCs from HIV-1-positive subjects with microbial peptides that were cross-reactive with immunodominant HIV-1 epitopes resulted in dramatic expansion of HIV-1-specific CD8+ T cells. Interestingly, the TCR repertoire of HIV-1-specific CD8+ T cells generated by ex vivo stimulation of PBMCs using HIV-1 peptide was different from that of cells stimulated with cross-reactive microbial peptides in some HIV-1-positive subjects. Despite these differences, CD8+ T cells stimulated with either HIV-1 or cross-reactive peptides effectively suppressed HIV-1 replication in autologous CD4+ T cells. These data suggest that exposure to cross-reactive microbial antigens can modulate HIV-1-specific immunity.

Position of residues in transmembrane peptides with respect to the lipid bilayer: A combined lipid NOEs and water chemical exchange approach in phospholipid bicelles

International Nuclear Information System (INIS)

Glover, Kerney Jebrell; Whiles, Jennifer A.; Vold, Regitze R.; Melacini, Giuseppe

2002-01-01

The model transmembrane peptide P16 was incorporated into small unaligned phospholipid bicelles, which provide a 'native-like' lipid bilayer compatible with high-resolution solution NMR techniques. Using amide-water chemical exchange and amide-lipid cross-relaxation measurements, the interactions between P16 and bicelles were investigated. Distinctive intermolecular NOE patterns observed in band-selective 2D-NOESY spectra of bicellar solutions with several lipid deuteration schemes indicated that P16 is preferentially interacting with the 'bilayered' region of the bicelle rather than with the rim. Furthermore, when amide-lipid NOEs were combined with amide-water chemical exchange cross-peaks of selectively 15 N-labeled P16 peptides, valuable information was obtained about the position of selected residues relative to the membrane-water interface. Specifically, three main classes were identified. Class I residues lie outside the bilayer and show amide-water exchange cross-peaks but no amide-lipid NOEs. Class II residues reside in the bilayer-water interface and show both amide-water exchange cross-peaks and amide-lipid NOEs. Class III residues are embedded within the hydrophobic core of the membrane and show no amide-water exchange cross-peaks but strong amide-lipid NOEs

Radiation-induced DNA-protein cross-links: Mechanisms and biological significance.

Science.gov (United States)

Nakano, Toshiaki; Xu, Xu; Salem, Amir M H; Shoulkamy, Mahmoud I; Ide, Hiroshi

2017-06-01

Ionizing radiation produces various DNA lesions such as base damage, DNA single-strand breaks (SSBs), DNA double-strand breaks (DSBs), and DNA-protein cross-links (DPCs). Of these, the biological significance of DPCs remains elusive. In this article, we focus on radiation-induced DPCs and review the current understanding of their induction, properties, repair, and biological consequences. When cells are irradiated, the formation of base damage, SSBs, and DSBs are promoted in the presence of oxygen. Conversely, that of DPCs is promoted in the absence of oxygen, suggesting their importance in hypoxic cells, such as those present in tumors. DNA and protein radicals generated by hydroxyl radicals (i.e., indirect effect) are responsible for DPC formation. In addition, DPCs can also be formed from guanine radical cations generated by the direct effect. Actin, histones, and other proteins have been identified as cross-linked proteins. Also, covalent linkages between DNA and protein constituents such as thymine-lysine and guanine-lysine have been identified and their structures are proposed. In irradiated cells and tissues, DPCs are repaired in a biphasic manner, consisting of fast and slow components. The half-time for the fast component is 20min-2h and that for the slow component is 2-70h. Notably, radiation-induced DPCs are repaired more slowly than DSBs. Homologous recombination plays a pivotal role in the repair of radiation-induced DPCs as well as DSBs. Recently, a novel mechanism of DPC repair mediated by a DPC protease was reported, wherein the resulting DNA-peptide cross-links were bypassed by translesion synthesis. The replication and transcription of DPC-bearing reporter plasmids are inhibited in cells, suggesting that DPCs are potentially lethal lesions. However, whether DPCs are mutagenic and induce gross chromosomal alterations remains to be determined. Copyright © 2017 Elsevier Inc. All rights reserved.

Chemical crosslinking and mass spectrometry studies of the structure and dynamics of membrane proteins and receptors.

Energy Technology Data Exchange (ETDEWEB)

Haskins, William E.; Leavell, Michael D.; Lane, Pamela; Jacobsen, Richard B.; Hong, Joohee; Ayson, Marites J.; Wood, Nichole L.; Schoeniger, Joseph S.; Kruppa, Gary Hermann; Sale, Kenneth L.; Young, Malin M.; Novak, Petr

2005-03-01

Membrane proteins make up a diverse and important subset of proteins for which structural information is limited. In this study, chemical cross-linking and mass spectrometry were used to explore the structure of the G-protein-coupled photoreceptor bovine rhodopsin in the dark-state conformation. All experiments were performed in rod outer segment membranes using amino acid 'handles' in the native protein sequence and thus minimizing perturbations to the native protein structure. Cysteine and lysine residues were covalently cross-linked using commercially available reagents with a range of linker arm lengths. Following chemical digestion of cross-linked protein, cross-linked peptides were identified by accurate mass measurement using liquid chromatography-fourier transform mass spectrometry and an automated data analysis pipeline. Assignments were confirmed and, if necessary, resolved, by tandem MS. The relative reactivity of lysine residues participating in cross-links was evaluated by labeling with NHS-esters. A distinct pattern of cross-link formation within the C-terminal domain, and between loop I and the C-terminal domain, emerged. Theoretical distances based on cross-linking were compared to inter-atomic distances determined from the energy-minimized X-ray crystal structure and Monte Carlo conformational search procedures. In general, the observed cross-links can be explained by re-positioning participating side-chains without significantly altering backbone structure. One exception, between C3 16 and K325, requires backbone motion to bring the reactive atoms into sufficient proximity for cross-linking. Evidence from other studies suggests that residues around K325 for a region of high backbone mobility. These findings show that cross-linking studies can provide insight into the structural dynamics of membrane proteins in their native environment.

The First MS-Cleavable, Photo-Thiol-Reactive Cross-Linker for Protein Structural Studies

Science.gov (United States)

Iacobucci, Claudio; Piotrowski, Christine; Rehkamp, Anne; Ihling, Christian H.; Sinz, Andrea

2018-04-01

Cleavable cross-linkers are gaining increasing importance for chemical cross-linking/mass spectrometry (MS) as they permit a reliable and automated data analysis in structural studies of proteins and protein assemblies. Here, we introduce 1,3-diallylurea (DAU) as the first CID-MS/MS-cleavable, photo-thiol-reactive cross-linker. DAU is a commercially available, inexpensive reagent that efficiently undergoes an anti-Markovnikov hydrothiolation with cysteine residues in the presence of a radical initiator upon UV-A irradiation. Radical cysteine cross-linking proceeds via an orthogonal "click reaction" and yields stable alkyl sulfide products. DAU reacts at physiological pH and cross-linking reactions with peptides, and proteins can be performed at temperatures as low as 4 °C. The central urea bond is efficiently cleaved upon collisional activation during tandem MS experiments generating characteristic product ions. This improves the reliability of automated cross-link identification. Different radical initiators have been screened for the cross-linking reaction of DAU using the thiol-containing compounds cysteine and glutathione. Our concept has also been exemplified for the biologically relevant proteins bMunc13-2 and retinal guanylyl cyclase-activating protein-2. [Figure not available: see fulltext.

Cross-linked structure of network evolution

International Nuclear Information System (INIS)

Bassett, Danielle S.; Wymbs, Nicholas F.; Grafton, Scott T.; Porter, Mason A.; Mucha, Peter J.

2014-01-01

We study the temporal co-variation of network co-evolution via the cross-link structure of networks, for which we take advantage of the formalism of hypergraphs to map cross-link structures back to network nodes. We investigate two sets of temporal network data in detail. In a network of coupled nonlinear oscillators, hyperedges that consist of network edges with temporally co-varying weights uncover the driving co-evolution patterns of edge weight dynamics both within and between oscillator communities. In the human brain, networks that represent temporal changes in brain activity during learning exhibit early co-evolution that then settles down with practice. Subsequent decreases in hyperedge size are consistent with emergence of an autonomous subgraph whose dynamics no longer depends on other parts of the network. Our results on real and synthetic networks give a poignant demonstration of the ability of cross-link structure to uncover unexpected co-evolution attributes in both real and synthetic dynamical systems. This, in turn, illustrates the utility of analyzing cross-links for investigating the structure of temporal networks

Cross-linked structure of network evolution

Energy Technology Data Exchange (ETDEWEB)

Bassett, Danielle S., E-mail: dsb@seas.upenn.edu [Department of Bioengineering, University of Pennsylvania, Philadelphia, Pennsylvania 19104 (United States); Department of Physics, University of California, Santa Barbara, California 93106 (United States); Sage Center for the Study of the Mind, University of California, Santa Barbara, California 93106 (United States); Wymbs, Nicholas F.; Grafton, Scott T. [Department of Psychology and UCSB Brain Imaging Center, University of California, Santa Barbara, California 93106 (United States); Porter, Mason A. [Oxford Centre for Industrial and Applied Mathematics, Mathematical Institute, University of Oxford, Oxford OX2 6GG (United Kingdom); CABDyN Complexity Centre, University of Oxford, Oxford, OX1 1HP (United Kingdom); Mucha, Peter J. [Carolina Center for Interdisciplinary Applied Mathematics, Department of Mathematics, University of North Carolina, Chapel Hill, North Carolina 27599 (United States); Department of Applied Physical Sciences, University of North Carolina, Chapel Hill, North Carolina 27599 (United States)

2014-03-15

We study the temporal co-variation of network co-evolution via the cross-link structure of networks, for which we take advantage of the formalism of hypergraphs to map cross-link structures back to network nodes. We investigate two sets of temporal network data in detail. In a network of coupled nonlinear oscillators, hyperedges that consist of network edges with temporally co-varying weights uncover the driving co-evolution patterns of edge weight dynamics both within and between oscillator communities. In the human brain, networks that represent temporal changes in brain activity during learning exhibit early co-evolution that then settles down with practice. Subsequent decreases in hyperedge size are consistent with emergence of an autonomous subgraph whose dynamics no longer depends on other parts of the network. Our results on real and synthetic networks give a poignant demonstration of the ability of cross-link structure to uncover unexpected co-evolution attributes in both real and synthetic dynamical systems. This, in turn, illustrates the utility of analyzing cross-links for investigating the structure of temporal networks.

Current status of accelerated corneal cross-linking

Directory of Open Access Journals (Sweden)

Michael Mrochen

2013-01-01

Full Text Available Corneal cross-linking with riboflavin is a technique to stabilize or reduce corneal ectasia, in diseases such as keratoconus and post-laser-assisted in situ keratomileusis (LASIK ectasia. There is an interest by patient as well as clinicians to reduce the overall treatment time. Especially, the introduction of corneal cross-linking in combination with corneal laser surgery demands a shorter treatment time to assure a sufficient patient flow. The principles and techniques of accelerated corneal cross-linking is discussed.

Efficient and Selective Chemical Labeling of Electrochemically Generated Peptides Based on Spirolactone Chemistry

NARCIS (Netherlands)

Zhang, Tao; Niu, Xiaoyu; Yuan, Tao; Tessari, Marco; de Vries, Marcel P.; Permentier, Hjalmar P.; Bischoff, Rainer

2016-01-01

Specific digestion of proteins is an essential step for mass spectrometry-based proteomics, and the chemical labeling of the resulting peptides is often used for peptide enrichment or the introduction of desirable tags. Cleavage of the peptide bond following electrochemical oxidation of Tyr or Trp

Shaped articles of cross-linked fluorocarbon polymers

International Nuclear Information System (INIS)

Gotcher, A.J.; Germeraad, P.B.

1981-01-01

A process is described which comprises (1) contacting (a) a shaped article of a polymeric composition wherein the polymer is a fluorocarbon polymer having a melting point of at least 200 0 C, the article having a tensile strength of at least 3,000 psi, with (b) a fluid composition comprising a cross-linking agent, until the article contains at least 2.5% by weight of the cross-linking agent; and (2) irradiating the shaped article with ionising radiation to a dosage not exceeding 50 Mrads under conditions such that the composition is cross-linked sufficiently to impart thereto an M 100 value of at least 300 psi, while maintaining a tensile strength of at least 3000 psi, the shaped article containing a specified proportion of the cross-linking agent. (author)

Using 1H and 13C NMR chemical shifts to determine cyclic peptide conformations: a combined molecular dynamics and quantum mechanics approach.

Science.gov (United States)

Nguyen, Q Nhu N; Schwochert, Joshua; Tantillo, Dean J; Lokey, R Scott

2018-05-10

Solving conformations of cyclic peptides can provide insight into structure-activity and structure-property relationships, which can help in the design of compounds with improved bioactivity and/or ADME characteristics. The most common approaches for determining the structures of cyclic peptides are based on NMR-derived distance restraints obtained from NOESY or ROESY cross-peak intensities, and 3J-based dihedral restraints using the Karplus relationship. Unfortunately, these observables are often too weak, sparse, or degenerate to provide unequivocal, high-confidence solution structures, prompting us to investigate an alternative approach that relies only on 1H and 13C chemical shifts as experimental observables. This method, which we call conformational analysis from NMR and density-functional prediction of low-energy ensembles (CANDLE), uses molecular dynamics (MD) simulations to generate conformer families and density functional theory (DFT) calculations to predict their 1H and 13C chemical shifts. Iterative conformer searches and DFT energy calculations on a cyclic peptide-peptoid hybrid yielded Boltzmann ensembles whose predicted chemical shifts matched the experimental values better than any single conformer. For these compounds, CANDLE outperformed the classic NOE- and 3J-coupling-based approach by disambiguating similar β-turn types and also enabled the structural elucidation of the minor conformer. Through the use of chemical shifts, in conjunction with DFT and MD calculations, CANDLE can help illuminate conformational ensembles of cyclic peptides in solution.

HPLC detection of loss rate and cell migration of HUVECs in a proanthocyanidin cross-linked recombinant human collagen-peptide (RHC)–chitosan scaffold

Energy Technology Data Exchange (ETDEWEB)

Zhang, Jing; Deng, Aipeng [School of Environmental and Biological Engineering, Nanjing University of Science and Technology, Nanjing 210094 (China); Yang, Yang [Faculty of Engineering, University of Nottingham, Nottingham NG7 2RD (United Kingdom); Gao, Lihu; Xu, Na; Liu, Xin; Hu, Lunxiang; Chen, Junhua [School of Environmental and Biological Engineering, Nanjing University of Science and Technology, Nanjing 210094 (China); Yang, Shulin, E-mail: yshulin@njust.edu.cn [School of Environmental and Biological Engineering, Nanjing University of Science and Technology, Nanjing 210094 (China)

2015-11-01

Porous scaffolds with appropriate pore structure, biocompatibility, mechanical property and processability play an important role in tissue engineering. In this paper, we fabricated a recombinant human collagen-peptide (RHC)–chitosan scaffold cross-linked by premixing 30% proanthocyanidin (PA) in one-step freeze-drying. To remove the residual acetic acid, optimized 0.2 M phosphate buffer of pH 6.24 with 30% ethanol (PBSE) was selected to neutralize the lyophilized scaffold followed by three times deionized water rinse. Ninhydrin assay was used to characterize the components loss during the fabrication process. To detect the exact RHC loss under optimized neutralization condition, high performance liquid chromatography (HPLC) equipped size exclusion chromatography column was used and the total RHC loss rate through PBSE rinse was 19.5 ± 5.08%. Fourier transform infrared spectroscopy (FT-IR) indicated hydrogen bonding among RHC, chitosan and PA, it also presented a probative but not strong hydrophobic interaction between phenyl rings of polyphenols and pyrrolidine rings of proline in RHC. Further, human umbilical vein endothelial cell (HUVEC) viability analyzed by a scanning electron microscope (SEM) and acridine orange/ethidium bromide (AO/EB) fluorescence staining exhibited that this scaffold could not only promote cell proliferation on scaffold surface but also permit cells migration into the scaffold. qRT-PCR exhibited that the optimized scaffold could stimulate angiogenesis associated genes VEGF and CD31 expression. These characterizations indicated that this scaffold can be considered as an ideal candidate for tissue engineering. - Highlights: • PA cross-linked recombinant human collagen–chitosan scaffold. • Fabrication in one-step lyophilization with neutralization. • HPLC detection of RHC loss rate • HUVEC proliferation and migration in scaffold • Angiogenesis associated gene expressions were increased in scaffold cell culturing.

Quantum molecular dynamics of methyl rotors in peptide links

International Nuclear Information System (INIS)

Del-Mar, Jon

2002-01-01

A particles wavefunction extends beyond the classically accessible regions of the potential energy surface. Quantum mechanical tunnelling is the result of this partial delocalisation, which enables the surpassing of classically inaccessible potential barriers. A particles mass is an important aspect, reflecting the tunnelling probability; a consequence of this is that a proton is ideally suited to this behaviour. Symmetrical molecular rotors such as Ch 3 provide a clear example of quantum mechanical tunnelling, seen in their motional spectrum. The advantage of the methyl rotor is that it's found in a wide range of organic compounds, giving a wide range in hindering potentials. It is effectively a proton rotor, and is easily observed using techniques such as Nuclear Magnetic Resonance (NMR), and Inelastic Neutron Scattering (INS). Both NMR and INS techniques are sensitive to molecular motion, and as they measure the tunnel frequencies in different energy windows, are complementary. Of central importance to many biological processes and structures is the peptide unit, -CONH-. Of particular significance are the intermolecular networks that are often formed by the NHO hydrogen bonds, the peptide links. The molecules were chosen for the research in this thesis to form a tractable model for polypeptides and alpha-helix proteins. Methyl rotor tunnelling frequencies have been used, which are very sensitive to the potential energy surface, as a probe of the electronic and molecular structure associated with the peptide links. Quantum chemistry calculations were then utilized to connect experiments to theory to learn about the hydrogen bond. (author)

Cross-linked aromatic cationic polymer electrolytes with enhanced stability for high temperature fuel cell applications

DEFF Research Database (Denmark)

Ma, Wenjia; Zhao, Chengji; Yang, Jingshuai

2012-01-01

Diamine-cross-linked membranes were prepared from cross-linkable poly(arylene ether ketone) containing pendant cationic quaternary ammonium group (QPAEK) solution by a facile and general thermal curing method using 4,4′-diaminodiphenylmethane with rigid framework and 1,6-diaminohexane with flexible...... anchoring of the molecule. Combining the excellent thermal stability, the addition of a small amount of diamines enhanced both the chemical and mechanical stability and the phosphoric acid doping (PA) ability of membranes. Fuel cell performance based on impregnated cross-linked membranes have been...... successfully operated at temperatures up to 120 °C and 180 °C with unhumidified hydrogen and air under ambient pressure, the maximum performance of diamine-cross-linked membrane is observed at 180 °C with a current density of 1.06 A cm−2 and the peak power density of 323 mW cm−2. The results also indicate...

Affect the Cross Linking Degree and Polymer Composition on the Mechanical Properties of Poly (vinyl alcohol/ Pullu-lan Films

Directory of Open Access Journals (Sweden)

Shemaa Abdul Satar Soud

2018-04-01

Full Text Available In this study Cross-linked PVA/Pullulan film was prepared. Cross-linking reaction done by addi-tion of gluteraldehyde at different reaction time (10,30and 60 min. Chemical interaction, me-chanical, thermal properties, water solubility and film morphology was studied for cross-linked PVA/Pullulan, PVA and Pullulan only. Thus FTIR investigated formation of hydrogen bonding between pullulan and PVA with (GA. Tensile strength, tensile modulus and elongation (% at break for PVA/Pullulan film was improved with addition of (GA as the reaction time proceed equivalent with increasing PVA content

Donor cross-linking for keratoplasty: a laboratory evaluation.

Science.gov (United States)

Mukherjee, Achyut; Hayes, Sally; Aslanides, Ioannis; Lanchares, Elena; Meek, Keith M

2015-12-01

This laboratory-based investigation compares the topographic outcomes of conventional penetrating keratoplasty with that of a novel procedure in which donor corneas are cross-linked prior to keratoplasty. Penetrating keratoplasty procedures with continuous running sutures were carried out in a porcine whole globe model. Sixty eyes were randomly paired as 'donor' and 'host' tissue before being assigned to one of two groups. In the cross-linked group, donor corneas underwent riboflavin/UVA cross-linking prior to being trephined and sutured to untreated hosts. In the conventional keratoplasty group, both host and donor corneas remained untreated prior to keratoplasty. Topographic and corneal wavefront measurements were performed following surgery, and technical aspects of the procedure evaluated. Mean keratometric astigmatism was significantly lower in the cross-linked donor group at 3.67D (SD 1.8 D), vs. 8.43 D (SD 2.4 D) in the conventional keratoplasty group (p < 0.005). Mean wavefront astigmatism was also significantly reduced in the cross-linked donor group 4.71 D (SD 2.1) vs. 8.29D (SD 3.6) in the conventional keratoplasty group (p < 0.005). Mean RMS higher order aberration was significantly lower in the cross-linked donor group at 1.79 um (SD 0.98), vs. 3.05 um (SD 1.9) in the conventional keratoplasty group (P = 0.02). Qualitative analysis revealed less tissue distortion at the graft-host junction in the cross-linked group. Cross-linking of donor corneas prior to keratoplasty reduces intraoperative induced astigmatism and aberrations in an animal model. Further studies are indicated to evaluate the implications of this potential modification of keratoplasty surgery.

Ionic interaction of myosin loop 2 with residues located beyond the N-terminal part of actin probed by chemical cross-linking.

Science.gov (United States)

Pliszka, Barbara; Martin, Brian M; Karczewska, Emilia

2008-02-01

To probe ionic contacts of skeletal muscle myosin with negatively charged residues located beyond the N-terminal part of actin, myosin subfragment 1 (S1) and actin split by ECP32 protease (ECP-actin) were cross-linked with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC). We have found that unmodified S1 can be cross-linked not only to the N-terminal part, but also to the C-terminal 36 kDa fragment of ECP-actin. Subsequent experiments performed on S1 cleaved by elastase or trypsin indicate that the cross-linking site in S1 is located within loop 2. This site is composed of Lys-636 and Lys-637 and can interact with negatively charged residues of the 36 kDa actin fragment, most probably with Glu-99 and Glu-100. Cross-links are formed both in the absence and presence of MgATP.P(i) analog, although the addition of nucleotide decreases the efficiency of the cross-linking reaction.

Biocatalytic cross-linking of pectic polysaccharides for designed food functionality

DEFF Research Database (Denmark)

Zaidel, Dayang Norulfairuz Abang; Meyer, Anne S.

2012-01-01

the mechanisms of formation of functional pectic polysaccharide cross-links, including covalent cross-links (notably phenolic esters and uronyl ester linkages) and non-covalent, ionic cross-links (which involve calcium and borate ester links). The treatise examines how such cross-links can be designed via......Recent research has demonstrated how cross-linking of pectic polysaccharides to obtain gel formation can be promoted by enzymatic catalysis reactions, and provide opportunities for functional upgrading of pectic polysaccharides present in agro-industrial sidestreams. This review highlights...... specific enzymatic reactions, and highlights the most recent data concerning enzyme catalyzed engineering of cross-links for in situ structural design of functional properties of foods....

Interactions of photoactive DNAs with terminal deoxynucleotidyl transferase: Identification of peptides in the DNA binding domain

International Nuclear Information System (INIS)

Farrar, Y.J.K.; Evans, R.K.; Beach, C.M.; Coleman, M.S.

1991-01-01

Terminal deoxynucleotidyl transferase (terminal transferase) was specifically modified in the DNA binding site by a photoactive DNA substrate (hetero-40-mer duplex containing eight 5-azido-dUMP residues at one 3' end). Under optimal photolabeling conditions, 27-40% of the DNA was covalently cross-linked to terminal transferase. The specificity of the DNA and protein interaction was demonstrated by protection of photolabeling at the DNA binding domain with natural DNA substrates. In order to recover high yields of modified peptides from limited amounts of starting material, protein modified with 32 P-labeled photoactive DNA and digested with trypsin was extracted 4 times with phenol followed by gel filtration chromatography. All peptides not cross-linked to DNA were extracted into the phenol phase while the photolyzed DNA and the covalently cross-linked peptides remained in the aqueous phase. The 32 P-containing peptide-DNA fraction was subjected to amino acid sequence analysis. Two sequences, Asp 221 -Lys 231 (peptide B8) and Cys 234 -Lys 249 (peptide B10), present in similar yield, were identified. Structure predictions placed the two peptides in an α-helical array of 39 angstrom which would accommodate a DNA helix span of 11 nucleotides. These peptides share sequence similarity with a region in DNA polymerase β that has been implicated in the binding of DNA template

Baking Performance of Phosphorylated Cross-Linked Resistant Starch in Low-Moisture Bakery Goods

Science.gov (United States)

Phosphorylated cross-linked resistant starch (RS) is a type 4 RS, which can be used for enhancing the benefits of dietary fiber. The baking performance of the RS was explored using wire-cut cookie baking and benchtop chemically-leavened cracker baking methods to produce low-moisture baked goods (coo...

Effects of cross-linking modification with phosphoryl chloride (POCl3 on pysiochemical properties of barely starch

Directory of Open Access Journals (Sweden)

Zahra Malekpour

2016-05-01

Full Text Available Chemical methods are one of the comon method in starch modification. This study aimed at investigating of cross-link affection of phosphoryl chloride with two different levels 0.5 and 1g.kg-1 in order to enhance funciotnal proeprties and physiochemical changes on extracted starch from barely variety Bahman which cultivates in Chahr-Mahal Bakhtiari Province of Iran. Obtained results indicated that cross-linking leads to reduce sweeling power of strach granuls compred to natural starch and the amount of reduciton increase via the substitituin level. Powerfull cross-linkingnetween starch chains casue more resistance of granules to seweeling which is increased by means of cross-linking dgree. Additioally,  investigationresults from synersis revealed that releasing water percentage in cross-linked starches increase in comparison to natural starches and this amount depends onthe amount of cross-link surface with a significantly difference in (α <0.05. Gelatinization temperature in both levels negligibly increased by modification where in low level of cross-linking was more. Furthermoe evaluating gelation temperatures of both natural and cross-linked modified starches showed that addition of phosphate groups in starch and creating extra coovalent bonds make granues more compressed reulting in slight increase of To, Tp, Tcin barely starch. Icreasing of temperature observed more in less concentration of cross-links. Evaluation of viscosity changes also revealed that this modification depending on increasing the amount of Phosphoryl Chloride led to increasing peak temperature, diminish peak and setback viscosity. Result also exhibited that in morphological level, cross-link causes to incidence changes in particles' diameter size. The comparison of diameter average and frequency between natural starch and cross-links starch exhibited that in cross-linkd treatment with 0.5% phosphoryl chloride, increase in frequency of granules with diameter of 6 - 10µm

Induction and repair of DNA cross-links induced by sulfur mustard in the A-549 cell line followed by a comet assay.

Science.gov (United States)

Jost, Petr; Svobodova, Hana; Stetina, Rudolf

2015-07-25

Sulfur mustard is a highly toxic chemical warfare agent with devastating impact on intoxicated tissues. DNA cross-links are probably the most toxic DNA lesions induced in the cell by sulfur mustard. The comet assay is a very sensitive method for measuring DNA damage. In the present study using the A-549 lung cell line, the comet assay protocol was optimized for indirect detection of DNA cross-links induced by sulfur mustard. The method is based on the additional treatment of the assayed cells containing cross-links with the chemical mutagen, styrene oxide. Alkali-labile adducts of styrene oxide cause DNA breaks leading to the formation of comets. A significant dose-dependent reduction of DNA migration of the comet's tail was found after exposing cells to sulfur mustard, indicative of the amount of sulfur mustard induced cross-links. The remarkable decrease of % tail DNA could be observed as early as 5min following exposure to sulfur mustard and the maximal effect was found after 30min, when DNA migration was reduced to the minimum. Sulfur mustard preincubated in culture medium without cells lost its ability to induce cross-links and had a half-life of about 15min. Pre-incubation longer than 30min does not lead to a significant increase in cross-links when applied to cells. However, the amount of cross-links is decreased during further incubation due to repair. The current modification of the comet assay provides a useful tool for detecting DNA cross-links induced by sulfur mustard and could be used for detection of other DNA cross-linking agents such as chemotherapeutic drugs. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Hydrogels Prepared from Cross-Linked Nanofibrillated Cellulose

Science.gov (United States)

Sandeep S. Nair; J.Y. Zhu; Yulin Deng; Arthur J. Ragauskas

2014-01-01

Nanocomposite hydrogels were developed by cross-linking nanofibrillated cellulose with poly(methyl vinyl ether-co-maleic acid) and polyethylene glycol. The cross-linked hydrogels showed enhanced water absorption and gel content with the addition of nanocellulose. In addition, the thermal stability, mechanical strength, and modulus increased with an increase in the...

DNA cross-linking by dehydromonocrotaline lacks apparent base sequence preference.

Science.gov (United States)

Rieben, W Kurt; Coulombe, Roger A

2004-12-01

Pyrrolizidine alkaloids (PAs) are ubiquitous plant toxins, many of which, upon oxidation by hepatic mixed-function oxidases, become reactive bifunctional pyrrolic electrophiles that form DNA-DNA and DNA-protein cross-links. The anti-mitotic, toxic, and carcinogenic action of PAs is thought to be caused, at least in part, by these cross-links. We wished to determine whether the activated PA pyrrole dehydromonocrotaline (DHMO) exhibits base sequence preferences when cross-linked to a set of model duplex poly A-T 14-mer oligonucleotides with varying internal and/or end 5'-d(CG), 5'-d(GC), 5'-d(TA), 5'-d(CGCG), or 5'-d(GCGC) sequences. DHMO-DNA cross-links were assessed by electrophoretic mobility shift assay (EMSA) of 32P endlabeled oligonucleotides and by HPLC analysis of cross-linked DNAs enzymatically digested to their constituent deoxynucleosides. The degree of DNA cross-links depended upon the concentration of the pyrrole, but not on the base sequence of the oligonucleotide target. Likewise, HPLC chromatograms of cross-linked and digested DNAs showed no discernible sequence preference for any nucleotide. Added glutathione, tyrosine, cysteine, and aspartic acid, but not phenylalanine, threonine, serine, lysine, or methionine competed with DNA as alternate nucleophiles for cross-linking by DHMO. From these data it appears that DHMO exhibits no strong base preference when forming cross-links with DNA, and that some cellular nucleophiles can inhibit DNA cross-link formation.

Composition of cross-linked 125I-follitropin-receptor complexes

Energy Technology Data Exchange (ETDEWEB)

Shin, J.; Ji, T.H.

1985-10-15

Both of the alpha and beta subunits of intact human follitropin (FSH) were radioiodinated with SVI-sodium iodide and chloramine-T and could be resolved on sodium dodecyl sulfate-polyacrylamide gels. Radioiodinated FSH was affinity-cross-linked with a cleavable (nondisulfide) homobifunctional reagent to its membrane receptor on the porcine granulosa cell surface as well as to a Triton X-100-solubilized form of the receptor. Cross-linked samples revealed three additional bands of slower electrophoretic mobility, corresponding to 65, 83, and 117 kDa, in addition to the hormone bands. The hormone alpha beta dimer band corresponded to 43 kDa. Formation of the three bands requires the SVI-hormone to bind specifically to the receptor with subsequent cross-linking. Binding was prevented by an excess of the native hormone but not by other hormones. A monofunctional analog of the cross-linking reagent failed to produce the three bands. Reagent concentration-dependent cross-linking revealed that their formation was sequential; smaller complexes formed first and then larger ones. When gels of cross-linked complexes were treated to cleave covalent cross-links and then electrophoresed in a second dimension, 18-, 22-, and 34-kDa components were released, in addition to the alpha and beta subunits of the hormone.

The release behavior and kinetic evaluation of tramadol HCl from chemically cross linked Ter polymeric hydrogels.

Science.gov (United States)

Malana, Muhammad A; Zohra, Rubab

2013-01-18

Hydrogels, being stimuli responsive are considered to be effective for targeted and sustained drug delivery. The main purpose for this work was to study the release behavior and kinetic evaluation of Tramadol HCl from chemically cross linked ter polymeric hydrogels. Ter-polymers of methacrylate, vinyl acetate and acrylic acid cross linked with ethylene glycol dimethacrylate (EGDMA) were prepared by free radical polymerization. The drug release rates, dynamic swelling behavior and pH sensitivity of hydrogels ranging in composition from 1-10 mol% EGDMA were studied. Tramadol HCl was used as model drug substance. The release behavior was investigated at pH 8 where all formulations exhibited non-Fickian diffusion mechanism. Absorbency was found to be more than 99% indicating good drug loading capability of these hydrogels towards the selected drug substance. Formulations designed with increasing amounts of EGDMA had a decreased equilibrium media content as well as media penetrating velocity and thus exhibited a slower drug release rate. Fitting of release data to different kinetic models indicate that the kinetic order shifts from the first to zero order as the concentration of drug was increased in the medium, showing gradual independency of drug release towards its concentration. Formulations with low drug content showed best fitness with Higuchi model whereas those with higher concentration of drug followed Hixson-Crowell model with better correlation values indicating that the drug release from these formulations depends more on change in surface area and diameter of tablets than that on concentration of the drug. Release exponent (n) derived from Korse-Meyer Peppas equation implied that the release of Tramadol HCl from these formulations was generally non-Fickian (n > 0.5 > 1) showing swelling controlled mechanism. The mechanical strength and controlled release capability of the systems indicate that these co-polymeric hydrogels have a great potential to

In Situ Cross-Linking of Stimuli-Responsive Hemicellulose Microgels during Spray Drying

Science.gov (United States)

2015-01-01

Chemical cross-linking during spray drying offers the potential for green fabrication of microgels with a rapid stimuli response and good blood compatibility and provides a platform for stimuli-responsive hemicellulose microgels (SRHMGs). The cross-linking reaction occurs rapidly in situ at elevated temperature during spray drying, enabling the production of microgels in a large scale within a few minutes. The SRHMGs with an average size range of ∼1–4 μm contain O-acetyl-galactoglucomannan as a matrix and poly(acrylic acid), aniline pentamer (AP), and iron as functional additives, which are responsive to external changes in pH, electrochemical stimuli, magnetic field, or dual-stimuli. The surface morphologies, chemical compositions, charge, pH, and mechanical properties of these smart microgels were evaluated using scanning electron microscopy, IR, zeta potential measurements, pH evaluation, and quantitative nanomechanical mapping, respectively. Different oxidation states were observed when AP was introduced, as confirmed by UV spectroscopy and cyclic voltammetry. Systematic blood compatibility evaluations revealed that the SRHMGs have good blood compatibility. This bottom-up strategy to synthesize SRHMGs enables a new route to the production of smart microgels for biomedical applications. PMID:25630464

In situ cross-linking of stimuli-responsive hemicellulose microgels during spray drying.

Science.gov (United States)

Zhao, Weifeng; Nugroho, Robertus Wahyu N; Odelius, Karin; Edlund, Ulrica; Zhao, Changsheng; Albertsson, Ann-Christine

2015-02-25

Chemical cross-linking during spray drying offers the potential for green fabrication of microgels with a rapid stimuli response and good blood compatibility and provides a platform for stimuli-responsive hemicellulose microgels (SRHMGs). The cross-linking reaction occurs rapidly in situ at elevated temperature during spray drying, enabling the production of microgels in a large scale within a few minutes. The SRHMGs with an average size range of ∼ 1-4 μm contain O-acetyl-galactoglucomannan as a matrix and poly(acrylic acid), aniline pentamer (AP), and iron as functional additives, which are responsive to external changes in pH, electrochemical stimuli, magnetic field, or dual-stimuli. The surface morphologies, chemical compositions, charge, pH, and mechanical properties of these smart microgels were evaluated using scanning electron microscopy, IR, zeta potential measurements, pH evaluation, and quantitative nanomechanical mapping, respectively. Different oxidation states were observed when AP was introduced, as confirmed by UV spectroscopy and cyclic voltammetry. Systematic blood compatibility evaluations revealed that the SRHMGs have good blood compatibility. This bottom-up strategy to synthesize SRHMGs enables a new route to the production of smart microgels for biomedical applications.

Effect of carbon nanotube dispersion on glass transition in cross-linked epoxy-carbon nanotube nanocomposites: role of interfacial interactions.

Science.gov (United States)

Khare, Ketan S; Khare, Rajesh

2013-06-20

We have used atomistic molecular simulations to study the effect of nanofiller dispersion on the glass transition behavior of cross-linked epoxy-carbon nanotube (CNT) nanocomposites. Specific chemical interactions at the interface of CNTs and cross-linked epoxy create an interphase region, whose impact on the properties of their nanocomposites increases with an increasing extent of dispersion. To investigate this aspect, we have compared the volumetric, structural, and dynamical properties of three systems: neat cross-linked epoxy, cross-linked epoxy nanocomposite containing dispersed CNTs, and cross-linked epoxy nanocomposite containing aggregated CNTs. We find that the nanocomposite containing dispersed CNTs shows a depression in the glass transition temperature (Tg) by ~66 K as compared to the neat cross-linked epoxy, whereas such a large depression is absent in the nanocomposite containing aggregated CNTs. Our results suggest that the poor interfacial interactions between the CNTs and the cross-linked epoxy matrix lead to a more compressible interphase region between the CNTs and the bulk matrix. An analysis of the resulting dynamic heterogeneity shows that the probability of percolation of immobile domains becomes unity near the Tg calculated from volumetric properties. Our observations also lend support to the conceptual analogy between polymer nanocomposites and the nanoconfinement of polymer thin films.

Importance of asparagine on the conformational stability and chemical reactivity of selected anti-inflammatory peptides

Energy Technology Data Exchange (ETDEWEB)

Soriano-Correa, Catalina, E-mail: csorico@comunidad.unam.mx [Química Computacional, Facultad de Estudios Superiores (FES)-Zaragoza, Universidad Nacional Autónoma de México (UNAM), Iztapalapa, C.P. 09230 México, D.F. (Mexico); Barrientos-Salcedo, Carolina [Laboratorio de Química Médica y Quimiogenómica, Facultad de Bioanálisis Campus Veracruz-Boca del Río, Universidad Veracruzana, C.P. 91700 Veracruz (Mexico); Campos-Fernández, Linda; Alvarado-Salazar, Andres [Química Computacional, Facultad de Estudios Superiores (FES)-Zaragoza, Universidad Nacional Autónoma de México (UNAM), Iztapalapa, C.P. 09230 México, D.F. (Mexico); Esquivel, Rodolfo O. [Departamento de Química, Universidad Autónoma Metropolitana-Iztapalapa (UAM-Iztapalapa), C.P. 09340 México, D.F. (Mexico)

2015-08-18

Highlights: • Asparagine plays an important role to anti-inflammatory effect of peptides. • The electron-donor substituent groups favor the formation of the hydrogen bonds, which contribute in the structural stability of peptides. • Chemical reactivity and the physicochemical features are crucial in the biological functions of peptides. - Abstract: Inflammatory response events are initiated by a complex series of molecular reactions that generate chemical intermediaries. The structure and properties of peptides and proteins are determined by the charge distribution of their side chains, which play an essential role in its electronic structure and physicochemical properties, hence on its biological functionality. The aim of this study was to analyze the effect of changing one central amino acid, such as substituting asparagine for aspartic acid, from Cys–Asn–Ser in aqueous solution, by assessing the conformational stability, physicochemical properties, chemical reactivity and their relationship with anti-inflammatory activity; employing quantum-chemical descriptors at the M06-2X/6-311+G(d,p) level. Our results suggest that asparagine plays a more critical role than aspartic acid in the structural stability, physicochemical features, and chemical reactivity of these tripeptides. Substituent groups in the side chain cause significant changes on the conformational stability and chemical reactivity, and consequently on their anti-inflammatory activity.

PH-Sensitive Nanogels Synthesised by Radiation-Induced Cross-Linking of Hydrogen-Bonded Interpolymer Complexes in Aqueous Solution

International Nuclear Information System (INIS)

Ulanski, P.; Kadłubowski, S.; Henke, A.; Olejnik, A.K.; Rokita, B.; Wach, R.; Rosiak, J.M.

2010-01-01

Nanogels, i.e., internally cross-linked hydrophilic polymeric particles of sub-micron sizes, gained much interest over the last years due to their possible application as components of advanced type of medicines, like drug carriers. It is expected that they can facilitate distribution and delivery of different types of biologically active substances (including proteins, peptides and oligonucleotides) in a controlled way within the human body. Nanogels and their bigger analogues – microgels, are mainly synthesised through free-radical cross-linking polymerization of monomers. This synthetic routine can be carried out in solution but more often emulsion techniques are preferred (mini- or microemulsion) due to easier size control and exclusion of the macrogelation process. Additionally, surfactant-free emulsion polymerization (SFEP) is the method of choice for the preparation of temperature-sensitive particles, mainly based on poly(N-isopropylacrylamide).Nanogels were also successfully prepared by intramolecular cross-linking of single macromolecules. More recently, covalent stabilization was utilized to obtain the self-assembled structures like micelles of amphiphilic block copolymers, held by relatively weak physical interactions. Due to low stability of these polymolecular systems against dilution or temperature changes, different chemistry-based strategies to turn them into permanent nanopaticles were proposed in the literature (e.g., independent stabilization of a core or a shell of the micelles)

PH-Sensitive Nanogels Synthesised by Radiation-Induced Cross-Linking of Hydrogen-Bonded Interpolymer Complexes in Aqueous Solution

Energy Technology Data Exchange (ETDEWEB)

Ulanski, P.; Kadłubowski, S.; Henke, A.; Olejnik, A. K.; Rokita, B.; Wach, R.; Rosiak, J.M., E-mail: slawekka@mitr.p.lodz.pl [Technical University of Lodz, Wroblewskiego 15, 93-590 Lodz (Poland)

2010-07-01

Nanogels, i.e., internally cross-linked hydrophilic polymeric particles of sub-micron sizes, gained much interest over the last years due to their possible application as components of advanced type of medicines, like drug carriers. It is expected that they can facilitate distribution and delivery of different types of biologically active substances (including proteins, peptides and oligonucleotides) in a controlled way within the human body. Nanogels and their bigger analogues – microgels, are mainly synthesised through free-radical cross-linking polymerization of monomers. This synthetic routine can be carried out in solution but more often emulsion techniques are preferred (mini- or microemulsion) due to easier size control and exclusion of the macrogelation process. Additionally, surfactant-free emulsion polymerization (SFEP) is the method of choice for the preparation of temperature-sensitive particles, mainly based on poly(N-isopropylacrylamide).Nanogels were also successfully prepared by intramolecular cross-linking of single macromolecules. More recently, covalent stabilization was utilized to obtain the self-assembled structures like micelles of amphiphilic block copolymers, held by relatively weak physical interactions. Due to low stability of these polymolecular systems against dilution or temperature changes, different chemistry-based strategies to turn them into permanent nanopaticles were proposed in the literature (e.g., independent stabilization of a core or a shell of the micelles)

MeV ion beam interaction with polymer films containing cross-linking agents

International Nuclear Information System (INIS)

Evelyn, A. L.

1999-01-01

Polymer films containing cross linking enhancers were irradiated with MeV alpha particles to determine the effects of MeV ion beam interaction on these materials. The contributed effects from the electronic and nuclear stopping powers were separated by irradiating stacked thin films of polyvinyl chloride (PVC), polystyrene (PS) and polyethersulfone (PES). This layered system allowed most of the effects of the electronic energy deposited to be experienced by the first layers and the last layers to receive most of the effects of the nuclear stopping power. RGA, Raman microprobe analysis, RBS and FTIR measured changes in the chemical structures of the irradiated films. The characterization resolved the effects of the stopping powers on the PVC, PS and PES and the results were compared with those from previously studied polymers that did not contain any cross linking agents

Glutaraldehyde cross-linking of amniotic membranes affects their nanofibrous structures and limbal epithelial cell culture characteristics.

Science.gov (United States)

Lai, Jui-Yang; Ma, David Hui-Kang

2013-01-01

Given that the cells can sense nanometer dimensions, the chemical cross-linking-mediated alteration in fibrillar structure of collagenous tissue scaffolds is critical to determining their cell culture performances. This article explores, for the first time, the effect of nanofibrous structure of glutaraldehyde (GTA) cross-linked amniotic membrane (AM) on limbal epithelial cell (LEC) cultivation. Results of ninhydrin assays demonstrated that the amount of new cross-links formed between the collagen chains is significantly increased with increasing the cross-linking time from 1 to 24 hours. By transmission electron microscopy, the AM treated with GTA for a longer duration exhibited a greater extent of molecular aggregation, thereby leading to a considerable increase in nanofiber diameter and resistance against collagenase degradation. In vitro biocompatibility studies showed that the samples cross-linked with GTA for 24 hours are not well-tolerated by the human corneal epithelial cell cultures. When the treatment duration is less than 6 hours, the biological tissues cross-linked with GTA for a longer time may cause slight reductions in 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt, and anti-inflammatory activities. Nevertheless, significant collagen molecular aggregation also enhances the stemness gene expression, indicating a high ability of these AM matrices to preserve the progenitors of LECs in vitro. It is concluded that GTA cross-linking of collagenous tissue materials may affect their nanofibrous structures and corneal epithelial stem cell culture characteristics. The AM treated with GTA for 6 hours holds promise for use as a niche for the expansion and transplantation of limbal epithelial progenitor cells.

MgADP-induced changes in the structure of myosin S1 near the ATPase-related thiol SH1 probed by cross-linking

International Nuclear Information System (INIS)

Rajasekharan, K.N.; Mayadevi, M.; Agarwal, R.; Burke, M.

1990-01-01

The structural consequence of MgADP binding at the vicinity of the ATPase-related thiol SH1 (Cys-707) have been examined by subjecting myosin subfragment 1, premodified at SH2 (Cys-697) with N-ethylmaleimide (NEM), to reaction with the bifunctional reagent p-phenylenedimaleimide (pPDM) in the presence and absence of MgADP. By monitoring the changes in the Ca 2+ -ATPase activity as a function of reaction time, it appears that the reagent rapidly modifies SH1 irrespective of whether MgADP is present or not. In the absence of nucleotide, only extremely low levels of cross-linking to the 50-kDa middle segment of S1 can be detected, while in the presence of MgADP substantial cross-linking to this segment is observed. A similar cross-link is also formed if MgADP is added subsequent to the reaction of the SH2-NEM-premodified S1 with pPDM in the absence of nucleotide. Isolation of the labeled tryptic peptide from the cross-linked adduct formed with [ 14 C]pPDM, and subsequent partial sequence analyses, indicates that the cross-link is made from SH1 to Cys-522. Moreover, it appears that this cross-link results in the trapping of MgADP in this S1 species. These data suggest that the binding of MgADP results in a change in the structure of S1 in the vicinity of the SH1 thiol relative to the 50-kDa domain which enables Cys-522 to adopt the appropriate configuration to enable it to be cross-linked to SH1 by pPDM

Development of a Nucleoprotein-Based Enzyme-Linked Immunosorbent Assay Using a Synthetic Peptide Antigen for Detection of Avian Metapneumovirus Antibodies in Turkey Sera

Science.gov (United States)

Alvarez, Rene; Njenga, M. Kariuki; Scott, Melissa; Seal, Bruce S.

2004-01-01

Avian metapneumoviruses (aMPV) cause an upper respiratory tract disease with low mortality but high morbidity, primarily in commercial turkeys, that can be exacerbated by secondary infections. There are three types of aMPV, of which type C is found only in the United States. The aMPV nucleoprotein (N) amino acid sequences of serotypes A, B, and C were aligned for comparative analysis. On the basis of the predicted antigenicity of consensus sequences, five aMPV-specific N peptides were synthesized for development of a peptide antigen enzyme-linked immunosorbent assay (aMPV N peptide-based ELISA) to detect aMPV-specific antibodies among turkeys. Sera from naturally and experimentally infected turkeys were used to demonstrate the presence of antibodies reactive to the chemically synthesized aMPV N peptides. Subsequently, aMPV N peptide 1, which had the sequence 10-DLSYKHAILKESQYTIKRDV-29, with variations at only three amino acids among aMPV serotypes, was evaluated as a universal aMPV ELISA antigen. Data obtained with the peptide-based ELISA correlated positively with total aMPV viral antigen-based ELISAs, and the peptide ELISA provided higher optical density readings. The results indicated that aMPV N peptide 1 can be used as a universal ELISA antigen to detect antibodies for all aMPV serotypes. PMID:15013970

Linking algal growth inhibition to chemical activity

DEFF Research Database (Denmark)

Schmidt, Stine N.; Mayer, Philipp

Unitless chemical activity, expressing the energetic level of a compound relative to its energetic level in pure liquid [0-1], has proven useful to quantify the effective exposure to hydrophobic organic compounds through both aerial and aqueous media. Several studies have linked toxicity to chemi......Unitless chemical activity, expressing the energetic level of a compound relative to its energetic level in pure liquid [0-1], has proven useful to quantify the effective exposure to hydrophobic organic compounds through both aerial and aqueous media. Several studies have linked toxicity...... to chemical activity, as opposed to e.g. the total concentration. Baseline toxicity (narcosis) for neutral hydrophobic organic compounds has been shown to initiate in the narrow chemical activity range of 0.01 to 0.1. This presentation focuses on linking algal growth inhibition to chemical activity......-polar liquids were applied to challenge the chemical activity range for baseline toxicity. For each compound, the effective activity (Ea50) was estimated as the ratio of the effective concentration (EC50) and water solubility. Of these ratios, 90% were within the expected chemical activity range of 0.01 to 0...

Study of the Conformational State of Non-Cross-Linked and Cross-Linked Poly(alkylmethyldiallylammonium chlorides) in Aqueous Solution by Fluorescence Probing

NARCIS (Netherlands)

Wang, Guang-Jia; Engberts, Jan B.F.N.

The aggregation behaviour of novel non-cross-linked and cross-linked poly(alkylmethyldiallylammonium chlorides) in aqueous solutions has been investigated by fluorescence spectroscopy using pyrene as a probe. These copolymers were found to exhibit similar aggregate properties as the corresponding

Highly conductive carbon nanotube buckypapers with improved doping stability via conjugational cross-linking.

Science.gov (United States)

Chen, I-Wen Peter; Liang, Richard; Zhao, Haibo; Wang, Ben; Zhang, Chuck

2011-12-02

Carbon nanotube (CNT) sheets or buckypapers have demonstrated promising electrical conductivity and mechanical performance. However, their electrical conductivity is still far below the requirements for engineering applications, such as using as a substitute for copper mesh, which is currently used in composite aircraft structures for lightning strike protection. In this study, different CNT buckypapers were stretched to increase their alignment, and then subjected to conjugational cross-linking via chemical functionalization. The conjugationally cross-linked buckypapers (CCL-BPs) demonstrated higher electrical conductivity of up to 6200 S cm( - 1), which is more than one order increase compared to the pristine buckypapers. The CCL-BPs also showed excellent doping stability in over 300 h in atmosphere and were resistant to degradation at elevated temperatures. The tensile strength of the stretched CCL-BPs reached 220 MPa, which is about three times that of pristine buckypapers. We attribute these property improvements to the effective and stable conjugational cross-links of CNTs, which can simultaneously improve the electrical conductivity, doping stability and mechanical properties. Specifically, the electrical conductivity increase resulted from improving the CNT alignment and inter-tube electron transport capability. The conjugational cross-links provide effective 3D conductive paths to increase the mobility of electrons among individual nanotubes. The stable covalent bonding also enhances the thermal stability and load transfer. The significant electrical and mechanical property improvement renders buckypaper a multifunctional material for various applications, such as conducting composites, battery electrodes, capacitors, etc.

Radiation cross-linked plastics: a versatile material solution for packaging, automotive, Electrotechnic and Electronics

International Nuclear Information System (INIS)

Rouif, Sophie

2004-01-01

Used since the beginning of the 1970s for the production of halogen-free and heat-resistant cables and wires, for conditioning polyethylene hot-water pipes or for the manufacture of heat shrinkable tubes and of tyres, radiation cross-linking is developing fastly today on the scale of plastic-moulded parts, and not only by the mean of EB, but also under gamma rays. Indeed, it improves considerably the performances of a great number of plastics among thermoplastics, elastomers and thermoplastic elastomers (TPE). Radiation cross-linking reinforces the dimensional stability of polymers in chemically aggressive and high-temperature conditions. Radiation cross-linked-based engineering plastics offers OEM and end users in many branches of industry both technical and economical advantages in comparison with high-performances plastics. They constitute a technical and economical compromise between engineering plastics that failed and high-performances plastic, often over-tailored and expensive. This modern industrial technology gives way to new applications and perspectives in various sectors (packaging, automotive, electrotechnic and electronics, including connectors, surface-mounted devices, integrated circuits, 3D-MID, etc.) that are described in the paper

Modified gum arabic cross-linked gelatin scaffold for biomedical applications

International Nuclear Information System (INIS)

Sarika, P.R.; Cinthya, Kuriakose; Jayakrishnan, A.; Anilkumar, P.R.; James, Nirmala Rachel

2014-01-01

The present work deals with development of modified gum arabic cross-linked gelatin scaffold for cell culture. A new biocompatible scaffold was developed by cross-linking gelatin (Gel) with gum arabic, a polysaccharide. Gum arabic was subjected to periodate oxidation to obtain gum arabic aldehyde (GAA). GAA was reacted with gelatin under appropriate pH to prepare the cross-linked hydrogel. Cross-linking occurred due to Schiff's base reaction between aldehyde groups of oxidized gum arabic and amino groups of gelatin. The scaffold prepared from the hydrogel was characterized by swelling properties, degree of cross-linking, in vitro degradation and scanning electron microscopy (SEM). Cytocompatibility evaluation using L-929 and HepG2 cells confirmed non-cytotoxic and non-adherent nature of the scaffold. These properties are essential for generating multicellular spheroids and hence the scaffold is proposed to be a suitable candidate for spheroid cell culture. - Highlights: • Gum arabic cross-linked gelatin scaffold was developed for tissue engineering. • Cross-linking was achieved by Schiff's base reaction. • The scaffold is non-cytotoxic and non adherent to fibroblast and hepatocytes. • The scaffolds are potential candidates for spheroid cell culture

Modified gum arabic cross-linked gelatin scaffold for biomedical applications

Energy Technology Data Exchange (ETDEWEB)

Sarika, P.R. [Department of Chemistry, Indian Institute of Space Science and Technology, Valiamala, Thiruvananthapuram, Kerala 695 547 (India); Cinthya, Kuriakose [Tissue Culture Laboratory, Biomedical Technology Wing, Sree Chitra Tirunal Institute for Medical Sciences and Technology, Poojappura, Thiruvananthapuram, Kerala 695 012 (India); Jayakrishnan, A. [Department of Biotechnology, Indian Institute of Technology Madras, Chennai 600 036 (India); Anilkumar, P.R., E-mail: anilkumarpr@sctimst.ac.in [Tissue Culture Laboratory, Biomedical Technology Wing, Sree Chitra Tirunal Institute for Medical Sciences and Technology, Poojappura, Thiruvananthapuram, Kerala 695 012 (India); James, Nirmala Rachel, E-mail: nirmala@iist.ac.in [Department of Chemistry, Indian Institute of Space Science and Technology, Valiamala, Thiruvananthapuram, Kerala 695 547 (India)

2014-10-01

The present work deals with development of modified gum arabic cross-linked gelatin scaffold for cell culture. A new biocompatible scaffold was developed by cross-linking gelatin (Gel) with gum arabic, a polysaccharide. Gum arabic was subjected to periodate oxidation to obtain gum arabic aldehyde (GAA). GAA was reacted with gelatin under appropriate pH to prepare the cross-linked hydrogel. Cross-linking occurred due to Schiff's base reaction between aldehyde groups of oxidized gum arabic and amino groups of gelatin. The scaffold prepared from the hydrogel was characterized by swelling properties, degree of cross-linking, in vitro degradation and scanning electron microscopy (SEM). Cytocompatibility evaluation using L-929 and HepG2 cells confirmed non-cytotoxic and non-adherent nature of the scaffold. These properties are essential for generating multicellular spheroids and hence the scaffold is proposed to be a suitable candidate for spheroid cell culture. - Highlights: • Gum arabic cross-linked gelatin scaffold was developed for tissue engineering. • Cross-linking was achieved by Schiff's base reaction. • The scaffold is non-cytotoxic and non adherent to fibroblast and hepatocytes. • The scaffolds are potential candidates for spheroid cell culture.

Highly stable pyridinium-functionalized cross-linked anion exchange membranes for all vanadium redox flow batteries

Science.gov (United States)

Zeng, L.; Zhao, T. S.; Wei, L.; Zeng, Y. K.; Zhang, Z. H.

2016-11-01

It has recently been demonstrated that the use of anion exchange membranes (AEMs) in vanadium redox flow batteries (VRFBs) can reduce the migration of vanadium ions through the membrane due to the Donnan exclusion effect among the positively charged functional groups and vanadium ions. However, AEMs are plagued by low chemical stability in harsh chemical environments. Here we propose and fabricate a pyridinium-functionalized cross-linked AEM for VRFBs. The pyridinium-functionalized bromomethylated poly (2,6-dimethyl-1,4-phenylene oxide) exhibits a superior chemical stability as a result of the strengthened internal cross-linking networks and the chemical inertness of the polymer backbone. Therefore, the membrane exhibits littler decay in a harsh environment for 20 days during the course of an ex situ immersion test. A cycling test also demonstrates that the VRFB assembled with the membrane enable to retain 80% of the initial discharge capacity over 537 cycles with a capacity decay rate of 0.037% cycle-1. Meanwhile, the membrane also shows a low vanadium permeability and a reasonably high conductivity in supporting electrolytes. Hence, all the measurements and performance tests reported in this work suggest that the membrane is a promising AEM for redox flow batteries to achieve excellent cycling stability and superior cell performance.

Cross-linking of wheat gluten using a water-soluble carbodiimide

NARCIS (Netherlands)

Tropini, V.; Lens, J.P.; Mulder, W.J.; Silvestre, F.

2000-01-01

Wheat gluten was cross-linked using water-soluble 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide HCl (EDC). To enhance cross-linking, N-hydroxysuccinimide (NHS) was added to the reaction mixture. The cross-linking efficiency was evaluated by the decrease in the amount of amino groups, the solubility

The property distance index PD predicts peptides that cross-react with IgE antibodies

Science.gov (United States)

Ivanciuc, Ovidiu; Midoro-Horiuti, Terumi; Schein, Catherine H.; Xie, Liping; Hillman, Gilbert R.; Goldblum, Randall M.; Braun, Werner

2009-01-01

Similarities in the sequence and structure of allergens can explain clinically observed cross-reactivities. Distinguishing sequences that bind IgE in patient sera can be used to identify potentially allergenic protein sequences and aid in the design of hypo-allergenic proteins. The property distance index PD, incorporated in our Structural Database of Allergenic Proteins (SDAP, http://fermi.utmb.edu/SDAP/), may identify potentially cross-reactive segments of proteins, based on their similarity to known IgE epitopes. We sought to obtain experimental validation of the PD index as a quantitative predictor of IgE cross-reactivity, by designing peptide variants with predetermined PD scores relative to three linear IgE epitopes of Jun a 1, the dominant allergen from mountain cedar pollen. For each of the three epitopes, 60 peptides were designed with increasing PD values (decreasing physicochemical similarity) to the starting sequence. The peptides synthesized on a derivatized cellulose membrane were probed with sera from patients who were allergic to Jun a 1, and the experimental data were interpreted with a PD classification method. Peptides with low PD values relative to a given epitope were more likely to bind IgE from the sera than were those with PD values larger than 6. Control sequences, with PD values between 18 and 20 to all the three epitopes, did not bind patient IgE, thus validating our procedure for identifying negative control peptides. The PD index is a statistically validated method to detect discrete regions of proteins that have a high probability of cross-reacting with IgE from allergic patients. PMID:18950868

Synthesis and Characterization of Poly(hydroxyethyl methacrylate-co-methacrylic acid) Cross Linked Polymeric Network for the Delivery of Analgesic Agent

International Nuclear Information System (INIS)

Rashid, H.; Ahmad, M.; Minhas, M. U.; Aamir, M. F.; Sohail, M.

2015-01-01

Objective of the study was to synthesize a chemically cross-linked poly(hydroxyethylmethacrylate-co-methacrylic acid) hydrogel (p(HEMA-co-MAA hydrogel)) for pH-responsive, controlled drug delivery of Flurbiprofen. pH-sensitive hydrogel drug delivery system was synthesized with the help of poly hydroxyethyl methacrylate (HEMA) and methacrylic acid (MAA) by using ethylene glycol dimethacrylate (EGDMA) as crosslinker. The monomers (HEMA and MAA) were successfully cross linked through free radical polymerization process, initiated by benzoyl peroxide in an aqueous medium. All formulations were loaded with Flurbiprofen as a model drug and evaluated at pH 1.2 and pH 7.4 to investigate pH-responsive nature of the system. Fourier transform infrared spectroscopy (FTIR) was performed to confirm the cross-linking of copolymer while thermo gravimetric analysis (TGA) and differential scanning calorimetry (DSC) were performed to evaluate the thermal stability of the system. Swelling studies and in-vitro release studies were carried out to evaluate pH-responsive nature of the hydrogels. FTIR confirmed that monomers were successfully cross-linked to form a copolymer. Hydrogel system showed less swelling at lower pH while at higher pH, it showed higher swelling, releasing drug in the same fashion. It was concluded that a stable hydrogel network was chemically cross-linked showing pH-responsive nature and thus, synthesized p(HEMA-co-MAA) hydrogels can be successfully employed as potential candidate for controlled drug delivery. (author)

Immobilization of enzymes on radiation-modified gelatine gel by using a chemical cross-linking agent

International Nuclear Information System (INIS)

Bachmann, S.; Gebicka, L.; Galant, S.

1981-01-01

Investigations into the effect of ionizing radiation on the gelatine gels have shown that water-insoluble gel can be formed under suitable irradiation conditions. To establish the optimal conditions for the processing of the insoluble gel, the yield of cross-linking has been determined for gelatine solutions and its gels irradiated with various doses in the absence and in the presence of oxygen. Glucose isomerase (GI) was used as a test enzyme for immobilization on the gelatine gel. This enzyme which catalyses the isomerization of glucose to fructose has been used on the commercial-scale production of high fructose syrups. The support matrix chosen for the enzyme immobilization has been obtained by irradiating 4% wt/vol. de-aerated gelatine gel at a dose of 1.5 x 10 4 kGy at 15 0 C. Actinoplanes missouriensis cells containing GI were mixed with gelatine gel particles and cross-linked with glutaraldehyde. It was found that the immobilized GI can be successfully applied in the continuous isomerization of glucose to fructose. (author)

Photoreactivities and thermal properties of psoralen cross-links

International Nuclear Information System (INIS)

Yeung, A.T.; Jones, B.K.; Chu, C.T.

1988-01-01

The authors have studied the photoreaction of 8-methoxypsoralen (8-MOP), 4,5',8-trimethylpsoralen (TMP), and 4'-(hydroxymethyl)-4,5',8-trimethylpsoralen (HMT) with a pair of 18-base-long oligonucleotides in which a 14-base region is complementary. Only one 5'TpA site, favored for both monoadduct and cross-link formation with psoralen, is present in this oligonucleotide pair. They have used this model system to demonstrate, for the first time, strand specificity in the photoreaction of psoralen with DNA. They found that the two types of cross-links which form at this site have large differences in thermal stabilities. In addition, the denaturation of each cross-links isomer duplex occurred in at least three stages, which can be visualized as three bands in thermal equilibrium under the conditions of a denaturing polyacrylamide gel. This novel observation suggests that there are several domains differing in thermal stability in a psoralen cross-link

Collagen cross-linking in thin corneas

Directory of Open Access Journals (Sweden)

Prema Padmanabhan

2013-01-01

Full Text Available Collagen cross-linking (CXL has become the standard of care for progressive keratoconus, after numerous clinical studies have established its efficacy and safety in suitably selected eyes. The standard protocol is applicable in eyes which have a minimum corneal thickness of 400 μm after epithelial debridement. This prerequisite was stipulated to protect the corneal endothelium and intraocular tissues from the deleterious effect of ultraviolet-A (UVA radiation. However, patients with keratoconus often present with corneal thickness of less than 400 μm and could have otherwise benefited from this procedure. A few modifications of the standard procedure have been suggested to benefit these patients without a compromise in safety. Transepithelial cross-linking, pachymetry-guided epithelial debridement before cross-linking, and the use of hypoosmolar riboflavin are some of the techniques that have been attempted. Although clinical data is limited at the present time, these techniques are worth considering in patients with thin corneas. Further studies are needed to scientifically establish their efficacy and safety.

Cross-linked polyelectrolyte multilayers for marine antifouling applications

NARCIS (Netherlands)

Zhu, X.; Janczewski, D.; Lee, S.S.C.; Teo, S.L-M.; Vancso, Gyula J.

2013-01-01

A polyionic multilayer film was fabricated by layer-by-layer (LbL) sequential deposition followed by cross-linking under mild conditions on a substrate surface to inhibit marine fouling. A novel polyanion, featuring methyl ester groups for an easy cross-linking was used as a generic solution for

In vivo oxidation in remelted highly cross-linked retrievals.

Science.gov (United States)

Currier, B H; Van Citters, D W; Currier, J H; Collier, J P

2010-10-20

Elimination of free radicals to prevent oxidation has played a major role in the development and product differentiation of the latest generation of highly cross-linked ultra-high molecular weight polyethylene bearing materials. In the current study, we (1) examined oxidation in a series of retrieved remelted highly cross-linked ultra-high molecular weight polyethylene bearings from a number of device manufacturers and (2) compared the retrieval results with findings for shelf-stored control specimens. The hypothesis was that radiation-cross-linked remelted ultra-high molecular weight polyethylene would maintain oxidative stability in vivo comparable with the stability during shelf storage and in published laboratory aging tests. Fifty remelted highly cross-linked ultra-high molecular weight polyethylene acetabular liners and nineteen remelted highly cross-linked ultra-high molecular weight polyethylene tibial inserts were received after retrieval from twenty-one surgeons from across the U.S. Thirty-two of the retrievals had been in vivo for two years or more. Each was measured for oxidation with use of Fourier transform infrared spectroscopy. A control series of remelted highly cross-linked ultra-high molecular weight polyethylene acetabular liners from three manufacturers was analyzed with electron paramagnetic resonance spectroscopy to measure free radical content and with Fourier transform infrared spectroscopy to measure oxidation initially and after eight to nine years of shelf storage in air. The never-implanted, shelf-aged controls had no measurable free-radical content initially or after eight to nine years of shelf storage. The never-implanted controls showed no increase in oxidation during shelf storage. Oxidation measurements showed measurable oxidation in 22% of the retrieved remelted highly cross-linked liners and inserts after an average of two years in vivo. Because never-implanted remelted highly cross-linked ultra-high molecular weight

Rapid detection of DNA-interstrand and DNA-protein cross-links in mammalian cells by gravity-flow alkaline elution

International Nuclear Information System (INIS)

Hincks, J.R.; Coulombe, R.A. Jr.

1989-01-01

Alkaline elution is a sensitive and commonly used technique to detect cellular DNA damage in the form of DNA strand breaks and DNA cross-links. Conventional alkaline elution procedures have extensive equipment requirements and are tedious to perform. Our laboratory recently presented a rapid, simplified, and sensitive modification of the alkaline elution technique to detect carcinogen-induced DNA strand breaks. In the present study, we have further modified this technique to enable the rapid characterization of chemically induced DNA-interstrand and DNA-protein associated cross-links in cultured epithelial cells. Cells were exposed to three known DNA cross-linking agents, nitrogen mustard (HN 2 ), mitomycin C (MMC), or ultraviolet irradiation (UV). One hour exposures of HN 2 at 0.25, 1.0, and 4.0 microM or of MMC at 20, 40, and 60 microM produced a dose-dependent induction of total DNA cross-links by these agents. Digestion with proteinase K revealed that HN 2 and MMC induced both DNA-protein cross-links and DNA-interstrand cross-links. Ultraviolet irradiation induced both DNA cross-links and DNA strand breaks, the latter of which were either protein and nonprotein associated. The results demonstrate that gravity-flow alkaline elution is a sensitive and accurate method to characterize the molecular events of DNA cross-linking. Using this procedure, elution of DNA from treated cells is completed in 1 hr, and only three fractions per sample are analyzed. This method may be useful as a rapid screening assay for genotoxicity and/or as an adjunct to other predictive assays for potential mutagenic or carcinogenic agents

Heating tubes of cross-linked polyethylene

International Nuclear Information System (INIS)

Knoeppler, H.; Hoffmann, M.

1981-01-01

Oxygen permeability of plastic tubes for floor heating systems was measured as a function of the reduced oxygen content of water in plastic tubes at a flow rate of 0.5 m/s and a temperature of 30 0 C and as a function of oxygen uptake of low-oxygen water in floor heating tubes. Pipes of VEP, periodically cross-linked polyethylene (Engels process), polypropylene copolymeride, and polybutene were compared. The permeability of periodically cross-linked polyethylene is twice as high as that of VEP. Measurements, results, and consequences for floor heating systems are discussed. (KH) [de

Synthesis and Catalytic Properties of Non-Cross-Linked and Cross-Linked Poly(alkylmethyldiallylammonium bromides) Having Decyl, Octyl, and Hexyl Side Chains

NARCIS (Netherlands)

Wang, G.J; Engberts, J.B.F.N.

1995-01-01

A family of non-cross-linked and cross-linked copolymers containing decyl, octyl, and hexyl groups as side chains ((CL)-CopolC1-10, (CL)-CopolC1-8, and (CL)-CopolC1-6, respectively) were synthesized by radical-initiated cyclocopolymerization of alkylmethyldiallylammonium bromide monomers without and

Multi-stimulus-responsive shape-memory polymer nanocomposite network cross-linked by cellulose nanocrystals.

Science.gov (United States)

Liu, Ye; Li, Ying; Yang, Guang; Zheng, Xiaotong; Zhou, Shaobing

2015-02-25

In this study, we developed a thermoresponsive and water-responsive shape-memory polymer nanocomposite network by chemically cross-linking cellulose nanocrystals (CNCs) with polycaprolactone (PCL) and polyethylene glycol (PEG). The nanocomposite network was fully characterized, including the microstructure, cross-link density, water contact angle, water uptake, crystallinity, thermal properties, and static and dynamic mechanical properties. We found that the PEG[60]-PCL[40]-CNC[10] nanocomposite exhibited excellent thermo-induced and water-induced shape-memory effects in water at 37 °C (close to body temperature), and the introduction of CNC clearly improved the mechanical properties of the mixture of both PEG and PCL polymers with low molecular weights. In addition, Alamar blue assays based on osteoblasts indicated that the nanocomposites possessed good cytocompatibility. Therefore, this thermoresponsive and water-responsive shape-memory nanocomposite could be potentially developed into a new smart biomaterial.

Effects of Genipin Concentration on Cross-Linked Chitosan Scaffolds for Bone Tissue Engineering: Structural Characterization and Evidence of Biocompatibility Features

Directory of Open Access Journals (Sweden)

Simona Dimida

2017-01-01

Full Text Available Genipin (GN is a natural molecule extracted from the fruit of Gardenia jasminoides Ellis according to modern microbiological processes. Genipin is considered as a favorable cross-linking agent due to its low cytotoxicity compared to widely used cross-linkers; it cross-links compounds with primary amine groups such as proteins, collagen, and chitosan. Chitosan is a biocompatible polymer that is currently studied in bone tissue engineering for its capacity to promote growth and mineral-rich matrix deposition by osteoblasts in culture. In this work, two genipin cross-linked chitosan scaffolds for bone repair and regeneration were prepared with different GN concentrations, and their chemical, physical, and biological properties were explored. Scanning electron microscopy and mechanical tests revealed that nonremarkable changes in morphology, porosity, and mechanical strength of scaffolds are induced by increasing the cross-linking degree. Also, the degradation rate was shown to decrease while increasing the cross-linking degree, with the high cross-linking density of the scaffold disabling the hydrolysis activity. Finally, basic biocompatibility was investigated in vitro, by evaluating proliferation of two human-derived cell lines, namely, the MG63 (human immortalized osteosarcoma and the hMSCs (human mesenchymal stem cells, as suitable cell models for bone tissue engineering applications of biomaterials.

Hotspot autoimmune T cell receptor binding underlies pathogen and insulin peptide cross-reactivity

Science.gov (United States)

Cole, David K.; Bulek, Anna M.; Dolton, Garry; Schauenberg, Andrea J.; Szomolay, Barbara; Trimby, Andrew; Jothikumar, Prithiviraj; Fuller, Anna; Skowera, Ania; Rossjohn, Jamie; Zhu, Cheng; Miles, John J.; Wooldridge, Linda; Rizkallah, Pierre J.; Sewell, Andrew K.

2016-01-01

The cross-reactivity of T cells with pathogen- and self-derived peptides has been implicated as a pathway involved in the development of autoimmunity. However, the mechanisms that allow the clonal T cell antigen receptor (TCR) to functionally engage multiple peptide–major histocompatibility complexes (pMHC) are unclear. Here, we studied multiligand discrimination by a human, preproinsulin reactive, MHC class-I–restricted CD8+ T cell clone (1E6) that can recognize over 1 million different peptides. We generated high-resolution structures of the 1E6 TCR bound to 7 altered peptide ligands, including a pathogen-derived peptide that was an order of magnitude more potent than the natural self-peptide. Evaluation of these structures demonstrated that binding was stabilized through a conserved lock-and-key–like minimal binding footprint that enables 1E6 TCR to tolerate vast numbers of substitutions outside of this so-called hotspot. Highly potent antigens of the 1E6 TCR engaged with a strong antipathogen-like binding affinity; this engagement was governed though an energetic switch from an enthalpically to entropically driven interaction compared with the natural autoimmune ligand. Together, these data highlight how T cell cross-reactivity with pathogen-derived antigens might break self-tolerance to induce autoimmune disease. PMID:27183389

Practical application of thermoreversibly Cross-linked rubber products

Science.gov (United States)

Polgar, L. M.; Picchioni, F.; de Ruiter, E.; van Duin, M.

2017-07-01

Currently, rubber products cannot simply be reprocessed after their product life, due to the irreversible cross-linking methods traditionally applied. The purpose of this work is to investigate how thermoreversible cross-linking of rubbers via Diels Alder chemistry can be used for the development of recyclable rubber products. Unfortunately, the applicability of the thermoreversible EPM-g-furan/BM system appears to be limited to room temperature applications, because of the rapid deterioration of the compression set at elevated temperatures compared to irreversibly cross-linked EPM. However, the use of EPM rubber modified with thiophene or cyclopentadiene moieties may extend the temperature application range and results in rubber products with acceptable properties. Finally, rubber products generally comprise fillers such as silica, carbon black or fibers. In this context, the reinforcing effect of short cut aramid fibers on the material properties of the newly developed thermoreversibly cross-linked EPM rubbers was also studied. The material properties of the resulting products were found to be comparable to those of a fiber reinforced, peroxide cured reference sample.

Identification of cross-linked amino acids in the protein pair HmaL23-HmaL29 from the 50S ribosomal subunit of the archaebacterium Haloarcula marismortui.

Science.gov (United States)

Bergmann, U; Wittmann-Liebold, B

1993-03-23

50S ribosomal subunits from the extreme halophilic archaebacterium Haloarcula marismortui were treated with the homobifunctional protein-protein cross-linking reagents diepoxybutane (4 A) and dithiobis(succinimidyl propionate) (12 A). The dominant product with both cross-linking reagents was identified on the protein level as HmaL23-HmaL29, which is homologous to the protein pair L23-L29 from Escherichia coli [Walleczek, J., Martin, T., Redl, B., Stöffler-Meilicke, M., & Stöffler, G. (1989) Biochemistry 28, 4099-4105] and from Bacillus stearothermophilus [Brockmöller, J., & Kamp, R. M. (1986) Biol. Chem. Hoppe-Seyler 367, 925-935]. To reveal the exact cross-linking site in HmaL23-HmaL29, the cross-linked complex was purified on a preparative scale by conventional and high-performance liquid chromatography. After endoproteolytic fragmentation of the protein pair, the amino acids engaged in cross-link formation were unambiguously identified by N-terminal sequence analysis and mass spectrometry of the cross-linked peptides. The cross-link is formed between lysine-57 in the C-terminal region of HmaL29 and the alpha-amino group of the N-terminal serine in protein HmaL23, irrespective of the cross-linking reagent. This result demonstrates that the N-terminal region of protein HmaL23 and the C-terminal domain of HmaL29 are highly flexible so that the distance between the two polypeptide chains can vary by at least 8 A. Comparison of our cross-linking results with those obtained with B. stearothermophilus revealed that the fine structure within this ribosomal domain is at least partially conserved.

Site-directed cross-linking: establishing the dimeric structure of the aspartate receptor of bacterial chemotaxis

International Nuclear Information System (INIS)

Milligan, D.L.; Koshland, D.E. Jr.

1988-01-01

Cysteine residues introduced at specific locations in the aspartate receptor of Salmonella typhimurium provide anchor points for cross-linking and serve as chemical markers for structural studies of this oligomeric receptor. These markers have been used to measure the rate of subunit exchange between oligomeric receptors and to show that ligand binding inhibits this exchange. The cysteine-containing receptors can be oxidatively cross-linked to completion within the oligomeric receptor, indicating that the receptor has an even number of subunits. Based on this observation, a technique has been developed that can be used to determine the oligomeric structure of proteins under a variety of experimental conditions. The technique involves the measurement of the effect of dilution by cysteineless receptor subunits on cross-linking and reveals that the aspartate receptor is dimeric in detergent solution, in a mixed-micelle system, and in reconstituted membrane vesicles. Binding of aspartate does not change the oligomeric structure of the receptor, indicating that transmembrane signaling occurs within an oligomeric receptor of constant size

Sub- T g Cross-Linking of a Polyimide Membrane for Enhanced CO 2 Plasticization Resistance for Natural Gas Separation

KAUST Repository

Qiu, Wulin

2011-08-09

Decarboxylation-induced thermal cross-linking occurs at elevated temperatures (∼15 °C above glass transition temperature) for 6FDA-DAM:DABA polyimides, which can stabilize membranes against swelling and plasticization in aggressive feed streams. Despite this advantage, such a high temperature might result in collapse of substructure and transition layers in the asymmetric structure of a hollow fibers based on such a material. In this work, the thermal cross-linking of the 6FDA-DAM:DABA at temperatures much below the glass transition temperature (∼387 °C by DSC) was demonstrated. This sub-Tg cross-linking capability enables extension to asymmetric structures useful for large scale membranes. The resulting polymer membranes were characterized by swelling in known solvents for the un-cross-linked materials, TGA analysis, and permeation tests of aggressive gas feed stream at higher pressure. The annealing temperature and time clearly influence the degree of cross-linking of the membranes, and results in a slight difference in selectivity for membranes under various cross-linking conditions. Results indicate that the sub-Tg thermal cross-linking of 6FDA-DAM:DABA dense film membrane can be carried out completely even at a temperature as low as 330 °C. Permeabilities were tested for the polyimide membranes using both pure gases (He, O2, N2, CH4, CO2) and mixed gases (CO2/CH4). The selectivity of the cross-linked membrane can be maintained even under very aggressive CO2 operating conditions that are not possible without cross-linking. Moreover, the plasticization resistance was demonstrated up to 700 psia for pure CO 2 gas or 1000 psia for 50% CO2 mixed gas feeds. © 2011 American Chemical Society.

UV cross-linking of polypeptides associated with 3'-terminal exons

International Nuclear Information System (INIS)

Stolow, D.T.; Berget, S.M.

1990-01-01

Association of nuclear proteins with chimeric vertebrate precursor RNAs containing both polyadenylation signals and an intron was examined by UV cross-linking. One major difference in cross-linking pattern was observed between this chimeric precursor RNA and precursors containing only polyadenylation or splicing signals. The heterogeneous nuclear ribonucleoprotein (hnRNP) polypeptide C cross-linked strongly to sequences downstream of the A addition site in polyadenylation precursor RNA containing only the polyadenylation signal from the simian virus 40 (SV40) late transcription unit. In contrast, the hnRNP C polypeptide cross-linked to chimeric RNA containing the same SV40 late poly(A) cassette very poorly, at a level less than 5% of that observed with the precursor RNA containing just the poly(A) site. Observation that cross-linking of the hnRNP C polypeptide to elements within the SV40 late poly(A) site was altered by the presence of an upstream intron suggests differences in the way nuclear factors associate with poly(A) sites in the presence and absence of an upstream intron. Cross-linking of C polypeptide to chimeric RNA increased with RNAs mutated for splicing or polyadenylation consensus sequences and under reaction conditions (high magnesium) that inhibited polyadenylation. Furthermore, cross-linking of hnRNP C polypeptide to precursors containing just the SV40 late poly(A) site was eliminated in the presence of competing poly(U); polyadenylation, however, was unaffected. Correlation of loss of activity with high levels of hnRNP C polypeptide cross-linking raises questions about the specificity of the interaction between the hnRNP C polypeptide and polyadenylation precursor RNAs in vitro

Tailoring the properties of cholecyst-derived extracellular matrix using carbodiimide cross-linking.

LENUS (Irish Health Repository)

Burugapalli, Krishna

2009-01-01

Modulation of properties of extracellular matrix (ECM) based scaffolds is key for their application in the clinical setting. In the present study, cross-linking was used as a tool for tailoring the properties of cholecyst-derived extracellular matrix (CEM). CEM was cross-linked with varying cross-linking concentrations of N,N-(3-dimethyl aminopropyl)-N\\'-ethyl carbodiimide (EDC) in the presence of N-hydroxysuccinimide (NHS). Shrink temperature measurements and ATR-FT-IR spectra were used to determine the degree of cross-linking. The effect of cross-linking on degradation was tested using the collagenase assay. Uniaxial tensile properties and the ability to support fibroblasts were also evaluated as a function of cross-linking. Shrink temperature increased from 59 degrees C for non-cross-linked CEM to 78 degrees C for the highest EDC cross-linking concentration, while IR peak area ratios for the free -NH(2) group at 3290 cm(-1) to that of the amide I band at 1635 cm(-1) decreased with increasing EDC cross-linking concentration. Collagenase assay demonstrated that degradation rates for CEM can be tailored. EDC concentrations 0 to 0.0033 mmol\\/mg CEM were the cross-linking concentration range in which CEM showed varied susceptibility to collagenase degradation. Furthermore, cross-linking concentrations up to 0.1 mmol EDC\\/mg CEM did not have statistically significant effect on the uniaxial tensile strength, as well as morphology, viability and proliferation of fibroblasts on CEM. In conclusion, the degradation rates of CEM can be tailored using EDC-cross-linking, while maintaining the mechanical properties and the ability of CEM to support cells.

The spectra character of photodegraded the pyridinoline cross-links by Hypocrellin B

International Nuclear Information System (INIS)

Zhang Jucheng; Chen Rui; Liu Wei; Chen Zhuo; Shu Lidan; Liu Yingji

2011-01-01

Pyridinoline cross-links is one of the cross-link formation in collagen which in cell matrix, many research shown that this cross-link cause the fibrosis. Hypocrellin B (HB) is one of the nature photosensitizers, this work investigated the pyridinoline cross-link in collagen was photodegraded by HB. The result shown HB can degrade the pyridinoline cross-link with photo. This is to say, HB may be use as the photodynamic reagent to study the fibrosis.

Interactions of cross-linked and uncross-linked chitosan hydrogels ...

African Journals Online (AJOL)

The swelling equilibrium of Chitosan and sodium tripolyphosphate (NaTPP) cross-linked chitosan hydrogels in aqueous solutions of surfactants differing in structure and hydrophobicity at 250C is reported. Anionic surfactant sodium dodecylsulfate (SDS), the cationic surfactant hexadecyltrimethylammonium bromide (HTAB) ...

Cross-linking of dermal sheep collagen using a water-soluble carbodiimide

NARCIS (Netherlands)

Damink, LHHO; Dijkstra, PJ; vanLuyn, MJA; vanWachem, PB; Nieuwenhuis, P; Feijen, J

A cross-linking method for collagen-based biomaterials was developed using the water-soluble carbodiimide 1-ethyl-3-(3-dimethyl aminopropyl)carbodiimide hydrochloride (EDC). Cross-linking using EDC involves the activation of carboxylic acid groups to give O-acylisourea groups, which form cross-links

Cross-linking of dermal sheep collagen using a water-soluble carbodiimide

NARCIS (Netherlands)

Olde damink, L.H.H.; Olde Damink, L.H.H.; Dijkstra, Pieter J.; van Luyn, M.J.A.; van Wachem, P.B.; Nieuwenhuis, P.; Feijen, Jan

1996-01-01

A cross-linking method for collagen-based biomaterials was developed using the water-soluble carbodiimide 1-ethyl-3-(3-dimethyl aminopropyl)carbodiimide hydrochloride (EDC). Cross-linking using EDC involves the activation of carboxylic acid groups to give O-acylisourea groups, which form cross-links

Formaldehyde cross-linking and structural proteomics: Bridging the gap.

Science.gov (United States)

Srinivasa, Savita; Ding, Xuan; Kast, Juergen

2015-11-01

Proteins are dynamic entities constantly moving and altering their structures based on their functions and interactions inside and outside the cell. Formaldehyde cross-linking combined with mass spectrometry can accurately capture interactions of these rapidly changing biomolecules while maintaining their physiological surroundings. Even with its numerous established uses in biology and compatibility with mass spectrometry, formaldehyde has not yet been applied in structural proteomics. However, formaldehyde cross-linking is moving toward analyzing tertiary structure, which conventional cross-linkers have already accomplished. The purpose of this review is to describe the potential of formaldehyde cross-linking in structural proteomics by highlighting its applications, characteristics and current status in the field. Copyright © 2015 Elsevier Inc. All rights reserved.

Small Strain Topological Effects of Biopolymer Networks with Rigid Cross-Links

NARCIS (Netherlands)

Zagar, G.; Onck, P. R.; Van der Giessen, E.; Garikipati, K; Arruda, EM

2010-01-01

Networks of cross-linked filamentous biopolymers form topological structures characterized by L, T and X cross-link types of connectivity 2, 3 and 4, respectively. The distribution of cross-links over these three types proofs to be very important for the initial elastic shear stiffness of isotropic

Haptoglobin preferentially binds β but not α subunits cross-linked hemoglobin tetramers with minimal effects on ligand and redox reactions.

Science.gov (United States)

Jia, Yiping; Wood, Francine; Buehler, Paul W; Alayash, Abdu I

2013-01-01

Human hemoglobin (Hb) and haptoglobin (Hp) exhibit an extremely high affinity for each other, and the dissociation of Hb tetramers into dimers is generally believed to be a prerequisite for complex formation. We have investigated Hp interactions with native Hb, αα, and ββ cross-linked Hb (ααXLHb and ββXLHb, respectively), and rapid kinetics of Hb ligand binding as well as the redox reactivity in the presence of and absence of Hp. The quaternary conformation of ββ subunit cross-linking results in a higher binding affinity than that of αα subunit cross-linked Hb. However, ββ cross-linked Hb exhibits a four fold slower association rate constant than the reaction rate of unmodified Hb with Hp. The Hp contact regions in the Hb dimer interfaces appear to be more readily exposed in ββXLHb than ααXLHb. In addition, apart from the functional changes caused by chemical modifications, Hp binding does not induce appreciable effects on the ligand binding and redox reactions of ββXLHb. Our findings may therefore be relevant to the design of safer Hb-based oxygen therapeutics by utilizing this preferential binding of ββXLHb to Hp. This may ultimately provide a safe oxidative inactivation and clearance pathway for chemically modified Hbs in circulation.

Synthesis of a cyclic fibrin-like peptide and its analysis by fast atom bombardment mass spectrometry

International Nuclear Information System (INIS)

Young, J.D.; Costello, C.E.; Langenhove, A. van; Haber, E.; Matsueda, G.R.

1983-01-01

For immunochemical purposes, a cyclic 12 peptide was synthesized to model the γ-γ-chain cross-link site in human fibrin. The model was based upon the structure proposed by Chen and Doolittle which is characterized by two reciprocating epsilon-(γ-Glu)Lys bonds between adjacent fibrin γ-chains oriented in an antiparallel manner. To achieve the antiparallel orientation of the peptide backbone, Pro and Gly were inserted at positions 6 and 7 of the linear 12-peptide: acetyl-Gly-Glu-Gln-His-His-Pro-Gly-Gly-Gly-Ala-Lys-Gly-amide. The insertions were made to facilitate a reverse turn of the peptide during the last synthetic step, which was formation of the epsilon-(γ-Glu)Lys bond between Glu at position 2 and Lys at position 11 with diphenylphosphorylazide. The resulting cyclic peptide represented half of the symmetrical cross-linked region in clotted fibrin. Following purification by HPLC, both linear and cyclic 12-peptides were analyzed by fast atom bombardment mass spectrometry. Abundant molecular protonated ions were observed for both peptides. In addition, the amino acid sequence of the linear peptide and the location of the epsilon-(γ-Glu)Lys bond in the cyclized peptide could be verified. (author)

Cross-linked polymeric membranes for carbon dioxide separation

Science.gov (United States)

Hong, Tao; Chatterjee, Sabornie; Mahurin, Shannon Mark; Long, Brian Keith; Jiang, De-en; Mays, Jimmy Wayne; Sokolov, Alexei P.; Saito, Tomonori

2018-01-23

A membrane useful in gas separation, the membrane comprising a cross-linked polysiloxane structure having a cross-link density of about 0.1.times.10.sup.-5 mol/cm.sup.3 to about 6.times.10.sup.-5 mol/cm.sup.3, where, in particular embodiments, the cross-linked polysiloxane structure has the following general structure: ##STR00001## wherein R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5, and R.sup.6 are independently selected from hydrocarbon groups having at least 1 and up to 6 carbon atoms; A.sup.1 and A.sup.2 are independently selected from cyclic hydrocarbon groups; L.sup.1 and L.sup.2 are linking groups or covalent bonds; n is an integer of at least 1; r and s are independently selected from integers of at least 1; and p is an integer of at least 10. The invention also includes methods for making and using the above-described membranes for gas separation.

Glutaraldehyde cross-linking of amniotic membranes affects their nanofibrous structures and limbal epithelial cell culture characteristics

Directory of Open Access Journals (Sweden)

Lai JY

2013-10-01

Full Text Available Jui-Yang Lai,1–3 David Hui-Kang Ma4,5 1Institute of Biochemical and Biomedical Engineering, 2Biomedical Engineering Research Center, 3Molecular Medicine Research Center, Chang Gung University, Taoyuan, Taiwan; 4Limbal Stem Cell Laboratory, Department of Ophthalmology, Chang Gung Memorial Hospital, Taoyuan, Taiwan; 5Department of Chinese Medicine, Chang Gung University, Taoyuan, Taiwan Abstract: Given that the cells can sense nanometer dimensions, the chemical cross-linking-mediated alteration in fibrillar structure of collagenous tissue scaffolds is critical to determining their cell culture performances. This article explores, for the first time, the effect of nanofibrous structure of glutaraldehyde (GTA cross-linked amniotic membrane (AM on limbal epithelial cell (LEC cultivation. Results of ninhydrin assays demonstrated that the amount of new cross-links formed between the collagen chains is significantly increased with increasing the cross-linking time from 1 to 24 hours. By transmission electron microscopy, the AM treated with GTA for a longer duration exhibited a greater extent of molecular aggregation, thereby leading to a considerable increase in nanofiber diameter and resistance against collagenase degradation. In vitro biocompatibility studies showed that the samples cross-linked with GTA for 24 hours are not well-tolerated by the human corneal epithelial cell cultures. When the treatment duration is less than 6 hours, the biological tissues cross-linked with GTA for a longer time may cause slight reductions in 3-(4,5-dimethylthiazol-2-yl-5-(3-carboxymethoxyphenyl-2-(4-sulfophenyl-2H-tetrazolium, inner salt, and anti-inflammatory activities. Nevertheless, significant collagen molecular aggregation also enhances the stemness gene expression, indicating a high ability of these AM matrices to preserve the progenitors of LECs in vitro. It is concluded that GTA cross-linking of collagenous tissue materials may affect their nanofibrous

Collagen cross linking: Current perspectives

Directory of Open Access Journals (Sweden)

Srinivas K Rao

2013-01-01

Full Text Available Keratoconus is a common ectatic disorder occurring in more than 1 in 1,000 individuals. The condition typically starts in adolescence and early adulthood. It is a disease with an uncertain cause and its progression is unpredictable, but in extreme cases, vision deteriorates and can require corneal transplant surgery. Corneal collagen cross-linking (CCL with riboflavin (C3R is a recent treatment option that can enhance the rigidity of the cornea and prevent disease progression. Since its inception, the procedure has evolved with newer instrumentation, surgical techniques, and is also now performed for expanded indications other than keratoconus. With increasing experience, newer guidelines regarding optimization of patient selection, the spectrum of complications and their management, and combination procedures are being described. This article in conjunction with the others in this issue, will try and explore the uses of collagen cross-linking (CXL in its current form.

Development of new cross-linked polyethylene for atomic energy

International Nuclear Information System (INIS)

Fujimura, Shun-ichi; Ohya, Shingo; Kubo, Masaji; Tsutsumi, Yukihiro; Seguchi, Tadao.

1988-01-01

Cross-linked polyethylene is the material which is used most as the insulating material for electric wires and cables, but for the cables for nuclear power stations and the wiring materials within machinery and equipment, the cross-linked polyethylene which is hard to burn by mixing burning-retarding agent is frequently used as the disaster-preventing countermeasures. As the burning-retarding agent for cross-linked polyethylene, bromine system agent that gives high burning retardation, chlorine system agent that can prevent melting and dripping at the time of burning and so on have been used so as to meet the objective. However by the addition of burning-retarding agents, the electrical and mechanical characteristics of cross-linked polyethylene lower, therefore consideration must be given to the use. In this paper, the results of the examination on the application of condensed acenaphthylene bromide as a new burning-retarding agent to cross-linked polyethylene are reported. White lead was effective for catching HBr. It was confirmed that more than 30 parts of this agent ensured burning retardation. By mixing this agent, the tensile strength increased, but the elongation lowered. It was found that the good radiation resistance was obtained by adding this agent. (K.I.)

Arginine-rich intracellular delivery peptides noncovalently transport protein into living cells

International Nuclear Information System (INIS)

Wang, Y.-H.; Chen, C.-P.; Chan, M.-H.; Chang, M.; Hou, Y.-W.; Chen, H.-H.; Hsu, H.-R.; Liu, Kevin; Lee, H.-J.

2006-01-01

Plasma membranes of plant or animal cells are generally impermeable to peptides or proteins. Many basic peptides have previously been investigated and covalently cross-linked with cargoes for cellular internalization. In the current study, we demonstrate that arginine-rich intracellular delivery (AID) peptides are able to deliver fluorescent proteins or β-galactosidase enzyme into animal and plant cells, as well as animal tissue. Cellular internalization and transdermal delivery of protein could be mediated by effective and nontoxic AID peptides in a neither fusion protein nor conjugation fashion. Therefore, noncovalent AID peptides may provide a useful strategy to have active proteins function in living cells and tissues in vivo

Protein analysis by 31p NMR spectroscopy in ionic liquid: quantitative determination of enzymatically created cross-links.

Science.gov (United States)

Monogioudi, Evanthia; Permi, Perttu; Filpponen, Ilari; Lienemann, Michael; Li, Bin; Argyropoulos, Dimitris; Buchert, Johanna; Mattinen, Maija-Liisa

2011-02-23

Cross-linking of β-casein by Trichoderma reesei tyrosinase (TrTyr) and Streptoverticillium mobaraense transglutaminase (Tgase) was analyzed by (31)P nuclear magnetic resonance (NMR) spectroscopy in ionic liquid (IL). According to (31)P NMR, 91% of the tyrosine side chains were cross-linked by TrTyr at high dosages. When Tgase was used, no changes were observed because a different cross-linking mechanism was operational. However, this verified the success of the phosphitylation of phenolics within the protein matrix in the IL. Atomic force microscopy (AFM) in solid state showed that disk-shaped nanoparticles were formed in the reactions with average diameters of 80 and 20 nm for TrTyr and Tgase, respectively. These data further advance the current understanding of the action of tyrosinases on proteins on molecular and chemical bond levels. Quantitative (31)P NMR in IL was shown to be a simple and efficient method for the study of protein modification.

Expression of the cationic antimicrobial peptide lactoferricin fused with the anionic peptide in Escherichia coli.

Science.gov (United States)

Kim, Ha-Kun; Chun, Dae-Sik; Kim, Joon-Sik; Yun, Cheol-Ho; Lee, Ju-Hoon; Hong, Soon-Kwang; Kang, Dae-Kyung

2006-09-01

Direct expression of lactoferricin, an antimicrobial peptide, is lethal to Escherichia coli. For the efficient production of lactoferricin in E. coli, we developed an expression system in which the gene for the lysine- and arginine-rich cationic lactoferricin was fused to an anionic peptide gene to neutralize the basic property of lactoferricin, and successfully overexpressed the concatemeric fusion gene in E. coli. The lactoferricin gene was linked to a modified magainin intervening sequence gene by a recombinational polymerase chain reaction, thus producing an acidic peptide-lactoferricin fusion gene. The monomeric acidic peptide-lactoferricin fusion gene was multimerized and expressed in E. coli BL21(DE3) upon induction with isopropyl-beta-D-thiogalactopyranoside. The expression levels of the fusion peptide reached the maximum at the tetramer, while further increases in the copy number of the fusion gene substantially reduced the peptide expression level. The fusion peptides were isolated and cleaved to generate the separate lactoferricin and acidic peptide. About 60 mg of pure recombinant lactoferricin was obtained from 1 L of E. coli culture. The purified recombinant lactoferricin was found to have a molecular weight similar to that of chemically synthesized lactoferricin. The recombinant lactoferricin showed antimicrobial activity and disrupted bacterial membrane permeability, as the native lactoferricin peptide does.

Studies on Cross-linking of succinic acid with chitosan/collagen

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Tapas Mitra

2013-01-01

Full Text Available The present study summarizes the cross-linking property of succinic acid with chitosan /collagen. In detail, the chemistry behind the cross-linking and the improvement in mechanical and thermal properties of the cross-linked material were discussed with suitable instruments and bioinformatics tools. The concentration of succinic acid with reference to the chosen polymers was optimized. A 3D scaffold prepared using an optimized concentration of succinic acid (0.2% (w/v with chitosan (1.0% (w/v and similarly with collagen (0.5% (w/v, was subjected to surface morphology, FT-IR analysis, tensile strength assessment, thermal stability and biocompatibility. Results revealed, cross-linking with succinic acid impart appreciable mechanical strength to the scaffold material. In silico analysis suggested the prevalence of non-covalent interactions, which played a crucial role in improving the mechanical and thermal properties of the cross-linked scaffold. The resultant 3D scaffold may find application as wound dressing material, as an implant in clinical applications and as a tissue engineering material.

X-ray-mediated cross linking of protein and DNA

International Nuclear Information System (INIS)

Minsky, B.D.; Braun, A.

1977-01-01

Using a simple filter assay for the binding of BSA or lysozyme to DNA, two mechanisms of x-ray-mediated cross linking are shown to occur. One, a fast reaction, appears to involve a radical intermediate, is inhibited by high pH and salt, and seems to be enhanced by deoxygenation. The second mechanism, a slow time-dependent component, differs from the fast reaction in its stimulation by histidine, its inhibition by catalase, and the lack of an oxygen effect. Separate irradiation of DNA or water does not lead to cross linking. However, separate irradiation of protein leads to cross linking which proceeds with slow-component kinetics

Diagnosis of Zika Virus Infection by Peptide Array and Enzyme-Linked Immunosorbent Assay

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Nischay Mishra

2018-03-01

Full Text Available Zika virus (ZIKV is implicated in fetal stillbirth, microcephaly, intracranial calcifications, and ocular anomalies following vertical transmission from infected mothers. In adults, infection may trigger autoimmune inflammatory polyneuropathy. Transmission most commonly follows the bite of infected Aedes mosquitoes but may also occur through sexual intercourse or receipt of blood products. Definitive diagnosis through detection of viral RNA is possible in serum or plasma within 10 days of disease onset, in whole blood within 3 weeks of onset, and in semen for up to 3 months. Serological diagnosis is nonetheless critical because few patients have access to molecular diagnostics during the acute phase of infection and infection may be associated with only mild or inapparent disease that does not prompt molecular testing. Serological diagnosis is confounded by cross-reactivity of immune sera with other flaviviruses endemic in the areas where ZIKV has recently emerged. Accordingly, we built a high-density microarray comprising nonredundant 12-mer peptides that tile, with one-residue overlap, the proteomes of Zika, dengue, yellow fever, West Nile, Ilheus, Oropouche, and chikungunya viruses. Serological analysis enabled discovery of a ZIKV NS2B 20-residue peptide that had high sensitivity (96.0% and specificity (95.9% versus natural infection with or vaccination against dengue, chikungunya, yellow fever, West Nile, tick-borne encephalitis, or Japanese encephalitis virus in a microarray assay and an enzyme-linked immunosorbent assay (ELISA of early-convalescent-phase sera (2 to 3 weeks after onset of symptomatic infection.

Applicability of a particularly simple model to nonlinear elasticity of slide-ring gels with movable cross-links as revealed by unequal biaxial deformation.

Science.gov (United States)

Kondo, Yuuki; Urayama, Kenji; Kidowaki, Masatoshi; Mayumi, Koichi; Takigawa, Toshikazu; Ito, Kohzo

2014-10-07

The strain energy density function (F) of the polyrotaxane-based slide-ring (SR) gels with movable cross-links along the network strands is characterized by unequal biaxial stretching which can achieve various types of deformation. The SR gels as prepared without any post-preparation complication exhibit considerably smaller values of the ratio of the stresses (σy/σx) in the stretched (x) and constrained (y) directions in planar extension than classical chemical gels with heterogeneous and nearly homogeneous network structures do. This feature of the SR gels leads to the peculiar characteristic that the strain energy density function (F) has no explicit cross term of strains in different directions, which is in contrast to F with explicit strain cross terms for most chemical gels and elastomers. The biaxial stress-strain data of the SR gels are successfully described by F of the Gent model with only two parameters (small-strain shear modulus and a parameter representing ultimate elongation), which introduces the finite extensibility effect into the neo-Hookean model with no explicit cross term of strain. The biaxial data of the deswollen SR gels examined in previous study, which underwent a considerable reduction in volume from the preparation state, are also well described by the Gent model, which is in contrast to the case of the classical chemical gels that the stress-strain relations before and after large deswelling are not described by a common type of F due to a significant degree of collapse of the network strands in the deswollen state. These intriguing features of nonlinear elasticity of the SR gels originate from a novel function of the slidable cross-links that can maximize the arrangement entropy of cross-linked and non-cross-linked cyclic molecules in the deformed networks.

Seed-mediated synthesis of cross-linked Pt-NiO nanochains for methanol oxidation

Energy Technology Data Exchange (ETDEWEB)

Gu, Zhulan; Bin, Duan; Feng, Yue; Zhang, Ke; Wang, Jin; Yan, Bo; Li, Shumin; Xiong, Zhiping; Wang, Caiqin; Shiraishi, Yukihide; Du, Yukou, E-mail: duyk@suda.edu.cn

2017-07-31

Highlights: • Cross-linked Pt-NiO nanochains using seed-mediated growth method are synthesized. • The as-prepared catalysts exhibit higher electrocatalytic activity than Pt/C for MOR. • The Pt-NiO(1:1 by molar) catalyst shows the best electrocatalytic property towards MOR. - Abstract: A simple method was reported for employing NiO nanoparticles act as seeds and then different amounts of Pt{sup 2+} were reduced on the NiO nanoparticles, forming a cross-linked Pt-NiO nanocatalysts. These as-prepared catalysts were characterized using different physical-chemical techniques, including X-ray diffraction (XRD), transmission electron microscopy (TEM) and X-ray photoelectron spectroscopy (XPS). The results indicate that the morphology of the cross-linked Pt-NiO nanochain was successfully produced regardless of the molar ratio of Pt{sup 2+} to NiO precursors. The electrochemical characteristics of Pt-NiO nanochain catalysts were evaluated for the oxidation of methanol as a model reaction, which verify that the Pt-NiO catalysts show enhanced activity and high stability in comparison with the commercial Pt/C catalyst. The optimized ratio of Pt to NiO is 1:1, then tuned by simple adjusting the feed ratio of the precursors as well. The synthesized nanocatalysts will be found the great potential applications as electrocatalysts for fuel cells owe to their enhanced catalytic performance and long-term stability.

Seed-mediated synthesis of cross-linked Pt-NiO nanochains for methanol oxidation

International Nuclear Information System (INIS)

Gu, Zhulan; Bin, Duan; Feng, Yue; Zhang, Ke; Wang, Jin; Yan, Bo; Li, Shumin; Xiong, Zhiping; Wang, Caiqin; Shiraishi, Yukihide; Du, Yukou

2017-01-01

Highlights: • Cross-linked Pt-NiO nanochains using seed-mediated growth method are synthesized. • The as-prepared catalysts exhibit higher electrocatalytic activity than Pt/C for MOR. • The Pt-NiO(1:1 by molar) catalyst shows the best electrocatalytic property towards MOR. - Abstract: A simple method was reported for employing NiO nanoparticles act as seeds and then different amounts of Pt 2+ were reduced on the NiO nanoparticles, forming a cross-linked Pt-NiO nanocatalysts. These as-prepared catalysts were characterized using different physical-chemical techniques, including X-ray diffraction (XRD), transmission electron microscopy (TEM) and X-ray photoelectron spectroscopy (XPS). The results indicate that the morphology of the cross-linked Pt-NiO nanochain was successfully produced regardless of the molar ratio of Pt 2+ to NiO precursors. The electrochemical characteristics of Pt-NiO nanochain catalysts were evaluated for the oxidation of methanol as a model reaction, which verify that the Pt-NiO catalysts show enhanced activity and high stability in comparison with the commercial Pt/C catalyst. The optimized ratio of Pt to NiO is 1:1, then tuned by simple adjusting the feed ratio of the precursors as well. The synthesized nanocatalysts will be found the great potential applications as electrocatalysts for fuel cells owe to their enhanced catalytic performance and long-term stability.

Stabilized Sulfonated Aromatic Polymers by in situ Solvothermal Cross-Linking

Energy Technology Data Exchange (ETDEWEB)

Di Vona, Maria Luisa, E-mail: divona@uniroma2.it; Sgreccia, Emanuela [Dipartimento di Scienze e Tecnologie Chimiche, Università di Roma Tor Vergata, Rome (Italy); Narducci, Riccardo; Pasquini, Luca [Dipartimento di Scienze e Tecnologie Chimiche, Università di Roma Tor Vergata, Rome (Italy); MAtériaux Divisés, Interfaces, Réactivité, ELectrochimie (MADIREL – UMR 7246), Aix Marseille Université, Marseille (France); Hou, Hongying [Faculty of Material and Engineering, Kunming University of Science and Technology, Kunming (China); Knauth, Philippe [MAtériaux Divisés, Interfaces, Réactivité, ELectrochimie (MADIREL – UMR 7246), Aix Marseille Université, Marseille (France)

2014-10-10

The cross-link reaction via sulfone bridges of sulfonated polyether ether ketone (SPEEK) by thermal treatment at 180°C in presence of dimethylsulfoxide is discussed. The modifications of properties subsequent to the cross-linking are presented. The mechanical strength as well as the hydrolytic stability increased with the thermal treatment time, i.e., with the degree of cross-linking. The proton conductivity was determined as function of temperature, IEC, degree of cross-linking, and hydration number. The memory effect, which is the membrane ability to “remember” the water uptake reached at high temperature also at lower temperature, is exploited in order to achieve high values of conductivity. Membranes swelled at 110°C can reach a conductivity of 0.14 S/cm at 80°C with a hydration number (λ) of 73.

Hydroxyl radical induced cross-linking of cytosine and tyrosine in nucleohistone

International Nuclear Information System (INIS)

Gajewski, E.; Dizdaroglu, M.

1990-01-01

Hydroxyl radical induced formation of a DNA-protein cross-link involving cytosine and tyrosine in nucleohistone in buffered aqueous solution is reported. The technique of gas chromatography-mass spectrometry was used for this investigation. A γ-irradiated aqueous mixture of cytosine and tyrosine was first investigated in order to obtain gas chromatographic-mass spectrometric properties of possible cytosine-tyrosine cross-links. One cross-link was observed, and its structure was identified as the product from the formation of a covalent bond between carbon 6 of cytosine and carbon 3 of tyrosine. With the use of gas chromatography-mass spectrometry with selected-ion monitoring, this cytosine-tyrosine cross-link was identified in acidic hydrolysates of calf thymus nucleohistone γ-irradiated in N 2 O-saturated aqueous solution. The yield of this DNA-protein cross-link in nucleohistone was found to be a linear function of the radiation dose in the range of 100-500 Gy (J·kg -1 ). This yield amounted to 0.05 nmol·J -1 . Mechanisms underlying the formation of the cytosine-tyrosine cross-link in nucleohistone were proposed to involve radical-radical and/or radical addition reactions of hydroxyl adduct radicals of cytosine and tyrosine moieties, forming a covalent bond between carbon 6 of cytosine and carbon 3 of tyrosine. When oxygen was present in irradiated solutions, no cytosine-tyrosine cross-links were observed

Development of extremely low wear cross-link polyethylene for 30 years

International Nuclear Information System (INIS)

Oonishi, Hironobu; Fujita, Hiroshi; Kim, Seok-Cheol; Ito, Shigeru; Masuda, Shingo; Clarke, I.C.

2003-01-01

In this report we present our long-term developmental and clinical results with both highly cross-linked and extensively cross-linked polyethylene materials. Beginning in 1970s, we performed wear screening studies on ultra high molecular weight polyethylene (UHMWPE) (GUR412) sterilized by gamma-irradiation in air (range 0 to 10,000 kGy). From these scientific studies the 1,000 kGy dose (100 Mrad) appeared optimal, and so we began clinical use in 1971, and that continued into 1978. The radiographic wear-rates in patients with 1,000 kGy sockets, assessed by radiography, appeared 6-fold reduced compared to our standard UHMWPE sockets. Note also that we had not used any post-sterilization heat treatment for these pioneering extensively cross-linked polyethylene sockets. With clinical use now over 30 years, it was also clear that there was no adverse oxidation created by any free radicals present in our extensively cross-linked polyethylene sockets. With these encouraging clinical results, we further studied laboratory wear results with the modern UHMWPE resins, using the irradiation doses 1,000, 5,000, 10,000 and 15,000 kGy and with both saline and serum lubricants in hip simulators. These more recent studies demonstrated that the wear in extensively cross-linked polyethylene sockets was undetectable, less even than the measurement errors in the simulator techniques. It was unfortunate that the physical properties of such extensively cross-linked polyethylene sockets did not meet the current International Organization for Standardization (ISO) and American Society for Testing and Materials (ASTM) standards. Thus, despite the excellent wear performance of these materials, we decided to investigate also the properties of the 60 kGy irradiated UHMWPE. The polyethylene sheet (GUR1050) was first irradiated with 35 kGy under N2 and then heat treated to remove free radicals. The socket liners were then machined to shape and resterilized with 25 kGy under N2 gas. The

Reinforcement of Bacillus subtilis spores by cross-linking of outer coat proteins during maturation.

Science.gov (United States)

Abhyankar, Wishwas; Pandey, Rachna; Ter Beek, Alexander; Brul, Stanley; de Koning, Leo J; de Koster, Chris G

2015-02-01

Resistance characteristics of bacterial endospores towards various environmental stresses such as chemicals and heat are in part attributed to their coat proteins. Heat resistance is developed in a late stage of sporulation and during maturation of released spores. Using our gel-free proteomic approach and LC-FT-ICR-MS/MS analysis we have monitored the efficiency of the tryptic digestion of proteins in the coat during spore maturation over a period of eight days, using metabolically (15)N labeled mature spores as reference. The results showed that during spore maturation the loss of digestion efficiency of outer coat and crust proteins synchronized with the increase in heat resistance. This implicates that spore maturation involves chemical cross-linking of outer coat and crust layer proteins leaving the inner coat layer proteins unmodified. It appears that digestion efficiencies of spore surface proteins can be linked to their location within the coat and crust layers. We also attempted to study a possible link between spore maturation and the observed heterogeneity in spore germination. Copyright © 2014 Elsevier Ltd. All rights reserved.

UV induced DNA-protein cross links in vitro and in vivo

International Nuclear Information System (INIS)

Kornhauser, A.

1976-01-01

The review was not intended to cover all the past year's literature in this field; only selective material published in 1974 and 1975 has been surveyed. Covalent linkage of DNA and RNA to proteins induced by UV is considered, but DNA-membrade attachment, amino acids covalently bound to DNA as functions of growth conditions and protein non-covalently bound to DNA involved in cell regulation are excluded. Studies of DNA-protein cross-links upon UV irradiation in chemical model systems, bacteria and tissue culture systems, and an in vivo mammalian system are all surveyed. (U.K.)

Stabilized sulfonated aromatic polymers by in situ solvothermal cross-linking

Directory of Open Access Journals (Sweden)

Maria Luisa eDi Vona

2014-10-01

Full Text Available The cross-link reaction via sulfone bridges of sulfonated polyetheretherketone (SPEEK by thermal treatment at 180 °C in presence of dimethylsulfoxide (DMSO is discussed. The modifications of properties subsequent to the cross-linking are presented. The mechanical strength as well as the hydrolytic stability increased with the thermal treatment time, i.e., with the degree of cross-linking. The proton conductivity was determined as function of temperature, IEC, degree of cross-linking and hydration number. The memory effect, which is the membrane ability to remember the water uptake reached at high temperature also at lower temperature, is exploited in order to achieve high values of conductivity. Membranes swelled at 110 °C can reach a conductivity of 0.14 S/cm at 80°C with a hydration number ( of 73.

Evaluation of nanohydroxyapaptite (nano-HA) coated epigallocatechin-3-gallate (EGCG) cross-linked collagen membranes.

Science.gov (United States)

Chu, Chenyu; Deng, Jia; Man, Yi; Qu, Yili

2017-09-01

Collagen is the main component of extracellular matrix (ECM) with desirable biological activities and low antigenicity. Collagen materials have been widely utilized in guided bone regeneration (GBR) surgery due to its abilities to maintain space for hard tissue growth. However, pure collagen lacks optimal mechanical properties. In our previous study, epigallocatechin-3-gallate (EGCG) cross-linked collagen membranes, with better biological activities and enhanced mechanical properties, may promote osteoblast proliferation, but their effect on osteoblast differentiation is not very significant. Nanohydroxyapatite (nano-HA) is the main component of mineral bone, which possesses exceptional bioactivity properties including good biocompatibility, high osteoconductivity and osteoinductivity, non-immunogenicity and non-inflammatory behavior. Herein, by analyzing the physical and chemical properties as well as the effects on promoting bone regeneration, we have attempted to present a novel EGCG-modified collagen membrane with nano-HA coating, and have found evidence that the novel collagen membrane may promote bone regeneration with a better surface morphology, without destroying collagen backbone. To evaluate the surface morphologies, chemical and mechanical properties of pure collagen membranes, epigallocatechin-3-gallate (EGCG) cross-linked collagen membranes, nano-HA coated collagen membranes, nano-HA coated EGCG-collagen membranes, (ii) to evaluate the bone regeneration promoted by theses membranes. In the present study, collagen membranes were divided into 4 groups: (1) untreated collagen membranes (2) EGCG cross-linked collagen membranes (3) nano-HA modified collagen membranes (4) nano-HA modified EGCG-collagen membranes. Scanning electron microscope (SEM) and Fourier transform infrared spectroscopy (FTIR) were used to evaluate surface morphologies and chemical properties, respectively. Mechanical properties were determined by differential scanning calorimeter (DSC

A fossil protein chimera; difficulties in discriminating dinosaur peptide sequences from modern cross-contamination

Science.gov (United States)

Warwood, Stacey; van Dongen, Bart; Kitchener, Andrew C.; Manning, Phillip L.

2017-01-01

A decade ago, reports that organic-rich soft tissue survived from dinosaur fossils were apparently supported by proteomics-derived sequence information of exceptionally well-preserved bone. This initial claim to the sequencing of endogenous collagen peptides from an approximately 68 Myr Tyrannosaurus rex fossil was highly controversial, largely on the grounds of potential contamination from either bacterial biofilms or from laboratory practice. In a subsequent study, collagen peptide sequences from an approximately 78 Myr Brachylophosaurus canadensis fossil were reported that have remained largely unchallenged. However, the endogeneity of these sequences relies heavily on a single peptide sequence, apparently unique to both dinosaurs. Given the potential for cross-contamination from modern bone analysed by the same team, here we extract collagen from bone samples of three individuals of ostrich, Struthio camelus. The resulting LC–MS/MS data were found to match all of the proposed sequences for both the original Tyrannosaurus and Brachylophosaurus studies. Regardless of the true nature of the dinosaur peptides, our finding highlights the difficulty of differentiating such sequences with confidence. Our results not only imply that cross-contamination cannot be ruled out, but that appropriate measures to test for endogeneity should be further evaluated. PMID:28566488

Characterization of solid UV cross-linked PEGDA for biological applications

KAUST Repository

Castro, David

2013-10-20

This paper reports on solid UV cross-linked Poly(ethylene)-glycol-diacrylate (PEGDA) as a material for microfluidic devices for biological applications. We have evaluated biocompatibility of PEGDA through two separate means: 1) by examining cell viability and attachment on cross-linked PEGDA surfaces for cell culture applications, and 2) by determining if cross-linked PEGDA inhibits the polymerase chain reaction (PCR) processes for on-chip PCR. Through these studies a correlation has been found between degree of curing and cell viability, attachment, as well as on PCR outcome.

Characterization of Chemical and Physical Properties of Hydroxypropylated and Cross-linked Arrowroot (Marantha arundinacea Starch

Directory of Open Access Journals (Sweden)

Rijanti Rahaju Maulani

2013-12-01

Full Text Available The modern food industry and a variety of food products require tolerant starch as raw material for processing in a broad range of techniques, from preparation to storage and distribution. Dual modification of arrowroot starch using hydroxypropylation and cross-linking was carried out to overcome the lack of native arrowroot starch in food processing application. The modifications applied were: combined propylene oxide (8%, 10%, and 12%; sodium tri meta phosphate/STMP (1%, 2%, and 3%; and sodium tri poly phosphate/STPP (4%, 5%, and 6%. These modifications significantly affected the composition of the amylose and amylopectin and the amount of phosphorus in the granules. Higher amounts of phosphate salt gave a higher phosphorus content, which increased the degree of substitution (DS and the degree of cross-link. Arrowroot starch that was modified using a concentration of 8-10% propylene oxide and 1-2% STMP : 3-5% STPP produced a starch with < 0.4% phosphorus content. A higher concentration of propylene oxide provided a higher degree of hydroxypropyl. The changed physical properties of the modified granular arrowroot starch were examined through SEM testing, and its changed crystalline patterns through X-ray diffraction measurements. Especially, provision of a high concentration of propylene oxide (12% combined with 3% STMP : 6% STPP affected the granular morphology and the crystallinity.

Protein and Peptide Gas-phase Structure Investigation Using Collision Cross Section Measurements and Hydrogen Deuterium Exchange

Science.gov (United States)

Khakinejad, Mahdiar

Protein and peptide gas-phase structure analysis provides the opportunity to study these species outside of their explicit environment where the interaction network with surrounding molecules makes the analysis difficult [1]. Although gas-phase structure analysis offers a unique opportunity to study the intrinsic behavior of these biomolecules [2-4], proteins and peptides exhibit very low vapor pressures [2]. Peptide and protein ions can be rendered in the gas-phase using electrospray ionization (ESI) [5]. There is a growing body of literature that shows proteins and peptides can maintain solution structures during the process of ESI and these structures can persist for a few hundred milliseconds [6-9]. Techniques for monitoring gas-phase protein and peptide ion structures are categorized as physical probes and chemical probes. Collision cross section (CCS) measurement, being a physical probe, is a powerful method to investigate gas-phase structure size [3, 7, 10-15]; however, CCS values alone do not establish a one to one relation with structure(i.e., the CCS value is an orientationally averaged value [15-18]. Here we propose the utility of gas-phase hydrogen deuterium exchange (HDX) as a second criterion of structure elucidation. The proposed approach incudes extensive MD simulations to sample biomolecular ion conformation space with the production of numerous, random in-silico structures. Subsequently a CCS can be calculated for these structures and theoretical CCS values are compared with experimental values to produce a pool of candidate structures. Utilizing a chemical reaction model based on the gas-phase HDX mechanism, the HDX kinetics behavior of these candidate structures are predicted and compared to experimental results to nominate the best in-silico structures which match (chemically and physically) with experimental observations. For the predictive approach to succeed, an extensive technique and method development is essential. To combine CCS

Cross-linking of sodium caseinate-structured emulsion with transglutaminase alters postprandial metabolic and appetite responses in healthy young individuals.

Science.gov (United States)

Juvonen, Kristiina R; Macierzanka, Adam; Lille, Martina E; Laaksonen, David E; Mykkänen, Hannu M; Niskanen, Leo K; Pihlajamäki, Jussi; Mäkelä, Kari A; Mills, Clare E N; Mackie, Alan R; Malcolm, Paul; Herzig, Karl-Heinz; Poutanen, Kaisa S; Karhunen, Leila J

2015-08-14

The physico-chemical and interfacial properties of fat emulsions influence lipid digestion and may affect postprandial responses. The aim of the present study was to determine the effects of the modification of the interfacial layer of a fat emulsion by cross-linking on postprandial metabolic and appetite responses. A total of fifteen healthy individuals (26.5 (sem 6.9) years and BMI 21.9 (sem 2.0) kg/m2) participated in a cross-over design experiment in which they consumed two isoenergetic (1924 kJ (460 kcal)) and isovolumic (250 g) emulsions stabilised with either sodium caseinate (Cas) or transglutaminase-cross-linked sodium caseinate (Cas-TG) in a randomised order. Blood samples were collected from the individuals at baseline and for 6 h postprandially for the determination of serum TAG and plasma NEFA, cholecystokinin (CCK), glucagon-like peptide 1 (GLP-1), glucose and insulin responses. Appetite was assessed using visual analogue scales. Postprandial TAG and NEFA responses and gastric emptying (GE) rates were comparable between the emulsions. CCK increased more after the ingestion of Cas-TG than after the ingestion of Cas (P< 0.05), while GLP-1 responses did not differ between the two test emulsions. Glucose and insulin profiles were lower after consuming Cas-TG than after consuming Cas (P< 0.05). The overall insulin, glucose and CCK responses, expressed as areas above/under the curve, did not differ significantly between the Cas and Cas-TG meal conditions. Satiety ratings were reduced and hunger, desire to eat and thirst ratings increased more after the ingestion of Cas-TG than after the ingestion of Cas (P< 0.05). The present results suggest that even a subtle structural modification of the interfacial layer of a fat emulsion can alter the early postprandial profiles of glucose, insulin, CCK, appetite and satiety through decreased protein digestion without affecting significantly on GE or overall lipid digestion.

Vitamin C and Poly(ethylene glycol) Protect Concentrated Poly(vinyl alcohol) Solutions against Radiation Cross-linking

International Nuclear Information System (INIS)

Oral, E.

2006-01-01

There is a need for an injectable material to augment damaged cartilage. We propose to make such self-associating poly(vinyl alcohol) (PVA) hydrogels. Physical associations can be formed in PVA using a gellant such as polyethylene glycol (PEG). The injectability of PVA solutions is compromised when sterilized due to chemical cross-linking. We hypothesized that an anticross-linking agent could prevent cross-linking of irradiated PVA solutions. PVA (17.5 wt/v %, MW= 115,000 g/mol) was prepared in water at 90 degree. PEG (MW=400 g/mol) was added at a ratio of PEG unit to PVA unit of 17, 86, 290, and 639 mol/mol. PVA solutions (17.5 wt/v %, MW= 16,000, 61,000, 81,000 and 115,000 g/mol) were also prepared. Vitamin C was added at a molar ratio of vitamin C to PVA unit of 0.75-10.4. Solutions were poured into syringes and γ-irradiated. The viscosity of injectable solutions was determined by using the bubble tube. Gel content of cross-linked samples was measured by boiling gels in water for 6 hours, drying at 90 degree and calculating the ratio of dry weight to 'as is' weight

Recyclable cross-linked anion exchange membrane for alkaline fuel cell application

Science.gov (United States)

Hou, Jianqiu; Liu, Yazhi; Ge, Qianqian; Yang, Zhengjin; Wu, Liang; Xu, Tongwen

2018-01-01

Cross-linking can effectively solve the conductivity-swelling dilemma in anion exchange membranes (AEMs) but will generate solid wastes. To address this, we developed an AEM cross-linked via disulfide bonds, bearing quaternary ammonium groups, which can be easily recycled. The membrane (RC-QPPO) with IEC of 1.78 mmol g-1, when cross-linked, showed enhanced mechanical properties and good hydroxide conductivity (24.6 mS cm-1 at 30 °C). Even at higher IEC value (2.13 mmol g-1), it still has low water uptake, low swelling ratio and delivers a peak power density of 150 mW cm-2 at 65 °C. Exploiting the formation of disulfide bonds from -SH groups, the membrane can be readily cross-linked in alkaline condition and recycled by reversibly breaking disulfide bonds with dithiothreitol (DTT). The recycled membrane solution can be directly utilized to cast a brand-new AEM. By washing away the residual DTT with water and exposure to air, it can be cross-linked again and this process is repeatable. During the recycling and cross-linking processes, the membrane showed a slight IEC decrease of 5% due to functional group degradation. The strategy presented here is promising in enhancing AEM properties and reducing the impact of artificial polymers on the environment.

Yield and Failure Behavior Investigated for Cross-Linked Phenolic Resins Using Molecular Dynamics

Science.gov (United States)

Monk, Joshua D.; Lawson, John W.

2016-01-01

Molecular dynamics simulations were conducted to fundamentally evaluate the yield and failure behavior of cross-linked phenolic resins at temperatures below the glass transition. Yield stress was investigated at various temperatures, strain rates, and degrees of cross-linking. The onset of non-linear behavior in the cross-linked phenolic structures was caused by localized irreversible molecular rearrangements through the rotation of methylene linkers followed by the formation or annihilation of neighboring hydrogen bonds. The yield stress results, with respect to temperature and strain rate, could be fit by existing models used to describe yield behavior of amorphous glasses. The degree of cross-linking only indirectly influences the maximum yield stress through its influence on glass transition temperature (Tg), however there is a strong relationship between the degree of cross-linking and the failure mechanism. Low cross-linked samples were able to separate through void formation, whereas the highly cross-linked structures exhibited bond scission.

Combination of supramolecular cross-linking with covalent cross-linking through epoxide ring-opening including gel studies

NARCIS (Netherlands)

Hofmeier, H.; El-Ghayoury, A.; Schubert, U.S.

2003-01-01

Terpolymers based on poly(methyl methacrylate), containing terpyridinemoieties as well as epoxide groups, were synthesized via free-radical polymerization. The products were cross-linked non-covalently with iron(II) ions and covalently by treatment with AlCl3. Both steps could be combined in

Electric long-term behaviour of polyethylene insulations for medium-voltage cables cross-linked chemically or by radiation

International Nuclear Information System (INIS)

Scharff, C.; Eberhardt, M.

1989-01-01

The electric long-term behaviour of cross-linked polyethylene (CLPE) was studied at room temperature by measuring the channel operating time under needling and the total charge before channel operation. It is found that the decomposition products formed in CLPE act as tension stabilizers. They have a dominating influence on the electric long-term behaviour

Phosphorylation of actin-binding protein (ABP-280; filamin) by tyrosine kinase p56lck modulates actin filament cross-linking.

Science.gov (United States)

Pal Sharma, C; Goldmann, Wolfgang H

2004-01-01

Actin-binding protein (ABP-280; filamin) is a phosphoprotein present in the periphery of the cytoplasm where it can cross-link actin filaments, associate with lipid membranes, and bind to membrane surface receptors. Given its function and localization in the cell, we decided to investigate the possibility of whether it serves as substrate for p56lck, a lymphocyte-specific member of the src family of protein tyrosine kinases associated with cell surface glycoproteins. The interaction of p56lck with membrane glycoproteins is important for cell development and functional activation. Here, we show that purified p56lck interacts and catalyzes in vitro kinase reactions. Tyrosine phosphorylation by p56lck is restricted to a single peptide of labeled ABP-280 shown by protease digest. The addition of phorbol ester to cells results in the inhibition of phosphorylation of ABP-280 by p56lck. These results show a decrease in phosphorylation suggesting conformationally induced regulation. Dynamic light scattering confirmed increased actin filament cross-linking due to phosphorylation of ABP-280 by p56lck.

UV-induced cross-linking of abscisic acid to binding proteins

International Nuclear Information System (INIS)

Cornelussen, M.H.M.; Karssen, C.M.; Loon, L.C. van

1995-01-01

Conditions for UV-induced cross-linking of abscisic acid (ABA) through its enone chromophore to binding proteins were evaluated. The effects of a UV-light band between 260 and 530 nm on both unconjugated and protein-conjugated ABA, as well as on anti-ABA antibodies as models of ABA-binding proteins were determined. UV irradiation caused both isomerization and photolysis of ABA, but increasing the lower irradiation boundary to 345 nm strongly reduced photolysis and largely prevented isomerization. When conjugated to alkaline phosphatase (AP), ABA remained stable when using either a 320 or a 345 nm filter. At these wavelengths both binding of ABA to antibodies as well as AP enzymatic activity were maintained. UV-induced cross-linking of monoclonal anti-ABA antibodies to immobilized ABA was analysed by immunoassays. Optimal cross-linking was achieved after a 5 min irradiation period at 0°, using a long pass, cut-on filter to quench wavelengths below 290 nm. This cross-linking faithfully reflected cognate binding activity. (author)

Combination of supramolecular cross-linking with covalent cross-linking through epoxide ring-opening including gel studies

NARCIS (Netherlands)

Hofmeier, H.; El-Ghayoury, A.; Schubert, U.S.

2003-01-01

Terpolymers based on poly(methyl methacrylate), containing terpyridine-moieties as well as epoxide groups, were synthesized via free-radical polymeri-zation. The products were cross-linked non-covalently with iron(II) ions and cova-lently by treatment with AlCl3. Both steps could be combined in

Cross-Linked Poly-4-vinylpyridines as Useful Supports in Metal Catalysis: Micro- and Nanometer Scale Morphology.

Czech Academy of Sciences Publication Activity Database

D'Archivio, A.A.; Tauro, L.; Galantini, L.; Panatta, A.; Tettamanti, E.; Giammatteo, M.; Jeřábek, Karel; Corain, B.

2007-01-01

Roč. 268, 1-2 (2007) , s. 176-184 ISSN 1381-1169 R&D Projects: GA AV ČR(CZ) KSK4050111 Grant - others:MURS(IT) 2001038991 Institutional research plan: CEZ:AV0Z40720504 Keywords : cross-linked functional polymers * poly-4-vinylpyridines * supported Pt(0) nanoclusters Subject RIV: CI - Industrial Chemistry, Chemical Engineering Impact factor: 2.707, year: 2007

A fossil protein chimera; difficulties in discriminating dinosaur peptide sequences from modern cross-contamination.

Science.gov (United States)

Buckley, Michael; Warwood, Stacey; van Dongen, Bart; Kitchener, Andrew C; Manning, Phillip L

2017-05-31

A decade ago, reports that organic-rich soft tissue survived from dinosaur fossils were apparently supported by proteomics-derived sequence information of exceptionally well-preserved bone. This initial claim to the sequencing of endogenous collagen peptides from an approximately 68 Myr Tyrannosaurus rex fossil was highly controversial, largely on the grounds of potential contamination from either bacterial biofilms or from laboratory practice. In a subsequent study, collagen peptide sequences from an approximately 78 Myr Brachylophosaurus canadensis fossil were reported that have remained largely unchallenged. However, the endogeneity of these sequences relies heavily on a single peptide sequence, apparently unique to both dinosaurs. Given the potential for cross-contamination from modern bone analysed by the same team, here we extract collagen from bone samples of three individuals of ostrich, Struthio camelus The resulting LC-MS/MS data were found to match all of the proposed sequences for both the original Tyrannosaurus and Brachylophosaurus studies. Regardless of the true nature of the dinosaur peptides, our finding highlights the difficulty of differentiating such sequences with confidence. Our results not only imply that cross-contamination cannot be ruled out, but that appropriate measures to test for endogeneity should be further evaluated. © 2017 The Authors.

Surface chemical immobilization of bioactive peptides on synthetic polymers for cardiac tissue engineering.

Science.gov (United States)

Rosellini, Elisabetta; Cristallini, Caterina; Guerra, Giulio D; Barbani, Niccoletta

2015-01-01

The aim of this work was the development of new synthetic polymeric systems, functionalized by surface chemical modification with bioactive peptides, for myocardial tissue engineering. Polycaprolactone and a poly(ester-ether-ester) block copolymer synthesized in our lab, polycaprolactone-poly(ethylene oxide)-polycaprolactone (PCL-PEO-PCL), were used as the substrates to be modified. Two pentapeptides, H-Gly-Arg-Gly-Asp-Ser-OH (GRGDS) from fibronectin and H-Tyr-Ile-Gly-Ser-Arg-OH (YIGSR) from laminin, were used for the functionalization. Polymeric membranes were obtained by casting from solutions and then functionalized by means of alkaline hydrolysis and subsequent coupling of the bioactive molecules through 1-(3-dimethylaminopropyl)-3-ethylcarbodimide hydrochloride/N-hydroxysuccinimide chemistry. The hydrolysis conditions, in terms of hydrolysis time, temperature, and sodium hydroxide concentration, were optimized for the two materials. The occurrence of the coupling reaction was demonstrated by infrared spectroscopy, as the presence on the functionalized materials of the absorption peaks typical of the two peptides. The peptide surface density was determined by chromatographic analysis and the distribution was studied by infrared chemical imaging. The results showed a nearly homogeneous peptide distribution, with a density above the minimum value necessary to promote cell adhesion. Preliminary in vitro cell culture studies demonstrated that the introduction of the bioactive molecules had a positive effect on improving C2C12 myoblasts growth on the synthetic materials.

General protein-protein cross-linking.

Science.gov (United States)

Alegria-Schaffer, Alice

2014-01-01

This protocol describes a general protein-to-protein cross-linking procedure using the water-soluble amine-reactive homobifunctional BS(3) (bis[sulfosuccinimidyl] suberate); however, the protocol can be easily adapted using other cross-linkers of similar properties. BS(3) is composed of two sulfo-NHS ester groups and an 11.4 Å linker. Sulfo-NHS ester groups react with primary amines in slightly alkaline conditions (pH 7.2-8.5) and yield stable amide bonds. The reaction releases N-hydroxysuccinimide (see an application of NHS esters on Labeling a protein with fluorophores using NHS ester derivitization). © 2014 Elsevier Inc. All rights reserved.

Seed-mediated synthesis of cross-linked Pt-NiO nanochains for methanol oxidation

Science.gov (United States)

Gu, Zhulan; Bin, Duan; Feng, Yue; Zhang, Ke; Wang, Jin; Yan, Bo; Li, Shumin; Xiong, Zhiping; Wang, Caiqin; Shiraishi, Yukihide; Du, Yukou

2017-07-01

A simple method was reported for employing NiO nanoparticles act as seeds and then different amounts of Pt2+ were reduced on the NiO nanoparticles, forming a cross-linked Pt-NiO nanocatalysts. These as-prepared catalysts were characterized using different physical-chemical techniques, including X-ray diffraction (XRD), transmission electron microscopy (TEM) and X-ray photoelectron spectroscopy (XPS). The results indicate that the morphology of the cross-linked Pt-NiO nanochain was successfully produced regardless of the molar ratio of Pt2+ to NiO precursors. The electrochemical characteristics of Pt-NiO nanochain catalysts were evaluated for the oxidation of methanol as a model reaction, which verify that the Pt-NiO catalysts show enhanced activity and high stability in comparison with the commercial Pt/C catalyst. The optimized ratio of Pt to NiO is 1:1, then tuned by simple adjusting the feed ratio of the precursors as well. The synthesized nanocatalysts will be found the great potential applications as electrocatalysts for fuel cells owe to their enhanced catalytic performance and long-term stability.

Novel magnetic cross-linked lipase aggregates for improving the resolution of (R, S)-2-octanol.

Science.gov (United States)

Liu, Ying; Guo, Chen; Liu, Chun-Zhao

2015-03-01

Novel magnetic cross-linked lipase aggregates were fabricated by immobilizing the cross-linked lipase aggregates onto magnetic particles with a high number of -NH2 terminal groups using p-benzoquinone as the cross-linking agent. At the optimal fabrication conditions, 100% of immobilization efficiency and 139% of activity recovery of the magnetic cross-linked lipase aggregates were achieved. The magnetic cross-linked lipase aggregates were able to efficiently resolve (R, S)-2-octanol, and retained 100% activity and 100% enantioselectivity after 10 cycles of reuse, whereas the cross-linked lipase aggregates only retained about 50% activity and 70% enantioselectivity due to insufficient cross-linking. These results provide a great potential for industrial applications of the magnetic cross-linked lipase aggregates. © 2014 Wiley Periodicals, Inc.

Thermal Stability of RNA Phage Virus-Like Particles Displaying Foreign Peptides

Directory of Open Access Journals (Sweden)

Peabody David S

2011-05-01

Full Text Available Abstract Background To be useful for genetic display of foreign peptides a viral coat protein must tolerate peptide insertions without major disruption of subunit folding and capsid assembly. The folding of the coat protein of RNA phage MS2 does not normally tolerate insertions in its AB-loop, but an engineered single-chain dimer readily accepts them as long as they are restricted to one of its two halves. Results Here we characterize the effects of peptide insertions on the thermal stabilities of MS2 virus-like particles (VLPs displaying a variety of different peptides in one AB-loop of the coat protein single-chain dimer. These particles typically denature at temperatures around 5-10°C lower than unmodified VLPs. Even so, they are generally stable up to about 50°C. VLPs of the related RNA phage PP7 are cross-linked with intersubunit disulfide bonds and are therefore significantly more stable. An AB-loop insertion also reduces the stability of PP7 VLPs, but they only begin to denature above about 70°C. Conclusions VLPs assembled from MS2 single-chain dimer coat proteins with peptide insertions in one of their AB-loops are somewhat less stable than the wild-type particle, but still resist heating up to about 50°C. Because they possess disulfide cross-links, PP7-derived VLPs provide an alternate platform with even higher stability.

A Comparative Study of the Characteristics of Cross-Linked, Oxidized and Dual-Modified Rice Starches

OpenAIRE

Xiao, Hua-Xi; Lin, Qin-Lu; Liu, Gao-Qiang; Yu, Feng-Xiang

2012-01-01

Rice starch was cross-linked with epichlorohydrin (0.3%, w/w, on a dry starch basis) and oxidized with sodium hypochlorite (2.5% w/w), respectively. Two dual-modified rice starch samples (oxidized cross-linked rice starch and cross-linked oxidized rice starch) were obtained by the oxidation of cross-linked rice starch and the cross-linking of oxidized rice starch at the same level of reagents. The physicochemical properties of native rice starch, cross-linked rice starch and oxidized rice sta...

Effects of Supercritical CO 2 Conditioning on Cross-Linked Polyimide Membranes

KAUST Repository

Kratochvil, Adam M.; Koros, William J.

2010-01-01

The effects of supercritical CO2 (scCO2) conditioning on high-performance cross-linked polyimide membranes is examined through gas permeation and sorption experiments. Under supercritical conditions, the cross-linked polymers do not exhibit a

PeptideNavigator: An interactive tool for exploring large and complex data sets generated during peptide-based drug design projects.

Science.gov (United States)

Diller, Kyle I; Bayden, Alexander S; Audie, Joseph; Diller, David J

2018-01-01

There is growing interest in peptide-based drug design and discovery. Due to their relatively large size, polymeric nature, and chemical complexity, the design of peptide-based drugs presents an interesting "big data" challenge. Here, we describe an interactive computational environment, PeptideNavigator, for naturally exploring the tremendous amount of information generated during a peptide drug design project. The purpose of PeptideNavigator is the presentation of large and complex experimental and computational data sets, particularly 3D data, so as to enable multidisciplinary scientists to make optimal decisions during a peptide drug discovery project. PeptideNavigator provides users with numerous viewing options, such as scatter plots, sequence views, and sequence frequency diagrams. These views allow for the collective visualization and exploration of many peptides and their properties, ultimately enabling the user to focus on a small number of peptides of interest. To drill down into the details of individual peptides, PeptideNavigator provides users with a Ramachandran plot viewer and a fully featured 3D visualization tool. Each view is linked, allowing the user to seamlessly navigate from collective views of large peptide data sets to the details of individual peptides with promising property profiles. Two case studies, based on MHC-1A activating peptides and MDM2 scaffold design, are presented to demonstrate the utility of PeptideNavigator in the context of disparate peptide-design projects. Copyright © 2017 Elsevier Ltd. All rights reserved.

Computer simulation of randomly cross-linked polymer networks

International Nuclear Information System (INIS)

Williams, Timothy Philip

2002-01-01

In this work, Monte Carlo and Stochastic Dynamics computer simulations of mesoscale model randomly cross-linked networks were undertaken. Task parallel implementations of the lattice Monte Carlo Bond Fluctuation model and Kremer-Grest Stochastic Dynamics bead-spring continuum model were designed and used for this purpose. Lattice and continuum precursor melt systems were prepared and then cross-linked to varying degrees. The resultant networks were used to study structural changes during deformation and relaxation dynamics. The effects of a random network topology featuring a polydisperse distribution of strand lengths and an abundance of pendant chain ends, were qualitatively compared to recent published work. A preliminary investigation into the effects of temperature on the structural and dynamical properties was also undertaken. Structural changes during isotropic swelling and uniaxial deformation, revealed a pronounced non-affine deformation dependant on the degree of cross-linking. Fractal heterogeneities were observed in the swollen model networks and were analysed by considering constituent substructures of varying size. The network connectivity determined the length scales at which the majority of the substructure unfolding process occurred. Simulated stress-strain curves and diffraction patterns for uniaxially deformed swollen networks, were found to be consistent with experimental findings. Analysis of the relaxation dynamics of various network components revealed a dramatic slowdown due to the network connectivity. The cross-link junction spatial fluctuations for networks close to the sol-gel threshold, were observed to be at least comparable with the phantom network prediction. The dangling chain ends were found to display the largest characteristic relaxation time. (author)

Permanent Set of Cross-Linking Networks: Comparison of Theory with Molecular Dynamics Simulations

DEFF Research Database (Denmark)

Rottach, Dana R.; Curro, John G.; Budzien, Joanne

2006-01-01

The permanent set of cross-linking networks is studied by molecular dynamics. The uniaxial stress for a bead-spring polymer network is investigated as a function of strain and cross-link density history, where cross-links are introduced in unstrained and strained networks. The permanent set...

Solvent-free covalent functionalization of multi-walled carbon nanotubes and nanodiamond with diamines: Looking for cross-linking effects

International Nuclear Information System (INIS)

Basiuk, Elena V.; Basiuk, Vladimir A.; Meza-Laguna, Víctor; Contreras-Torres, Flavio F.; Martínez, Melchor; Rojas-Aguilar, Aarón; Salerno, Marco

2012-01-01

Highlights: ► Diamines were used for one-step functionalization of nanotubes and nanodiamond. ► We found experimental evidences of cross-linking effects in these nanomaterials. ► We found a strong orientation effect in the functionalized carbon nanotubes. - Abstract: The covalent functionalization of carbon nanomaterials with diamines is a way to enhance the mechanical strength of nanocomposites due to cross-linking effects, to form complex networks for nanotube-based electronic circuits, as well as is important for a number of biomedical applications. The main goal of the present work was to covalently functionalize pristine multi-walled carbon nanotubes and nanodiamond with three aliphatic diamines (1,8-diaminooctane, 1,10-diaminodecane and 1,12-diaminododecane) and one aromatic diamine (1,5-diaminonaphthalene), by employing a simple one-step solvent-free methodology, which is based on thermal instead of chemical activation. We looked for experimental evidences of cross-linking effects in the carbon nanomaterials synthesized by using solubility/dispersibility tests, atomic force microscopy, scanning and transmission electron microscopy, as well as Fourier-transform infrared spectroscopy and thermogravimetric analysis for additional characterization.

Mapping the signal peptide binding and oligomer contact sites of the core subunit of the pea twin arginine protein translocase.

Science.gov (United States)

Ma, Xianyue; Cline, Kenneth

2013-03-01

Twin arginine translocation (Tat) systems of thylakoid and bacterial membranes transport folded proteins using the proton gradient as the sole energy source. Tat substrates have hydrophobic signal peptides with an essential twin arginine (RR) recognition motif. The multispanning cpTatC plays a central role in Tat operation: It binds the signal peptide, directs translocase assembly, and may facilitate translocation. An in vitro assay with pea (Pisum sativum) chloroplasts was developed to conduct mutagenesis and analysis of cpTatC functions. Ala scanning mutagenesis identified mutants defective in substrate binding and receptor complex assembly. Mutations in the N terminus (S1) and first stromal loop (S2) caused specific defects in signal peptide recognition. Cys matching between substrate and imported cpTatC confirmed that S1 and S2 directly and specifically bind the RR proximal region of the signal peptide. Mutations in four lumen-proximal regions of cpTatC were defective in receptor complex assembly. Copurification and Cys matching analyses suggest that several of the lumen proximal regions may be important for cpTatC-cpTatC interactions. Surprisingly, RR binding domains of adjacent cpTatCs directed strong cpTatC-cpTatC cross-linking. This suggests clustering of binding sites on the multivalent receptor complex and explains the ability of Tat to transport cross-linked multimers. Transport of substrate proteins cross-linked to the signal peptide binding site tentatively identified mutants impaired in the translocation step.

Targeting a Cross-Reactive Gly m 5 Soy Peptide as Responsible for Hypersensitivity Reactions in a Milk Allergy Mouse Model

Science.gov (United States)

Curciarello, Renata; Smaldini, Paola L.; Candreva, Angela M.; González, Virginia; Parisi, Gustavo; Cauerhff, Ana; Barrios, Ivana; Blanch, Luis Bruno; Fossati, Carlos A.

2014-01-01

Background Cross-reactivity between soybean allergens and bovine caseins has been previously reported. In this study we aimed to map epitopes of the major soybean allergen Gly m 5 that are co-recognized by casein specific antibodies, and to identify a peptide responsible for the cross-reactivity. Methods Cow's milk protein (CMP)-specific antibodies were used in different immunoassays (immunoblotting, ELISA, ELISA inhibition test) to evaluate the in vitro recognition of soybean proteins (SP). Recombinant Gly m 5 (α), a truncated fragment containing the C-terminal domain (α-T) and peptides of α-T were obtained and epitope mapping was performed with an overlapping peptide assay. Bioinformatics tools were used for epitope prediction by sequence alignment, and for modelling the cross-recognized soy proteins and peptides. The binding of SP to a monoclonal antibody was studied by surface Plasmon resonance (SPR). Finally, the in vivo cross-recognition of SP was assessed in a mouse model of milk allergy. Results Both α and α-T reacted with the different CMP-specific antibodies. α-T contains IgG and IgE epitopes in several peptides, particularly in the peptide named PA. Besides, we found similar values of association and dissociation constants between the α-casein specific mAb and the different milk and soy components. The food allergy mouse model showed that SP and PA contain the cross-reactive B and T epitopes, which triggered hypersensitivity reactions and a Th2-mediated response on CMP-sensitized mice. Conclusions Gly m 5 is a cross-reactive soy allergen and the α-T portion of the molecule contains IgG and IgE immunodominant epitopes, confined to PA, a region with enough conformation to be bound by antibodies. These findings contribute to explain the intolerance to SP observed in IgE-mediated CMA patients, primarily not sensitised to SP, as well as it sets the basis to propose a mucosal immunotherapy for milk allergy using this soy peptide. PMID:24416141

The conformational stability and flexibility of insulin with an additional intramolecular cross-link

International Nuclear Information System (INIS)

Brems, D.N.; Brown, P.L.; Nakagawa, S.H.; Tager, H.S.

1991-01-01

The conformational stability and flexibility of insulin containing a cross-link between the alpha-amino group of the A-chain to the epsilon-amino group of Lys29 of the B-chain was examined. The cross-link varied in length from 2 to 12 carbon atoms. The conformational stability was determined by guanidine hydrochloride-induced equilibrium denaturation and flexibility was assessed by H2O/D2O amide exchange. The cross-link has substantial effects on both conformational stability and flexibility which depend on its length. In general, the addition of a cross-link enhances conformational stability and decreases flexibility. The optimal length for enhanced stability and decreased flexibility was the 6-carbon link. For the 6-carbon link the Gibbs free energy of unfolding was 8.0 kcal/mol compared to 4.5 kcal/mol for insulin, and the amide exchange rate decreased by at least 3-fold. A very short cross-link (i.e. the 2-carbon link) caused conformational strain that was detectable by a lack of stabilization in the Gibbs free energy of unfolding and enhancement in the amide exchange rate compared to insulin. The effect of the cross-link length on insulin hydrodynamic properties is discussed relative to previously obtained receptor binding results

WICH, a member of WASP-interacting protein family, cross-links actin filaments

International Nuclear Information System (INIS)

Kato, Masayoshi; Takenawa, Tadaomi

2005-01-01

In yeast, Verprolin plays an important role in rearrangement of the actin cytoskeleton. There are three mammalian homologues of Verprolin, WIP, CR16, and WICH, and all of them bind actin and Wiskott-Aldrich syndrome protein (WASP) and/or neural-WASP. Here, we describe a novel function of WICH. In vitro co-sedimentation analysis revealed that WICH not only binds to actin filaments but also cross-links them. Fluorescence and electron microscopy detected that this cross-linking results in straight bundled actin filaments. Overexpression of WICH alone in cultured fibroblast caused the formation of thick actin fibers. This ability of WICH depended on its own actin cross-linking activity. Importantly, the actin cross-linking activity of WICH was modified through a direct association with N-WASP. Taken together, these data suggest that WICH induces a bundled form of actin filament with actin cross-linking activity and the association with N-WASP suppresses that activity. WICH thus appears to be a novel actin bundling protein

Unique and cross-reactive T cell epitope peptides of the major Bahia grass pollen allergen, Pas n 1.

Science.gov (United States)

Etto, Tamara; de Boer, Carmela; Prickett, Sara; Gardner, Leanne M; Voskamp, Astrid; Davies, Janet M; O'Hehir, Robyn E; Rolland, Jennifer M

2012-01-01

Bahia grass pollen (BaGP) is a major cause of allergic rhinitis. Subcutaneous allergen-specific immunotherapy is effective for grass pollen allergy, but is unsuitable for patients with moderate to severe asthma due to the risk of anaphylaxis. T cell-reactive but IgE nonreactive peptides provide a safer treatment option. This study aimed to identify and characterize dominant CD4(+) T cell epitope peptides of the major BaGP allergen, Pas n 1. Pas n 1-specific T cell lines generated from the peripheral blood of BaGP-allergic subjects were tested for proliferative and cytokine response to overlapping 20-mer Pas n 1 peptides. Cross-reactivity to homologous peptides from Lol p 1 and Cyn d 1 of Ryegrass and Bermuda grass pollen, respectively, was assessed using Pas n 1 peptide-specific T cell clones. MHC class II restriction of Pas n 1 peptide T cell recognition was determined by HLA blocking assays and peptide IgE reactivity tested by dot blotting. Three Pas n 1 peptides showed dominant T cell reactivity; 15 of 18 (83%) patients responded to one or more of these peptides. T cell clones specific for dominant Pas n 1 peptides showed evidence of species-specific T cell reactivity as well as cross-reactivity with other group 1 grass pollen allergens. The dominant Pas n 1 T cell epitope peptides showed HLA binding diversity and were non-IgE reactive. The immunodominant T cell-reactive Pas n 1 peptides are candidates for safe immunotherapy for individuals, including those with asthma, who are allergic to Bahia and possibly other grass pollens. Copyright © 2012 S. Karger AG, Basel.

Transglutaminase catalyzed cross-linking of sodium caseinate improves oxidative stability of flaxseed oil emulsion.

Science.gov (United States)

Ma, Hairan; Forssell, Pirkko; Kylli, Petri; Lampi, Anna-Maija; Buchert, Johanna; Boer, Harry; Partanen, Riitta

2012-06-20

Sodium caseinate was modified by transglutaminase catalyzed cross-linking reaction prior to the emulsification process in order to study the effect of cross-linking on the oxidative stability of protein stabilized emulsions. The extent of the cross-linking catalyzed by different dosages of transglutaminase was investigated by following the ammonia production during the reaction and using SDS-PAGE gel. O/W emulsions prepared with the cross-linked and non-cross-linked sodium caseinates were stored for 30 days under the same conditions. Peroxide value measurement, oxygen consumption measurement, and headspace gas chromatography analysis were used to study the oxidative stability of the emulsions. The emulsion made of the cross-linked sodium caseinate showed an improved oxidative stability with reduced formation of fatty acid hydroperoxides and volatiles and a longer period of low rate oxygen consumption. The improving effect of transglutaminase catalyzed cross-linking could be most likely attributed to the enhanced physical stability of the interfacial protein layer against competitive adsorption by oil oxidation products.

Combined cross-linked enzyme aggregates of horseradish peroxidase and glucose oxidase for catalyzing cascade chemical reactions.

Science.gov (United States)

Nguyen, Le Truc; Yang, Kun-Lin

2017-05-01

Cascade reactions involved unstable intermediates are often encountered in biological systems. In this study, we developed combined cross-linked enzyme aggregates (combi-CLEA) to catalyze a cascade reaction which involves unstable hydrogen peroxide as an intermediate. The combi-CLEA contains two enzymes̶ glucose oxidase (GOx) and horseradish peroxidase (HRP) which are cross-linked together as solid aggregates. The first enzyme GOx catalyzes the oxidation of glucose and produces hydrogen peroxide, which is used by the second enzyme HRP to oxidize 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS). The apparent reaction rate of the cascade reaction reaches 10.5±0.5μM/min when the enzyme ratio is 150:1 (GOx:HRP). Interestingly, even in the presence of catalase, an enzyme that quickly decomposes hydrogen peroxide, the reaction rate only decreases by 18.7% to 8.3±0.3μM/min. This result suggests that the intermediate hydrogen peroxide is not decomposed by catalase due to a short diffusion distance between GOx and HRP in the combi-CLEA. Scanning electron microscopy images suggest that combi-CLEA particles are hollow spheres and have an average diameter around 250nm. Because of their size, combi-CLEA particles can be entrapped inside a nylon membrane for detecting glucose by using the cascade reaction. Copyright © 2017 Elsevier Inc. All rights reserved.

A novel strategy for preparing mechanically robust ionically cross-linked alginate hydrogels

International Nuclear Information System (INIS)

Jejurikar, Aparna; Lawrie, Gwen; Groendahl, Lisbeth; Martin, Darren

2011-01-01

The properties of alginate films modified using two cross-linker ions (Ca 2+ and Ba 2+ ), comparing two separate cross-linking techniques (the traditional immersion (IM) method and a new strategy in a pressure-assisted diffusion (PD) method), are evaluated. This was achieved through measuring metal ion content, water uptake and film stability in an ionic solution ([Ca 2+ ] = 2 mM). Characterization of the internal structure and mechanical properties of hydrated films were established by cryogenic scanning electron microscopy and tensile testing, respectively. It was found that gels formed by the PD technique possessed greater stability and did not exhibit any delamination after 21 day immersion as compared to gels formed by the IM technique. The Ba 2+ cross-linked gels possessed significantly higher cross-linking density as reflected in lower water content, a more dense internal structure and higher Young's modulus compared to Ca 2+ cross-linked gels. For the Ca 2+ cross-linked gels, a large improvement in the mechanical properties was observed in gels produced by the PD technique and this was attributed to thicker pore walls observed within the hydrogel structure. In contrast, for the Ba 2+ cross-linked gels, the PD technique resulted in gels that had lower tensile strength and strain energy density and this was attributed to phase separation and larger macropores in this gel.

Antimicrobial Peptide Human Neutrophil Peptide 1 as a Potential Link Between Chronic Inflammation and Ductal Adenocarcinoma of the Pancreas.

Science.gov (United States)

Pausch, Thomas; Adolph, Sarah; Felix, Klaus; Bauer, Andrea S; Bergmann, Frank; Werner, Jens; Hartwig, Werner

Defensins are antimicrobial peptides playing a role in innate immunity, in epithelial cell regeneration, and in carcinogenesis of inflammation-triggered malignancies. We analyzed this role in pancreatic ductal adenocarcinoma (PDAC) in the context of its association with chronic pancreatitis (CP). Human tissue of healthy pancreas, CP, and PDAC was screened for defensins by immunohistochemistry. Defensin α 1 (human neutrophil peptide 1 [HNP-1]) expression was validated using mass spectrometry and microarray analysis. Human neutrophil peptide 1 expression and influences of proinflammatory cytokines (tumor necrosis factor α, interleukin 1β, and interferon γ) were studied in human pancreatic cancer cells (Colo 357, T3M4, PANC-1) and normal human pancreatic duct epithelial cells (HPDE). Accumulation of HNP-1 in malignant pancreatic ductal epithelia was seen. Spectrometry showed increased expression of HNP-1 in CP and even more in PDAC. At RNA level, no significant regulation was found. In cancer cells, HNP-1 expression was significantly higher than in HPDE. Proinflammatory cytokines significantly led to increased HNP-1 levels in culture supernatants and decreased levels in lysates of cancer cells. In HPDE cytokines significantly decreased HNP-1 levels. Inflammatory regulation of HNP-1 in PDAC tissue and cells indicates that HNP-1 may be a link between chronic inflammation and malignant transformation in the pancreas.

Into the intracellular logistics of cross-presentation

Directory of Open Access Journals (Sweden)

Charlotte eSadaka

2012-02-01

Full Text Available The induction of cytotoxic CD8+ T cell responses requires the presentation of antigenic peptides by MHC class I molecules (MHC I. MHC I usually present peptides derived from endogenous proteins. However, some subtypes of dendritic cells (DCs have developed the ability to efficiently present peptides derived from exogenous antigens on MHC I via a process called cross-presentation. Cross-presentation is intimately linked to the induction of anti-viral, -bacterial and -tumor cytotoxic T cell (CTL responses, as well as a wide variety of CTL-mediated diseases and transplant rejections. The molecular and cellular mechanisms underlying cross-presentation have been studied intensively since its original description, yet understanding of this process is incomplete and on the forefront of immunological research. Numerous pathways and models, some of them conflicting, have been described so far. Here, we review the various pathways reported as involved in cross-presentation, highlighting the complexity of this process. We also discuss in detail the different intracellular steps required, from antigen capture and routing, to processing and finally peptide loading, emphasizing the need for a better understanding of the cell biology of this phenomenon.

Cross-linking methods of electrospun fibrinogen scaffolds for tissue engineering applications

International Nuclear Information System (INIS)

Sell, Scott A; Garg, Koyal; McClure, Michael J; Bowlin, Gary L; Francis, Michael P; Simpson, David G

2008-01-01

The purpose of this study was to enhance the mechanical properties and slow the degradation of an electrospun fibrinogen scaffold, while maintaining the scaffold's high level of bioactivity. Three different cross-linkers were used to achieve this goal: glutaraldehyde vapour, 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) in ethanol and genipin in ethanol. Scaffolds with a fibrinogen concentration of 120 mg ml -1 were electrospun and cross-linked with one of the aforementioned cross-linkers. Mechanical properties were determined through uniaxial tensile testing performed on scaffolds incubated under standard culture conditions for 1 day, 7 days and 14 days. Cross-linked scaffolds were seeded with human foreskin fibroblasts (BJ-GFP-hTERT) and cultured for 7, 14 and 21 days, with histology and scanning electron microscopy performed upon completion of the time course. Mechanical testing revealed significantly increased peak stress and modulus values for the EDC and genipin cross-linked scaffolds, with significantly slowed degradation. However, cross-linking with EDC and genipin was shown to have some negative effect on the bioactivity of the scaffolds as cell migration throughout the thickness of the scaffold was slowed.

Evaluation of a high-throughput peptide reactivity format assay for assessment of the skin sensitization potential of chemicals

Directory of Open Access Journals (Sweden)

Chin Lin eWong

2016-03-01

Full Text Available The direct peptide reactivity assay (DPRA is a validated method for in vitro assessment of the skin sensitization potential of chemicals. In the present work, we describe a peptide reactivity assay using 96-well plate format and systematically identified the optimal assay conditions for accurate and reproducible classification of chemicals with known sensitizing capacity. The aim of the research is to ensure that the analytical component of the peptide reactivity assay is robust, accurate and reproducible in accordance with criteria that are used for the validation of bioanalytical methods. Analytical performance was evaluated using quality control samples (QCs; heptapeptides at low, medium and high concentrations and incubation of control chemicals (chemicals with known sensitization capacity, weak, moderate, strong, extreme and non-sensitizers with each of three synthetic heptapeptides, viz Cor1-C420 (Ac-NKKCDLF, cysteine- (Ac-RFAACAA and lysine- (Ac-RFAAKAA containing heptapeptides. The optimal incubation temperature for all three heptapeptides was 25°C. Apparent heptapeptide depletion was affected by vial material composition. Incubation of test chemicals with Cor1-C420, showed that peptide depletion was unchanged in polypropylene vials over 3-days storage in an autosampler but this was not the case for borosilicate glass vials. For cysteine-containing heptapeptide, the concentration was not stable by day 3 post-incubation in borosilicate glass vials. Although the lysine-containing heptapeptide concentration was unchanged in both polypropylene and borosilicate glass vials, the apparent extent of lysine-containing heptapeptide depletion by ethyl acrylate, differed between polypropylene (24.7% and glass (47.3% vials. Additionally, the peptide-chemical complexes for Cor1-C420-cinnamaldehyde and cysteine-containing heptapeptide-2,4-dinitrochlorobenzene were partially reversible during 3-days of autosampler storage. These observations further

Ocean acidification affects marine chemical communication by changing structure and function of peptide signalling molecules.

Science.gov (United States)

Roggatz, Christina C; Lorch, Mark; Hardege, Jörg D; Benoit, David M

2016-12-01

Ocean acidification is a global challenge that faces marine organisms in the near future with a predicted rapid drop in pH of up to 0.4 units by the end of this century. Effects of the change in ocean carbon chemistry and pH on the development, growth and fitness of marine animals are well documented. Recent evidence also suggests that a range of chemically mediated behaviours and interactions in marine fish and invertebrates will be affected. Marine animals use chemical cues, for example, to detect predators, for settlement, homing and reproduction. But, while effects of high CO 2 conditions on these behaviours are described across many species, little is known about the underlying mechanisms, particularly in invertebrates. Here, we investigate the direct influence of future oceanic pH conditions on the structure and function of three peptide signalling molecules with an interdisciplinary combination of methods. NMR spectroscopy and quantum chemical calculations were used to assess the direct molecular influence of pH on the peptide cues, and we tested the functionality of the cues in different pH conditions using behavioural bioassays with shore crabs (Carcinus maenas) as a model system. We found that peptide signalling cues are susceptible to protonation in future pH conditions, which will alter their overall charge. We also show that structure and electrostatic properties important for receptor binding differ significantly between the peptide forms present today and the protonated signalling peptides likely to be dominating in future oceans. The bioassays suggest an impaired functionality of the signalling peptides at low pH. Physiological changes due to high CO 2 conditions were found to play a less significant role in influencing the investigated behaviour. From our results, we conclude that the change of charge, structure and consequently function of signalling molecules presents one possible mechanism to explain altered behaviour under future oceanic p

Dehydration of an azeotrope of ethanol/water by sodium carboxymethylcellulose membranes cross-linked with organic or inorganic cross-linker

Directory of Open Access Journals (Sweden)

2010-11-01

Full Text Available To control the swelling of sodium carboxymethylcellulose (CMCNa membranes, mixtures of CMCNa and glutaraldehyde (GA and mixtures of CMCNa as an organic component and tetraethoxysilane (TEOS as an inorganic component were prepared, and CMCNa/GA cross-linked membranes and CMCNa/TEOS hybrid membranes were formed. In the separation of an ethanol/water azeotrope by pervaporation (PV, the effects of the GA or TEOS content on the water/ethanol selectivity and permeability of these CMCNa/GA cross-linked and CMCNa/TEOS hybrid membranes were investigated. Cross-linked and hybrid membranes containing up to 10 wt% GA or 10 wt% TEOS exhibited higher water/ethanol selectivity than CMCNa membrane without any cross-linker. This resulted from both increased density and depressed swelling of the membranes by the formation of a cross-linked structure. The relationship between the structure of the CMCNa/GA cross-linked membranes and CMCNa/TEOS hybrid membranes and their permeation and separation characteristics for an ethanol/water azeotrope during PV is discussed in detail.

Rheological properties of dispersions of enzymatically cross-linked apo-α-lactalbumin

NARCIS (Netherlands)

Saricay, Yunus; Wierenga, Peter A.; Vries, de Renko

2016-01-01

The enzymatic cross-linking of apo-α-lactalbumin (α-LA) with horseradish peroxidase (HRP) leads to the formation of hydrophilic protein aggregates with controlled size and architecture. We explore the rheological properties of dispersions of these HRP-cross-linked α-LA aggregates with a

Models for stiffening in cross-linked biopolymer networks : A comparative study

NARCIS (Netherlands)

van Dillen, T.; Onck, P. R.; Van der Giessen, E.

In a recent publication, we studied the mechanical stiffening behavior in two-dimensional (2D) cross-linked networks of semiflexible biopolymer filaments under simple shear [Onck, P.R., Koeman, T., Van Dillen, T., Van der Giessen, E., 2005. Alternative explanation of stiffening in cross-linked

Cross-Linked Amylose Bio-Plastic: A Transgenic-Based Compostable Plastic Alternative

Directory of Open Access Journals (Sweden)

Domenico Sagnelli

2017-09-01

Full Text Available Bio-plastics and bio-materials are composed of natural or biomass derived polymers, offering solutions to solve immediate environmental issues. Polysaccharide-based bio-plastics represent important alternatives to conventional plastic because of their intrinsic biodegradable nature. Amylose-only (AO, an engineered barley starch with 99% amylose, was tested to produce cross-linked all-natural bioplastic using normal barley starch as a control. Glycerol was used as plasticizer and citrate cross-linking was used to improve the mechanical properties of cross-linked AO starch extrudates. Extrusion converted the control starch from A-type to Vh- and B-type crystals, showing a complete melting of the starch crystals in the raw starch granules. The cross-linked AO and control starch specimens displayed an additional wide-angle diffraction reflection. Phospholipids complexed with Vh-type single helices constituted an integrated part of the AO starch specimens. Gas permeability tests of selected starch-based prototypes demonstrated properties comparable to that of commercial Mater-Bi© plastic. The cross-linked AO prototypes had composting characteristics not different from the control, indicating that the modified starch behaves the same as normal starch. The data shows the feasibility of producing all-natural bioplastic using designer starch as raw material.

Cross-Linked Amylose Bio-Plastic: A Transgenic-Based Compostable Plastic Alternative

Science.gov (United States)

Sagnelli, Domenico; Kemmer, Gerdi Christine; Holse, Mette; Hebelstrup, Kim H.; Bao, Jinsong; Stelte, Wolfgang; Bjerre, Anne-Belinda; Blennow, Andreas

2017-01-01

Bio-plastics and bio-materials are composed of natural or biomass derived polymers, offering solutions to solve immediate environmental issues. Polysaccharide-based bio-plastics represent important alternatives to conventional plastic because of their intrinsic biodegradable nature. Amylose-only (AO), an engineered barley starch with 99% amylose, was tested to produce cross-linked all-natural bioplastic using normal barley starch as a control. Glycerol was used as plasticizer and citrate cross-linking was used to improve the mechanical properties of cross-linked AO starch extrudates. Extrusion converted the control starch from A-type to Vh- and B-type crystals, showing a complete melting of the starch crystals in the raw starch granules. The cross-linked AO and control starch specimens displayed an additional wide-angle diffraction reflection. Phospholipids complexed with Vh-type single helices constituted an integrated part of the AO starch specimens. Gas permeability tests of selected starch-based prototypes demonstrated properties comparable to that of commercial Mater-Bi© plastic. The cross-linked AO prototypes had composting characteristics not different from the control, indicating that the modified starch behaves the same as normal starch. The data shows the feasibility of producing all-natural bioplastic using designer starch as raw material. PMID:28973963

Cross-Linked Amylose Bio-Plastic: A Transgenic-Based Compostable Plastic Alternative.

Science.gov (United States)

Sagnelli, Domenico; Hooshmand, Kourosh; Kemmer, Gerdi Christine; Kirkensgaard, Jacob J K; Mortensen, Kell; Giosafatto, Concetta Valeria L; Holse, Mette; Hebelstrup, Kim H; Bao, Jinsong; Stelte, Wolfgang; Bjerre, Anne-Belinda; Blennow, Andreas

2017-09-30

Bio-plastics and bio-materials are composed of natural or biomass derived polymers, offering solutions to solve immediate environmental issues. Polysaccharide-based bio-plastics represent important alternatives to conventional plastic because of their intrinsic biodegradable nature. Amylose-only (AO), an engineered barley starch with 99% amylose, was tested to produce cross-linked all-natural bioplastic using normal barley starch as a control. Glycerol was used as plasticizer and citrate cross-linking was used to improve the mechanical properties of cross-linked AO starch extrudates. Extrusion converted the control starch from A-type to Vh- and B-type crystals, showing a complete melting of the starch crystals in the raw starch granules. The cross-linked AO and control starch specimens displayed an additional wide-angle diffraction reflection. Phospholipids complexed with Vh-type single helices constituted an integrated part of the AO starch specimens. Gas permeability tests of selected starch-based prototypes demonstrated properties comparable to that of commercial Mater-Bi © plastic. The cross-linked AO prototypes had composting characteristics not different from the control, indicating that the modified starch behaves the same as normal starch. The data shows the feasibility of producing all-natural bioplastic using designer starch as raw material.

Riboflavin/UVA Collagen Cross-Linking-Induced Changes in Normal and Keratoconus Corneal Stroma

Science.gov (United States)

Hayes, Sally; Boote, Craig; Kamma-Lorger, Christina S.; Rajan, Madhavan S.; Harris, Jonathan; Dooley, Erin; Hawksworth, Nicholas; Hiller, Jennifer; Terill, Nick J.; Hafezi, Farhad; Brahma, Arun K.; Quantock, Andrew J.; Meek, Keith M.

2011-01-01

Purpose To determine the effect of Ultraviolet-A collagen cross-linking with hypo-osmolar and iso-osmolar riboflavin solutions on stromal collagen ultrastructure in normal and keratoconus ex vivo human corneas. Methods Using small-angle X-ray scattering, measurements of collagen D-periodicity, fibril diameter and interfibrillar spacing were made at 1 mm intervals across six normal post-mortem corneas (two above physiological hydration (swollen) and four below (unswollen)) and two post-transplant keratoconus corneal buttons (one swollen; one unswollen), before and after hypo-osmolar cross-linking. The same parameters were measured in three other unswollen normal corneas before and after iso-osmolar cross-linking and in three pairs of swollen normal corneas, in which only the left was cross-linked (with iso-osmolar riboflavin). Results Hypo-osmolar cross-linking resulted in an increase in corneal hydration in all corneas. In the keratoconus corneas and unswollen normal corneas, this was accompanied by an increase in collagen interfibrillar spacing (priboflavin solutions are more likely a consequence of treatment-induced changes in tissue hydration rather than cross-linking. PMID:21850225

Design and Preparation of Cross-Linked Polystyrene Nanoparticles for Elastomer Reinforcement

Directory of Open Access Journals (Sweden)

Ming Lu

2010-01-01

Full Text Available Cross-linked polystyrene (PS particles in a latex form were synthesized by free radical emulsion polymerization. The nano-PS-filled elastomer composites were prepared by the energy-saving latex compounding method. Results showed that the PS particles took a spherical shape in the size of 40–60 nm with a narrow size distribution, and the glass-transition temperature of the PS nanoparticles increased with the cross-linking density. The outcomes from the mechanical properties demonstrated that when filled into styrene-butadiene rubber (SBR, nitrile-butadiene rubber (NBR, and natural rubber (NR, the cross-linked PS nano-particles exhibited excellent reinforcing capabilities in all the three matrices, and the best in the SBR matrix. In comparison with that of the carbon black filled composites, another distinguished advantage of the cross-linked PS particles filled elastomer composites was found to be light weight in density, which could help to save tremendous amount of energy when put into end products.

Cross-linking of rubber in the presence of multi-functional cross-linking aids via thermoreversible Diels-Alder chemistry

NARCIS (Netherlands)

Polgar, L. M.; Fortunato, G.; Araya-Hermosilla, R.; van Duin, M.; Pucci, A.; Picchioni, F.

Furan-functionalized polyketone (PK-FU) was added to a furan-functionalized ethylene-propylene rubber (EPM-FU). The mixture was subsequently cross-linked with a bismaleimide through Diels-Alder chemistry in order to improve the mechanical properties of the rubber. Infrared spectroscopy showed the

Solution processed organic light-emitting diodes using the plasma cross-linking technology

Energy Technology Data Exchange (ETDEWEB)

He, Kongduo [Department of Light Sources and Illuminating Engineering, Fudan University, Shanghai 200433 (China); Liu, Yang [Department of Light Sources and Illuminating Engineering, Fudan University, Shanghai 200433 (China); Engineering Research Center of Advanced Lighting Technology, Ministry of Education, Shanghai 200433 (China); Gong, Junyi; Zeng, Pan; Kong, Xun; Yang, Xilu; Yang, Cheng; Yu, Yan [Department of Light Sources and Illuminating Engineering, Fudan University, Shanghai 200433 (China); Liang, Rongqing [Department of Light Sources and Illuminating Engineering, Fudan University, Shanghai 200433 (China); Engineering Research Center of Advanced Lighting Technology, Ministry of Education, Shanghai 200433 (China); Ou, Qiongrong, E-mail: qrou@fudan.edu.cn [Department of Light Sources and Illuminating Engineering, Fudan University, Shanghai 200433 (China); Engineering Research Center of Advanced Lighting Technology, Ministry of Education, Shanghai 200433 (China)

2016-09-30

Highlights: • Mixed acetylene and Ar plasma treatment makes the organic film surface cross-linked. • The plasma treatment for 30 s does not affect the performance of OLEDs. • Cross-linking surface can resist rinsing and corrosion of organic solvent. • The surface morphology is nearly unchanged after plasma treatment. • The plasma cross-linking method can realize solution processed multilayer OLEDs. - Abstract: Solution processed multilayer organic light-emitting diodes (OLEDs) present challenges, especially regarding dissolution of the first layer during deposition of a second layer. In this work, we first demonstrated a plasma cross-linking technology to produce a solution processed OLED. The surfaces of organic films can be cross-linked after mixed acetylene and Ar plasma treatment for several tens of seconds and resist corrosion of organic solvent. The film thickness and surface morphology of emissive layers (EMLs) with plasma treatment and subsequently spin-rinsed with chlorobenzene are nearly unchanged. The solution processed triple-layer OLED is successfully fabricated and the current efficiency increases 50% than that of the double-layer OLED. Fluorescent characteristics of EMLs are also observed to investigate factors influencing the efficiency of the triple-layer OLED. Plasma cross-linking technology may open up a new pathway towards fabrication of all-solution processed multilayer OLEDs and other soft electronic devices.

Formulation and Characterization of Glutaraldehyde Cross-Linked ...

African Journals Online (AJOL)

... drug/polymer ratio, volume of cross linking agent and volume of surfactant were ... The microspheres were characterized for entrapment efficiency, drug loading, ... size distribution (105 – 219 μm) and an entrapment efficiency of up to 73 %.

Characterization of the functional epitope on the urokinase receptor. Complete alanine scanning mutagenesis supplemented by chemical cross-linking

DEFF Research Database (Denmark)

Gårdsvoll, Henrik; Gilquin, Bernard; Le Du, Marie Hélène

2006-01-01

a comprehensive alanine scanning mutagenesis of uPAR combined with low resolution distance constraints defined within the complex using chemical cross-linkers as molecular rulers. The kinetic rate constants for the interaction between pro-uPA and 244 purified uPAR mutants with single-site replacements were...

Structuring of Amide Cross-Linked Non-Bridged and Bridged Alkyl-Based Silsesquioxanes.

Science.gov (United States)

Nunes, S C; de Zea Bermudez, V

2018-02-06

The development of sophisticated organized materials exhibiting enhanced properties is a challenging topic of the domain of organic/inorganic hybrid materials. This review, composed of four sections, reports the work we have carried out over the last 10 years on the synthesis of amide cross-linked alkyl/siloxane hybrids by means of sol-gel chemistry and self-directed assembly/self-organization routes relying on weak interactions (hydrophobic interactions and hydrogen bonding). The various as-produced lamellar structures displaying a myriad of morphologies, often closely resembling those found in natural materials, are discussed. The major role played by the synthetic conditions (pH, water content, co-solvent(s) nature/concentration and dopant presence/concentration), the alkyl chains (length and presence of ramification or not) and the number of the amide cross-links present in the precursor, is evidenced. Examples of highly organized hybrids structures incorporating ionic species (alkali and alkaline earth metal salts) and optically-active centers (organic dyes and lanthanide ions) are described. A useful qualitative relationship deduced between the emission quantum yield of the ordered hybrid materials and the degree of order of the hydrogen-bonded network is highlighted. © 2018 The Chemical Society of Japan & Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Preparation and Characterization of a Cross-linked Matrimid/Polyvinylidene Fluoride Composite Membrane for H2/N2 Separation

Directory of Open Access Journals (Sweden)

Mahmood Esmaeilipur

2017-01-01

Full Text Available A double layer composite membrane was fabricated by matrimid 5218 as a selective layer on polyvinylidene fluoride (PVDF, a porous asymmetric membrane, as a sublayer. The effect of chemical cross-linking of Matrimid 5218 by ethylenediamine (EDA was investigated on gas transport properties of the corresponding membrane. The permeability levels of hydrogen (H2 and nitrogen (N2 were measured through the fabricated composite membranes at 25°C under pressure range of 2-8 bar. Scanning electron microscopy (SEM was used for morphological observations of the composite membranes. The Matrimid membranes before and after cross-linking modification were characterized by the Fourier transform infrared (FTIR spectroscopy, X-ray diffraction (XRD and density measurement. The FTIR results showed the conversion of imide functional groups into amide through the crosslinking reaction in Matrimid. The XRD results demonstrated a reduction in d-spacing between the polymer chains through cross-linking reaction. Measuring the density of each membrane's partial selective layer and calculating the corresponding fractional free volume revealed an increase in the density and reduced free volumes in Matrimid through the cross-linking reaction. Moreover, by increasing the EDA concentration, the gas permeability in each membrane decreased significantly for nitrogen compared to hydrogen which could be related to lower gas diffusivity through chain packing due to cross-linking of the polymer. The H2/N2 selectivity at 2 bar increased through the cross-linking modification from 56.5 for the pure Matrimid to 79.4 for the composite membrane containing 12 wt% EDA. The effect of pressure on gas permeability through the composite membranes was investigated and the results found to be in agreement with the behavior of less soluble gases in the glassy polymers. Moreover, the H2/N2 selectivity decreased first at low EDA content (0-4 wt%, before reaching a constant value at 8 wt% EDA and

Fluorescence spectroscopic study of the aggregation behavior of non-cross-linked and cross-linked poly(alkylmethyldiallylammonium bromides) having decyl, octyl, and hexyl side chains in aqueous solution

NARCIS (Netherlands)

Wang, G.J; Engberts, J.B.F.N.

1996-01-01

The conformational state of a series of non-cross-linked and cross-linked poly(alkylmethyldiallylammonium bromides) bearing decyl, octyl, and hexyl side chains ((CL)-CopolC1-10, (CL)-CopolC1-8, and (CL)-CopolC1-6, respectively) in aqueous solutions were investigated by fluorescence spectroscopy

Assessment of UVA-Riboflavin Corneal Cross-Linking Using Small Amplitude Oscillatory Shear Measurements.

Science.gov (United States)

Aslanides, Ioannis M; Dessi, Claudia; Georgoudis, Panagiotis; Charalambidis, Georgios; Vlassopoulos, Dimitris; Coutsolelos, Athanassios G; Kymionis, George; Mukherjee, Achyut; Kitsopoulos, Theofanis N

2016-04-01

The effect of ultraviolet (UV)-riboflavin cross-linking (CXL) has been measured primarily using the strip extensometry technique. We propose a simple and reliable methodology for the assessment of CXL treatment by using an established rheologic protocol based on small amplitude oscillatory shear (SAOS) measurements. It provides information on the average cross-link density and the elastic modulus of treated cornea samples. Three fresh postmortem porcine corneas were used to study the feasibility of the technique, one serving as control and two receiving corneal collagen cross-linking treatment. Subsequently, five pairs of fresh postmortem porcine corneas received corneal collagen cross-linking treatment with riboflavin and UVA-irradiation (370 nm; irradiance of 3 mW/cm2) for 30 minutes (Dresden protocol); the contralateral porcine corneas were used as control samples. After the treatment, the linear viscoelastic moduli of the corneal samples were measured using SAOS measurements and the average cross-linking densities extracted. For all cases investigated, the dynamic moduli of the cross-linked corneas were higher compared to those of the corresponding control samples. The increase of the elastic modulus of the treated samples was between 122% and 1750%. The difference was statistically significant for all tested samples (P = 0.018, 2-tailed t-test). We report a simple and accurate methodology for quantifying the effects of cross-linking on porcine corneas treated with the Dresden protocol by means of SAOS measurements in the linear regime. The measured dynamic moduli, elastic and viscous modulus, represent the energy storage and energy dissipation, respectively. Hence, they provide a means to assess the changing physical properties of the cross-linked collagen networks after CXL treatment.

Carboxymethyl starch cross-linked by electron beam radiation in presence of acrylic acid sensitizer

International Nuclear Information System (INIS)

Doan Binh; Nguyen Thanh Duoc; Pham Thi Thu Hong

2013-01-01

Carboxymethyl starch (CMS) can be cross-linked by electron beam radiation to form a biocompatible and environment-friendly hydrogel at a high absorbed dose and a condensed CMS concentration. Acrylic acid (AAc) can be used as a sensitizer in order to reduce the absorbed doses to an acceptable certain level. At an absorbed dose of 3-4 kGy, the gel content of crosslinked CMS can be obtained about 50% with 5% (w/w) AAc concentration used. The compressive strength of CMS samples increased with increasing their cross-linked densities due to raising absorbed doses. The swelling ratio of cross-linked CMS was also attainable at a maximum of 50 times in the distilled water. The enzymatic degradation of cross-linked CMS was carried out in acetate buffer pH 4.6 with 0.1% α-amylase enzymatic solution incubated at 40℃ for 6 h. The crosslinked CMS samples were degraded slower than uncrosslinked CMS ones. The results indicated that the highly cross-linked CMS was almost fully degradable when the enzymatic hydrolysis was performed during 6 h. The FT IR spectra of cross-linked CMS in the presence of AAc were examined to observe the carboxyl group of AAc in the structure of cross-linked CMS. The hydrophilic of cross-linked CMS surface was determined by a contact-angle analysis. (authors)

Programmed Nanomaterial Assemblies in Large Scales: Applications of Synthetic and Genetically- Engineered Peptides to Bridge Nano-Assemblies and Macro-Assemblies

Energy Technology Data Exchange (ETDEWEB)

Matsui, Hiroshi

2014-09-09

Work is reported in these areas: Large-scale & reconfigurable 3D structures of precise nanoparticle assemblies in self-assembled collagen peptide grids; Binary QD-Au NP 3D superlattices assembled with collagen-like peptides and energy transfer between QD and Au NP in 3D peptide frameworks; Catalytic peptides discovered by new hydrogel-based combinatorial phage display approach and their enzyme-mimicking 2D assembly; New autonomous motors of metal-organic frameworks (MOFs) powered by reorganization of self-assembled peptides at interfaces; Biomimetic assembly of proteins into microcapsules on oil-in-water droplets with structural reinforcement via biomolecular recognition-based cross-linking of surface peptides; and Biomimetic fabrication of strong freestanding genetically-engineered collagen peptide films reinforced by quantum dot joints. We gained the broad knowledge about biomimetic material assembly from nanoscale to microscale ranges by coassembling peptides and NPs via biomolecular recognition. We discovered: Genetically-engineered collagen-like peptides can be self-assembled with Au NPs to generate 3D superlattices in large volumes (> μm{sup 3}); The assembly of the 3D peptide-Au NP superstructures is dynamic and the interparticle distance changes with assembly time as the reconfiguration of structure is triggered by pH change; QDs/NPs can be assembled with the peptide frameworks to generate 3D superlattices and these QDs/NPs can be electronically coupled for the efficient energy transfer; The controlled assembly of catalytic peptides mimicking the catalytic pocket of enzymes can catalyze chemical reactions with high selectivity; and, For the bacteria-mimicking swimmer fabrication, peptide-MOF superlattices can power translational and propellant motions by the reconfiguration of peptide assembly at the MOF-liquid interface.

Solvent-free covalent functionalization of multi-walled carbon nanotubes and nanodiamond with diamines: Looking for cross-linking effects

Energy Technology Data Exchange (ETDEWEB)

Basiuk, Elena V., E-mail: elenagd@unam.mx [Centro de Ciencias Aplicadas y Desarrollo Tecnologico, Universidad Nacional Autonoma de Mexico, Circuito Exterior, Ciudad Universitaria, 04510 Mexico, D.F. (Mexico); Nanophysics Department, Italian Institute of Technology, via Morego 30, 16163 Genova, Liguria (Italy); Basiuk, Vladimir A. [Nanophysics Department, Italian Institute of Technology, via Morego 30, 16163 Genova, Liguria (Italy); Instituto de Ciencias Nucleares, Universidad Nacional Autonoma de Mexico, Circuito Exterior, Ciudad Universitaria, 04510 Mexico, D.F. (Mexico); Facultad de Ciencias, Universidad Autonoma del Estado de Morelos, Av. Universidad 1001, Col. Chamilpa, 62209 Cuernavaca, Morelos (Mexico); Meza-Laguna, Victor; Contreras-Torres, Flavio F.; Martinez, Melchor [Centro de Ciencias Aplicadas y Desarrollo Tecnologico, Universidad Nacional Autonoma de Mexico, Circuito Exterior, Ciudad Universitaria, 04510 Mexico, D.F. (Mexico); Rojas-Aguilar, Aaron [Centro de Investigacion y de Estudios Avanzados, Instituto Politecnico Nacional, Av. Instituto Politecnico Nacional 2508, Col. San Pedro Zacatenco, 07360 Mexico, D.F. (Mexico); Salerno, Marco [Nanophysics Department, Italian Institute of Technology, via Morego 30, 16163 Genova, Liguria (Italy); and others

2012-10-15

Highlights: Black-Right-Pointing-Pointer Diamines were used for one-step functionalization of nanotubes and nanodiamond. Black-Right-Pointing-Pointer We found experimental evidences of cross-linking effects in these nanomaterials. Black-Right-Pointing-Pointer We found a strong orientation effect in the functionalized carbon nanotubes. - Abstract: The covalent functionalization of carbon nanomaterials with diamines is a way to enhance the mechanical strength of nanocomposites due to cross-linking effects, to form complex networks for nanotube-based electronic circuits, as well as is important for a number of biomedical applications. The main goal of the present work was to covalently functionalize pristine multi-walled carbon nanotubes and nanodiamond with three aliphatic diamines (1,8-diaminooctane, 1,10-diaminodecane and 1,12-diaminododecane) and one aromatic diamine (1,5-diaminonaphthalene), by employing a simple one-step solvent-free methodology, which is based on thermal instead of chemical activation. We looked for experimental evidences of cross-linking effects in the carbon nanomaterials synthesized by using solubility/dispersibility tests, atomic force microscopy, scanning and transmission electron microscopy, as well as Fourier-transform infrared spectroscopy and thermogravimetric analysis for additional characterization.

Role of special cross-links in structure formation of bacterial DNA polymer

Science.gov (United States)

Agarwal, Tejal; Manjunath, G. P.; Habib, Farhat; Lakshmi Vaddavalli, Pavana; Chatterji, Apratim

2018-01-01

Using data from contact maps of the DNA-polymer of Escherichia coli (E. Coli) (at kilobase pair resolution) as an input to our model, we introduce cross-links between monomers in a bead-spring model of a ring polymer at very specific points along the chain. Via suitable Monte Carlo simulations, we show that the presence of these cross-links leads to a particular organization of the chain at large (micron) length scales of the DNA. We also investigate the structure of a ring polymer with an equal number of cross-links at random positions along the chain. We find that though the polymer does get organized at the large length scales, the nature of the organization is quite different from the organization observed with cross-links at specific biologically determined positions. We used the contact map of E. Coli bacteria which has around 4.6 million base pairs in a single circular chromosome. In our coarse-grained flexible ring polymer model, we used 4642 monomer beads and observed that around 80 cross-links are enough to induce the large-scale organization of the molecule accounting for statistical fluctuations caused by thermal energy. The length of a DNA chain even of a simple bacterial cell such as E. Coli is much longer than typical proteins, hence we avoided methods used to tackle protein folding problems. We define new suitable quantities to identify the large scale structure of a polymer chain with a few cross-links.

Biomimetic acellular detoxified glutaraldehyde cross-linked bovine pericardium for tissue engineering

International Nuclear Information System (INIS)

Mathapati, Santosh; Bishi, Dillip Kumar; Guhathakurta, Soma; Cherian, Kotturathu Mammen; Venugopal, Jayarama Reddy; Ramakrishna, Seeram; Verma, Rama Shanker

2013-01-01

Glutaraldehyde (GLUT) processing, cellular antigens, calcium ions in circulation, and phospholipids present in the native tissue are predominantly responsible for calcification, degeneration, and lack of natural microenvironment for host progenitor cell migration in tissue implants. The study presents an improved methodology for adhesion and proliferation of endothelial progenitor cells (EPCs) without significant changes in biomechanical and biodegradation properties of the processed acellular bovine pericardium. The anti-calcification potential of the processed tissue was enhanced by detoxification of GLUT-cross-linked bovine pericardium by decellularization, pretreating it with ethanol or removing the free aldehydes by citric acid treatment and lyophilization. The treated tissues were assessed for biomechanical properties, GLUT ligand quantification, adhesion, proliferation of EPCs, and biodegradability. The results indicate that there was no significant change in biomechanical properties and biodegradability when enzymatic hydrolysis (p > 0.05) is employed in detoxified acellular GLUT cross-linked tissue (DBP–G–CA–ET), compared with the native detoxified GLUT cross-linked bovine pericardium (NBP–G–CA–ET). DBP–G–CA–ET exhibited a significant (p > 0.05) increase in the viability of EPCs and cell adhesion as compared to acellular GLUT cross-linked bovine pericardium (p < 0.05). Lyophilized acellular detoxified GLUT cross-linked bovine pericardium, employed in our study as an alternative to conventional GLUT cross-linked bovine pericardium, might provide longer durability and better biocompatibility, and reduce calcification. The developed bovine pericardium patches could be used in cardiac reconstruction and repair, arteriotomy, soft tissue repair, and general surgical procedures with tissue regeneration dimensions. - Highlights: ► We improved the quality of patch biomaterial for cardiovascular surgical procedures. ► Bovine pericardium was

Biomimetic acellular detoxified glutaraldehyde cross-linked bovine pericardium for tissue engineering

Energy Technology Data Exchange (ETDEWEB)

Mathapati, Santosh; Bishi, Dillip Kumar [Stem Cell and Molecular Biology Laboratory, Department of Biotechnology, Indian Institute of Technology Madras, Chennai (India); Frontier Lifeline Pvt Ltd. and Dr. K. M. Cherian Heart Foundation, Mogappair, Chennai (India); Healthcare and Energy Materials Laboratory, NUSNNI, Faculty of Engineering, National University of Singapore (Singapore); Guhathakurta, Soma [Departmet of Engineering Design, Indian Institute of Technology Madras, Chennai (India); Cherian, Kotturathu Mammen [Frontier Lifeline Pvt Ltd. and Dr. K. M. Cherian Heart Foundation, Mogappair, Chennai (India); Venugopal, Jayarama Reddy; Ramakrishna, Seeram [Healthcare and Energy Materials Laboratory, NUSNNI, Faculty of Engineering, National University of Singapore (Singapore); Verma, Rama Shanker, E-mail: vermars@iitm.ac.in [Stem Cell and Molecular Biology Laboratory, Department of Biotechnology, Indian Institute of Technology Madras, Chennai (India)

2013-04-01

Glutaraldehyde (GLUT) processing, cellular antigens, calcium ions in circulation, and phospholipids present in the native tissue are predominantly responsible for calcification, degeneration, and lack of natural microenvironment for host progenitor cell migration in tissue implants. The study presents an improved methodology for adhesion and proliferation of endothelial progenitor cells (EPCs) without significant changes in biomechanical and biodegradation properties of the processed acellular bovine pericardium. The anti-calcification potential of the processed tissue was enhanced by detoxification of GLUT-cross-linked bovine pericardium by decellularization, pretreating it with ethanol or removing the free aldehydes by citric acid treatment and lyophilization. The treated tissues were assessed for biomechanical properties, GLUT ligand quantification, adhesion, proliferation of EPCs, and biodegradability. The results indicate that there was no significant change in biomechanical properties and biodegradability when enzymatic hydrolysis (p > 0.05) is employed in detoxified acellular GLUT cross-linked tissue (DBP–G–CA–ET), compared with the native detoxified GLUT cross-linked bovine pericardium (NBP–G–CA–ET). DBP–G–CA–ET exhibited a significant (p > 0.05) increase in the viability of EPCs and cell adhesion as compared to acellular GLUT cross-linked bovine pericardium (p < 0.05). Lyophilized acellular detoxified GLUT cross-linked bovine pericardium, employed in our study as an alternative to conventional GLUT cross-linked bovine pericardium, might provide longer durability and better biocompatibility, and reduce calcification. The developed bovine pericardium patches could be used in cardiac reconstruction and repair, arteriotomy, soft tissue repair, and general surgical procedures with tissue regeneration dimensions. - Highlights: ► We improved the quality of patch biomaterial for cardiovascular surgical procedures. ► Bovine pericardium was

Thermoreversible cross-linking of maleated ethylene/propylene copolymers with diamines and amino-alcohols

NARCIS (Netherlands)

Mee, van der M.A.J.; Goossens, J.G.P.; Duin, van M.

2008-01-01

Maleated ethylene/propylene copolymers (MAn-g-EPM) were thermoreversibly cross-linked using diamines and amino-alcohols. Covalent cross-links are formed via the equilibrium reaction of the grafted anhydride groups with di-functional cross-linkers containing combinations of primary (1°) and secondary

SYNTHESIS AND CATALYTIC PROPERTIES OF CROSS-LINKED HYDROPHOBICALLY ASSOCIATING POLY(ALKYLMETHYLDIALLYLAMMONIUM BROMIDES)

NARCIS (Netherlands)

WANG, GJ; ENGBERTS, JBFN

1994-01-01

Cross-linked, hydrophobically associating homo- and copolymers were synthesized by free-radical cyclo(co)polymerization of alkylmethyldiallylammonium bromide monomers with a small amount of N,N'-methylenebisacrylamide in aqueous solution using ammonium persulfate as the initiator. The cross-linked

Conserved Binding Regions Provide the Clue for Peptide-Based Vaccine Development: A Chemical Perspective

Directory of Open Access Journals (Sweden)

Hernando Curtidor

2017-12-01

Full Text Available Synthetic peptides have become invaluable biomedical research and medicinal chemistry tools for studying functional roles, i.e., binding or proteolytic activity, naturally-occurring regions’ immunogenicity in proteins and developing therapeutic agents and vaccines. Synthetic peptides can mimic protein sites; their structure and function can be easily modulated by specific amino acid replacement. They have major advantages, i.e., they are cheap, easily-produced and chemically stable, lack infectious and secondary adverse reactions and can induce immune responses via T- and B-cell epitopes. Our group has previously shown that using synthetic peptides and adopting a functional approach has led to identifying Plasmodium falciparum conserved regions binding to host cells. Conserved high activity binding peptides’ (cHABPs physicochemical, structural and immunological characteristics have been taken into account for properly modifying and converting them into highly immunogenic, protection-inducing peptides (mHABPs in the experimental Aotus monkey model. This article describes stereo–electron and topochemical characteristics regarding major histocompatibility complex (MHC-mHABP-T-cell receptor (TCR complex formation. Some mHABPs in this complex inducing long-lasting, protective immunity have been named immune protection-inducing protein structures (IMPIPS, forming the subunit components in chemically synthesized vaccines. This manuscript summarizes this particular field and adds our recent findings concerning intramolecular interactions (H-bonds or π-interactions enabling proper IMPIPS structure as well as the peripheral flanking residues (PFR to stabilize the MHCII-IMPIPS-TCR interaction, aimed at inducing long-lasting, protective immunological memory.

Epoxides cross-linked hexafluoropropylidene polybenzimidazole membranes for application as high temperature proton exchange membranes

International Nuclear Information System (INIS)

Yang, Jingshuai; Xu, Yixin; Liu, Peipei; Gao, Liping; Che, Quantong; He, Ronghuan

2015-01-01

Covalently cross-linked hexafluoropropylidene polybenzimidazole (F 6 PBI) was prepared and used to fabricate high temperature proton exchange membranes with enhanced mechanical strength against thermoplastic distortion. Three different epoxides, i.e. bisphenol A diglycidyl ether (R 1 ), bisphenol A propoxylate diglycidyl ether (R 2 ) and poly(ethylene glycol) diglycidyl ether (R 3 ), were chosen as the cross-linkers to investigate the influence of their structures on the properties of the cross-linked F 6 PBI membranes. All the cross-linked F 6 PBI membranes displayed excellent stability towards the radical oxidation. Comparing with the pure F 6 PBI membrane, the cross-linked F 6 PBI membranes showed high acid doping level but less swelling after doping phosphoric acid at elevated temperatures. The mechanical strength at 130 °C was improved from 0.4 MPa for F 6 PBI membrane to a range of 0.8–2.0 MPa for the cross-linked F 6 PBI membranes with an acid doping level as high as around 14, especially for that crosslinking with the epoxide (R 3 ), which has a long linear structure of alkyl ether. The proton conductivity of the cross-linked membranes was increased accordingly due to the high acid doping levels. Fuel cell tests demonstrated the technical feasibility of the acid doped cross-linked F 6 PBI membranes for high temperature proton exchange membrane fuel cells

Antimicrobial Peptides from Plants

Directory of Open Access Journals (Sweden)

James P. Tam

2015-11-01

Full Text Available Plant antimicrobial peptides (AMPs have evolved differently from AMPs from other life forms. They are generally rich in cysteine residues which form multiple disulfides. In turn, the disulfides cross-braced plant AMPs as cystine-rich peptides to confer them with extraordinary high chemical, thermal and proteolytic stability. The cystine-rich or commonly known as cysteine-rich peptides (CRPs of plant AMPs are classified into families based on their sequence similarity, cysteine motifs that determine their distinctive disulfide bond patterns and tertiary structure fold. Cystine-rich plant AMP families include thionins, defensins, hevein-like peptides, knottin-type peptides (linear and cyclic, lipid transfer proteins, α-hairpinin and snakins family. In addition, there are AMPs which are rich in other amino acids. The ability of plant AMPs to organize into specific families with conserved structural folds that enable sequence variation of non-Cys residues encased in the same scaffold within a particular family to play multiple functions. Furthermore, the ability of plant AMPs to tolerate hypervariable sequences using a conserved scaffold provides diversity to recognize different targets by varying the sequence of the non-cysteine residues. These properties bode well for developing plant AMPs as potential therapeutics and for protection of crops through transgenic methods. This review provides an overview of the major families of plant AMPs, including their structures, functions, and putative mechanisms.

Cross-validation of commercial enzyme-linked immunosorbent assay and radioimmunoassay for porcine C-peptide concentration measurements in non-human primate serum.

Science.gov (United States)

Gresch, Sarah C; Mutch, Lucas A; Janecek, Jody L; Hegstad-Davies, Rebecca L; Graham, Melanie L

2017-09-01

C-peptide concentration is widely used as a marker of insulin secretion and is especially relevant in evaluating islet graft function following transplantation, because its measurement is not confounded by the presence of exogenous insulin. To address the shortage of human islet donors, the use of porcine islets has been proposed as a possible solution and the stringent pig-to-non-human primate (NHP) model is often the most relevant for pre-clinical evaluation of the potential for diabetes reversal resulting from an islet xenograft. The Millipore radioimmunoassay (RIA) was exclusively used to measure porcine C-peptide (PCP) until 2013 when the assay was discontinued and subsequently a commercially available enzyme-linked immunosorbent assay (ELISA) from Mercodia has been widely adopted. Both assays have been used in pre-clinical trials evaluating the therapeutic potential of xenograft products in reversing diabetes in the pig-to-NHP model, to interpret data in a comparable way it may be useful to perform a harmonization of C-peptide measurements. We performed a method comparison by determining the PCP concentration in 620 serum samples collected from 20 diabetic cynomolgus macaques transplanted with adult porcine islets. All analyses were performed according to manufacturer instructions. With both assays, we demonstrated an acceptable detection limit, precision, and recovery. Linearity of the ELISA met acceptance criteria at all concentrations tested while linearity of the RIA only met acceptance criteria at five of the eight concentrations tested. The RIA had a detection limit of 0.16 ng/mL, and recovery ranged from 82% to 96% and met linearity acceptance criteria at 0.35 ng/mL and from 0.78 to 2.33 ng/mL. The ELISA had a detection limit of 0.03 ng/mL, and recovery ranged from 81% to 115% and met linearity acceptance criteria from 0.08 to 0.85 ng/mL. Both assays had intra-assay precision assay precision ELISA demonstrated a significant correlation with RIA (R

21 CFR 177.1211 - Cross-linked polyacrylate copolymers.

Science.gov (United States)

2010-04-01

... Basic Components of Single and Repeated Use Food Contact Surfaces § 177.1211 Cross-linked polyacrylate... shall be determined using size exclusion chromatography or an equivalent method. When conducting the...

Radiation cross-linked PVC and its applications

International Nuclear Information System (INIS)

Lan Junming; Chen Ruyan; Jia Chaoxing; Li Min; Li Chengxin

1990-04-01

The radiation cross-linking technique is adopted for improving the polyvinyl chloride (PVC) heat-resistance and reducing its thermocontractibility. For examining its properties a small insulation sheath made from modified PVC material has been tested at 260 0 5 seconds. The results obtained were satisfactory

Chitosan Cross-linked Reconstituted Amniotic Collagen Membrane ...

Indian Academy of Sciences (India)

First page Back Continue Last page Overview Graphics. Chitosan Cross-linked Reconstituted Amniotic Collagen Membrane – An Excellent Cell Substratum. The KERATINOCYTE proliferation and Differentiation into multiple layers is due to the presence of type - IV collagen in the amnion. Cultured FIBROBLASTS had good ...

Optimization of Formaldehyde Cross-Linking for Protein Interaction Analysis of Non-Tagged Integrin β1

Directory of Open Access Journals (Sweden)

Cordula Klockenbusch

2010-01-01

Full Text Available Formaldehyde cross-linking of protein complexes combined with immunoprecipitation and mass spectrometry analysis is a promising technique for analysing protein-protein interactions, including those of transient nature. Here we used integrin β1 as a model to describe the application of formaldehyde cross-linking in detail, particularly focusing on the optimal parameters for cross-linking, the detection of formaldehyde cross-linked complexes, the utility of antibodies, and the identification of binding partners. Integrin β1 was found in a high molecular weight complex after formaldehyde cross-linking. Eight different anti-integrin β1 antibodies were used for pull-down experiments and no loss in precipitation efficiency after cross-linking was observed. However, two of the antibodies could not precipitate the complex, probably due to hidden epitopes. Formaldehyde cross-linked complexes, precipitated from Jurkat cells or human platelets and analyzed by mass spectrometry, were found to be composed of integrin β1, α4 and α6 or β1, α6, α2, and α5, respectively.

Optimization of Formaldehyde Cross-Linking for Protein Interaction Analysis of Non-Tagged Integrin β1

Science.gov (United States)

Klockenbusch, Cordula; Kast, Juergen

2010-01-01

Formaldehyde cross-linking of protein complexes combined with immunoprecipitation and mass spectrometry analysis is a promising technique for analysing protein-protein interactions, including those of transient nature. Here we used integrin β1 as a model to describe the application of formaldehyde cross-linking in detail, particularly focusing on the optimal parameters for cross-linking, the detection of formaldehyde cross-linked complexes, the utility of antibodies, and the identification of binding partners. Integrin β1 was found in a high molecular weight complex after formaldehyde cross-linking. Eight different anti-integrin β1 antibodies were used for pull-down experiments and no loss in precipitation efficiency after cross-linking was observed. However, two of the antibodies could not precipitate the complex, probably due to hidden epitopes. Formaldehyde cross-linked complexes, precipitated from Jurkat cells or human platelets and analyzed by mass spectrometry, were found to be composed of integrin β1, α4 and α6 or β1, α6, α2, and α5, respectively. PMID:20634879

Optical properties of thin films of zinc oxide quantum dots and polydimethylsiloxane: UV-blocking and the effect of cross-linking.

Science.gov (United States)

Eita, Mohamed; El Sayed, Ramy; Muhammed, Mamoun

2012-12-01

Thin films of polydimethylsiloxane (PDMS) and ZnO quantum dots (QDs) were built up as multilayers by spin-coating. The films are characterized by a UV-blocking ability that increases with increasing number of bilayers. Photoluminescence (PL) emission spectra of the thin films occur at 522 nm, which is the PL wavelength of the ZnO QDs dispersion, but with a lower intensity and a quantum yield (QY) less than 1% that of the dispersion. Cross-linking has introduced new features to the absorption spectra in that the absorption peak was absent. These changes were attributed to the morphological and structural changes revealed by transmission electron microscopy (TEM) and Fourier transform infrared spectroscopy (FTIR), respectively. TEM showed that the ZnO particle size in the film increased from 7 (±2.7) nm to 16 (±7.8) upon cross-linking. The FTIR spectra suggest that ZnO QDs are involved in the cross-linking of PDMS and that the surface of the ZnO QDs has been chemically modified. Copyright © 2012 Elsevier Inc. All rights reserved.

Gastrin receptor characterization: affinity cross-linking of the gastrin receptor on canine gastric parietal cells

International Nuclear Information System (INIS)

Matsumoto, M.; Park, J.; Yamada, T.

1987-01-01

The authors applied affinity cross-linking methods to label the gastrin receptor on isolated canine gastric parietal cells in order to elucidate the nature of its chemical structure. 125 I-labeled Leu 15 -gastrin and 125 I-labeled gastrin/sub 2-17/ bound to intact parietal cells and their membranes with equal affinity, and half-maximal inhibition of binding was obtained at an incubation concentration of 3.2 x 10 -10 M unlabeled gastrin. 125 I-gastrin/sub 2-17/ was cross-linked to plasma membranes or intact parietal cells by incubation in disuccinimidyl suberate. The membrane pellets were solubilized with or without dithiothreitol and applied to electrophoresis on 7.5% sodium dodecyl sulfate polyacrylamide gels. Autoradiograms revealed a band of labeling at M/sub r/ 76,000 and labeling of this band was inhibited in a dose-dependent fashion by addition of unlabeled gastrin to the incubation mixture. Dithiothreitol in concentrations as high as 100 mM did not later the electrophoretic mobility of the labeled band. After taking into account the molecular weight of 125 I-gastrin/sub 2-17/, the results suggest that the gastrin receptor on parietal cells is a single protein of M/sub r/ 74,000 without disulfide-linked subunits

Novel thermoplastic vulcanizates (TPVs based on silicone rubber and polyamide exploring peroxide cross-linking

Directory of Open Access Journals (Sweden)

K. Naskar

2014-04-01

Full Text Available Novel thermoplastic vulcanizates (TPVs based on silicone rubber (PDMS and polyamide (PA12 have been prepared by dynamic vulcanization process. The effect of dynamic vulcanization and influence of various types of peroxides as cross-linking agents were studied in detail. All the TPVs were prepared at a ratio of 50/50 wt% of silicone rubber and polyamide. Three structurally different peroxides, namely dicumyl peroxide (DCP, 3,3,5,7,7-pentamethyl 1,2,4-trioxepane (PMTO and cumyl hydroperoxide (CHP were taken for investigation. Though DCP was the best option for curing the silicone rubber, at high temperature it suffers from scorch safety. An inhibitor 2,2,6,6-tetramethylpiperidinyloxyl (TEMPO was added with DCP to stabilize the radicals in order to increase the scorch time. Though CHP (hydroperoxide had higher half life time than DCP at higher temperature, it has no significant effect on cross-linking of silicone rubber. PMTO showed prolonged scorch safety and better cross-linking efficiency rather than the other two. TPVs of DCP and PMTO were made up to 11 minutes of mixing. Increased values of tensile strength and elongation at break of PMTO cross-linked TPV indicate the superiority of PMTO. Scanning electron micrographs correlate with mechanical properties of the TPVs. High storage modulus (E' and lower loss tangent value of the PMTO cross-linked TPV indicate the higher degree of cross-linking which is also well supported by the overall cross-link density value. Thus PMTO was found to be the superior peroxide for cross-linking of silicone rubber at high temperature.

Fabrication of Si negative electrodes for Li-ion batteries (LIBs) using cross-linked polymer binders.

Science.gov (United States)

Jang, Suk-Yong; Han, Sien-Ho

2016-12-19

Currently, Si as an active material for LIBs has been attracting much attention due to its high theoretical specific capacity (3572 mAh g -1 ). However, a disadvantage when using a Si negative electrode for LIBs is the abrupt drop of its capabilities during the cycling process. Therefore, there have been a few studies of polymers such as poly(vinylidene fluoride) (PVdF), carboxymethyl cellulose (CMC), styrene butadiene rubber (SBR) and polyacrylic acid (PAA) given that the robust structure of a polymeric binder to LIBs anodes is a promising means by which to enhance the performance of high-capacity anodes. These studies essentially focused mainly on modifying of the linear-polymer component or on copolymers dissolved in solvents. Cross-linking polymers as a binder may be preferred due to their good scratch resistance, excellent chemical resistance and high levels of adhesion and resilience. However, because these types of polymers (with a rigid structure and cross-linking points) are also insoluble in general organic solvents, applying these types in this capacity is virtually impossible.

Thermal/chemical degradation of ceramic cross-flow filter materials

Energy Technology Data Exchange (ETDEWEB)

Alvin, M.A.; Lane, J.E.; Lippert, T.E.

1989-11-01

This report summarizes the 14-month, Phase 1 effort conducted by Westinghouse on the Thermal/Chemical Degradation of Ceramic Cross-Flow Filter Materials program. In Phase 1 expected filter process conditions were identified for a fixed-bed, fluid-bed, and entrained-bed gasification, direct coal fired turbine, and pressurized fluidized-bed combustion system. Ceramic cross-flow filter materials were also selected, procured, and subjected to chemical and physical characterization. The stability of each of the ceramic cross-flow materials was assessed in terms of potential reactions or phase change as a result of process temperature, and effluent gas compositions containing alkali and fines. In addition chemical and physical characterization was conducted on cross-flow filters that were exposed to the METC fluid-bed gasifier and the New York University pressurized fluidized-bed combustor. Long-term high temperature degradation mechanisms were proposed for each ceramic cross-flow material at process operating conditions. An experimental bench-scale test program is recommended to be conducted in Phase 2, generating data that support the proposed cross-flow filter material thermal/chemical degradation mechanisms. Papers on the individual subtasks have been processed separately for inclusion on the data base.

Surface characteristics determining the cell compatibility of ionically cross-linked alginate gels

International Nuclear Information System (INIS)

Machida-Sano, Ikuko; Hirakawa, Makoto; Matsumoto, Hiroki; Kamada, Mitsuki; Ogawa, Sakito; Satoh, Nao; Namiki, Hideo

2014-01-01

In this study we investigated differences in the characteristics determining the suitability of five types of ion (Fe 3+ , Al 3+ , Ca 2+ , Ba 2+ and Sr 2+ )-cross-linked alginate films as culture substrates for cells. Human dermal fibroblasts were cultured on each alginate film to examine the cell affinity of the alginates. Since cell behavior on the surface of a material is dependent on the proteins adsorbed to it, we investigated the protein adsorption ability and surface features (wettability, morphology and charge) related to the protein adsorption abilities of alginate films. We observed that ferric, aluminum and barium ion-cross-linked alginate films supported better cell growth and adsorbed higher amounts of serum proteins than other types. Surface wettability analysis demonstrated that ferric and aluminum ion-cross-linked alginates had moderate hydrophilic surfaces, while other types showed highly hydrophilic surfaces. The roughness was exhibited only on barium ion-cross-linked alginate surface. Surface charge measurements revealed that alginate films had negatively charged surfaces, and showed little difference among the five types of gel. These results indicate that the critical factors of ionically cross-linked alginate films determining the protein adsorption ability required for their cell compatibility may be surface wettability and morphology. (paper)

Polymeric micelles with ionic cores containing biodegradable cross-links for delivery of chemotherapeutic agents.

Science.gov (United States)

Kim, Jong Oh; Sahay, Gaurav; Kabanov, Alexander V; Bronich, Tatiana K

2010-04-12

Novel functional polymeric nanocarriers with ionic cores containing biodegradable cross-links were developed for delivery of chemotherapeutic agents. Block ionomer complexes (BIC) of poly(ethylene oxide)-b-poly(methacylic acid) (PEO-b-PMA) and divalent metal cations (Ca(2+)) were utilized as templates. Disulfide bonds were introduced into the ionic cores by using cystamine as a biodegradable cross-linker. The resulting cross-linked micelles with disulfide bonds represented soft, hydrogel-like nanospheres and demonstrated a time-dependent degradation in the conditions mimicking the intracellular reducing environment. The ionic character of the cores allowed to achieve a very high level of doxorubicin (DOX) loading (50% w/w) into the cross-linked micelles. DOX-loaded degradable cross-linked micelles exhibited more potent cytotoxicity against human A2780 ovarian carcinoma cells as compared to micellar formulations without disulfide linkages. These novel biodegradable cross-linked micelles are expected to be attractive candidates for delivery of anticancer drugs.

Physical Characterization Of High Amylose/Pectin Mixtures Cross-Linked With Sodium Trimetaphosphate

International Nuclear Information System (INIS)

Carbinatto, F.M.; Cury, B.S.F.; Evangelista, R.C.

2010-01-01

Some researches have reported that pectin and high amylose mixtures presented superior mechanical properties in relation to those of the isolated polymers. In this work, mixtures at different ratios (1:4; 1:1) of pectin and high amylose were crosslinked with sodium trimetaphosphate at different degrees by varying reaction conditions. All samples were characterized by rheological and X-ray diffraction analyses. Samples without cross-linker were prepared as control. The oscillatory dynamic tests showed that all samples exhibited predominant elastic behavior, although cross-linked samples presented higher G' values, suggesting that crosslinking by phosphorylation resulted in more strength structures. The diffractograms showed that cross-linked samples underwent structural modifications that resulted in increase of crystallinity due to cross-linking process. (author)

Universal antibodies against the highly conserved influenza fusion peptide cross-neutralize several subtypes of influenza A virus

Energy Technology Data Exchange (ETDEWEB)

Hashem, Anwar M. [Centre for Vaccine Evaluation, Biologics and Genetic Therapies Directorate, HPFB, Health Canada, Ottawa, ON (Canada); Department of Microbiology, Faculty of Medicine, King Abdulaziz University, Jeddah (Saudi Arabia); Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON (Canada); Van Domselaar, Gary [National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg, MB (Canada); Li, Changgui; Wang, Junzhi [National Institute for the Control of Pharmaceutical and Biological Products, Beijing (China); She, Yi-Min; Cyr, Terry D. [Centre for Vaccine Evaluation, Biologics and Genetic Therapies Directorate, HPFB, Health Canada, Ottawa, ON (Canada); Sui, Jianhua [Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Department of Medicine, Harvard Medical School, 44 Binney Street, Boston, MA 02115 (United States); He, Runtao [National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg, MB (Canada); Marasco, Wayne A. [Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, Department of Medicine, Harvard Medical School, 44 Binney Street, Boston, MA 02115 (United States); Li, Xuguang, E-mail: Sean.Li@hc-sc.gc.ca [Centre for Vaccine Evaluation, Biologics and Genetic Therapies Directorate, HPFB, Health Canada, Ottawa, ON (Canada); Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ottawa, ON (Canada)

2010-12-10

Research highlights: {yields} The fusion peptide is the only universally conserved epitope in all influenza viral hemagglutinins. {yields} Anti-fusion peptide antibodies are universal antibodies that cross-react with all influenza HA subtypes. {yields} The universal antibodies cross-neutralize different influenza A subtypes. {yields} The universal antibodies inhibit the fusion process between the viruses and the target cells. -- Abstract: The fusion peptide of influenza viral hemagglutinin plays a critical role in virus entry by facilitating membrane fusion between the virus and target cells. As the fusion peptide is the only universally conserved epitope in all influenza A and B viruses, it could be an attractive target for vaccine-induced immune responses. We previously reported that antibodies targeting the first 14 amino acids of the N-terminus of the fusion peptide could bind to virtually all influenza virus strains and quantify hemagglutinins in vaccines produced in embryonated eggs. Here we demonstrate that these universal antibodies bind to the viral hemagglutinins in native conformation presented in infected mammalian cell cultures and neutralize multiple subtypes of virus by inhibiting the pH-dependant fusion of viral and cellular membranes. These results suggest that this unique, highly-conserved linear sequence in viral hemagglutinin is exposed sufficiently to be attacked by the antibodies during the course of infection and merits further investigation because of potential importance in the protection against diverse strains of influenza viruses.

Universal antibodies against the highly conserved influenza fusion peptide cross-neutralize several subtypes of influenza A virus

International Nuclear Information System (INIS)

Hashem, Anwar M.; Van Domselaar, Gary; Li, Changgui; Wang, Junzhi; She, Yi-Min; Cyr, Terry D.; Sui, Jianhua; He, Runtao; Marasco, Wayne A.; Li, Xuguang

2010-01-01

Research highlights: → The fusion peptide is the only universally conserved epitope in all influenza viral hemagglutinins. → Anti-fusion peptide antibodies are universal antibodies that cross-react with all influenza HA subtypes. → The universal antibodies cross-neutralize different influenza A subtypes. → The universal antibodies inhibit the fusion process between the viruses and the target cells. -- Abstract: The fusion peptide of influenza viral hemagglutinin plays a critical role in virus entry by facilitating membrane fusion between the virus and target cells. As the fusion peptide is the only universally conserved epitope in all influenza A and B viruses, it could be an attractive target for vaccine-induced immune responses. We previously reported that antibodies targeting the first 14 amino acids of the N-terminus of the fusion peptide could bind to virtually all influenza virus strains and quantify hemagglutinins in vaccines produced in embryonated eggs. Here we demonstrate that these universal antibodies bind to the viral hemagglutinins in native conformation presented in infected mammalian cell cultures and neutralize multiple subtypes of virus by inhibiting the pH-dependant fusion of viral and cellular membranes. These results suggest that this unique, highly-conserved linear sequence in viral hemagglutinin is exposed sufficiently to be attacked by the antibodies during the course of infection and merits further investigation because of potential importance in the protection against diverse strains of influenza viruses.

Thiol-disulfide exchange in peptides derived from human growth hormone.

Science.gov (United States)

Chandrasekhar, Saradha; Epling, Daniel E; Sophocleous, Andreas M; Topp, Elizabeth M

2014-04-01

Disulfide bonds stabilize proteins by cross-linking distant regions into a compact three-dimensional structure. They can also participate in hydrolytic and oxidative pathways to form nonnative disulfide bonds and other reactive species. Such covalent modifications can contribute to protein aggregation. Here, we present experimental data for the mechanism of thiol-disulfide exchange in tryptic peptides derived from human growth hormone in aqueous solution. Reaction kinetics was monitored to investigate the effect of pH (6.0-10.0), temperature (4-50°C), oxidation suppressants [ethylenediaminetetraacetic acid (EDTA) and N2 sparging], and peptide secondary structure (amide cyclized vs. open form). The concentrations of free thiol containing peptides, scrambled disulfides, and native disulfide-linked peptides generated via thiol-disulfide exchange and oxidation reactions were determined using reverse-phase HPLC and liquid chromatography-mass spectrometry. Concentration versus time data were fitted to a mathematical model using nonlinear least squares regression analysis. At all pH values, the model was able to fit the data with R(2) ≥ 0.95. Excluding oxidation suppressants (EDTA and N2 sparging) resulted in an increase in the formation of scrambled disulfides via oxidative pathways but did not influence the intrinsic rate of thiol-disulfide exchange. In addition, peptide secondary structure was found to influence the rate of thiol-disulfide exchange. © 2014 Wiley Periodicals, Inc. and the American Pharmacists Association.

CrossLink: a novel method for cross-condition classification of cancer subtypes.

Science.gov (United States)

Ma, Chifeng; Sastry, Konduru S; Flore, Mario; Gehani, Salah; Al-Bozom, Issam; Feng, Yusheng; Serpedin, Erchin; Chouchane, Lotfi; Chen, Yidong; Huang, Yufei

2016-08-22

We considered the prediction of cancer classes (e.g. subtypes) using patient gene expression profiles that contain both systematic and condition-specific biases when compared with the training reference dataset. The conventional normalization-based approaches cannot guarantee that the gene signatures in the reference and prediction datasets always have the same distribution for all different conditions as the class-specific gene signatures change with the condition. Therefore, the trained classifier would work well under one condition but not under another. To address the problem of current normalization approaches, we propose a novel algorithm called CrossLink (CL). CL recognizes that there is no universal, condition-independent normalization mapping of signatures. In contrast, it exploits the fact that the signature is unique to its associated class under any condition and thus employs an unsupervised clustering algorithm to discover this unique signature. We assessed the performance of CL for cross-condition predictions of PAM50 subtypes of breast cancer by using a simulated dataset modeled after TCGA BRCA tumor samples with a cross-validation scheme, and datasets with known and unknown PAM50 classification. CL achieved prediction accuracy >73 %, highest among other methods we evaluated. We also applied the algorithm to a set of breast cancer tumors derived from Arabic population to assign a PAM50 classification to each tumor based on their gene expression profiles. A novel algorithm CrossLink for cross-condition prediction of cancer classes was proposed. In all test datasets, CL showed robust and consistent improvement in prediction performance over other state-of-the-art normalization and classification algorithms.

Physical and mechanical properties of gamma radiation cross-linked polyethylene

International Nuclear Information System (INIS)

Gonzalez, Maria E.; Romero, G.; Smolko, Eduardo E.

1999-01-01

Granulated LDPE 2003 polyethylene was extruded and irradiated under nitrogen with 150, 200 and 300 kGy gamma rays doses to produce cross-linking. The study of the physical and mechanical properties shows that the product has a high degree of molecular cross-linking, can be heated up to 200 C for 2 hours without deformation and that the mechanical properties improve. Preliminary aging tests indicate that after heating at 60 C for 4 weeks no physical or mechanical deterioration can be observed. (author)

Covalent-ionically cross-linked polyetheretherketone proton exchange membrane for direct methanol fuel cell

CSIR Research Space (South Africa)

Luo, H

2010-08-01

Full Text Available cross-linked PEEK-WC membrane, this covalent-ionically cross-linked PEEK-WC membrane exhibits extremely reduced water uptake and methanol permeability, but just slightly sacrificed proton conductivity. The proton conductivity of the covalent...

Dually cross-linked single network poly(acrylic acid) hydrogels with superior mechanical properties and water absorbency.

Science.gov (United States)

Zhong, Ming; Liu, Yi-Tao; Liu, Xiao-Ying; Shi, Fu-Kuan; Zhang, Li-Qin; Zhu, Mei-Fang; Xie, Xu-Ming

2016-06-28

Poly(acrylic acid) (PAA) hydrogels with superior mechanical properties, based on a single network structure with dual cross-linking, are prepared by one-pot free radical polymerization. The network structure of the PAA hydrogels is composed of dual cross-linking: a dynamic and reversible ionic cross-linking among the PAA chains enabled by Fe(3+) ions, and a sparse covalent cross-linking enabled by a covalent cross-linker (Bis). Under deformation, the covalently cross-linked PAA chains remain intact to maintain their original configuration, while the Fe(3+)-enabled ionic cross-linking among the PAA chains is broken to dissipate energy and then recombined. It is found that the mechanical properties of the PAA hydrogels are significantly influenced by the contents of covalent cross-linkers, Fe(3+) ions and water, which can be adjusted within a substantial range and thus broaden the applications of the hydrogels. Meanwhile, the PAA hydrogels have excellent recoverability based on the dynamic and reversible ionic cross-linking enabled by Fe(3+) ions. Moreover, the swelling capacity of the PAA hydrogels is as high as 1800 times in deionized water due to the synergistic effects of ionic and covalent cross-linkings. The combination of balanced mechanical properties, efficient recoverability, high swelling capacity and facile preparation provides a new method to obtain high-performance hydrogels.

Cross-linked compared with historical polyethylene in THA: an 8-year clinical study.

Science.gov (United States)

Geerdink, Carel H; Grimm, Bernd; Vencken, Wendy; Heyligers, Ide C; Tonino, Alphons J

2009-04-01

Wear particle-induced osteolysis is a major cause of aseptic loosening in THA. Increasing wear resistance of polyethylene (PE) occurs by increasing the cross-link density and early reports document low wear rates with such implants. To confirm longer-term reductions in wear we compared cross-linked polyethylene (irradiation in nitrogen, annealing) with historical polyethylene (irradiation in air) in a prospective, randomized clinical study involving 48 patients who underwent THAs with a minimum followup of 7 years (mean, 8 years; range, 7-9 years). The insert material was the only variable. The Harris hip score, radiographic signs of osteolysis, and polyethylene wear were recorded annually. Twenty-three historical and 17 moderately cross-linked polyethylene inserts were analyzed (five patients died, three were lost to followup). At 8 years, the wear rate was lower for cross-linked polyethylene (0.088 +/- 0.03 mm/year) than for the historical polyethylene (0.142 +/- 0.07 mm/year). This reduction (38%) did not diminish with time (33% at 5 years). Acetabular cyst formation was less frequent (39% versus 12%), affected fewer DeLee and Charnley zones (17% versus 4%), and was less severe for the cross-linked polyethylene. The only revision was for an aseptically loose cup in the historical polyethylene group. Moderately cross-linked polyethylene maintained its wear advantage with time and produced less osteolysis, showing no signs of aging at mid-term followup. Level I, therapeutic study. See Guidelines for Authors for a complete description of levels of evidence.

Multiple repair pathways mediate tolerance to chemotherapeutic cross-linking agents in vertebrate cells.

Science.gov (United States)

Nojima, Kuniharu; Hochegger, Helfrid; Saberi, Alihossein; Fukushima, Toru; Kikuchi, Koji; Yoshimura, Michio; Orelli, Brian J; Bishop, Douglas K; Hirano, Seiki; Ohzeki, Mioko; Ishiai, Masamichi; Yamamoto, Kazuhiko; Takata, Minoru; Arakawa, Hiroshi; Buerstedde, Jean-Marie; Yamazoe, Mitsuyoshi; Kawamoto, Takuo; Araki, Kasumi; Takahashi, Jun A; Hashimoto, Nobuo; Takeda, Shunichi; Sonoda, Eiichiro

2005-12-15

Cross-linking agents that induce DNA interstrand cross-links (ICL) are widely used in anticancer chemotherapy. Yeast genetic studies show that nucleotide excision repair (NER), Rad6/Rad18-dependent postreplication repair, homologous recombination, and cell cycle checkpoint pathway are involved in ICL repair. To study the contribution of DNA damage response pathways in tolerance to cross-linking agents in vertebrates, we made a panel of gene-disrupted clones from chicken DT40 cells, each defective in a particular DNA repair or checkpoint pathway, and measured the sensitivities to cross-linking agents, including cis-diamminedichloroplatinum (II) (cisplatin), mitomycin C, and melphalan. We found that cells harboring defects in translesion DNA synthesis (TLS), Fanconi anemia complementation groups (FANC), or homologous recombination displayed marked hypersensitivity to all the cross-linking agents, whereas NER seemed to play only a minor role. This effect of replication-dependent repair pathways is distinctively different from the situation in yeast, where NER seems to play a major role in dealing with ICL. Cells deficient in Rev3, the catalytic subunit of TLS polymerase Polzeta, showed the highest sensitivity to cisplatin followed by fanc-c. Furthermore, epistasis analysis revealed that these two mutants work in the same pathway. Our genetic comprehensive study reveals a critical role for DNA repair pathways that release DNA replication block at ICLs in cellular tolerance to cross-linking agents and could be directly exploited in designing an effective chemotherapy.

Impact of styrenic polymer one-step hyper-cross-linking on volatile organic compound adsorption and desorption performance.

Science.gov (United States)

Ghafari, Mohsen; Atkinson, John D

2018-06-05

A novel one-step hyper-cross-linking method, using 1,2-dichloroethane (DCE) and 1,6-dichlorohexane (DCH) cross-linkers, expands the micropore volume of commercial styrenic polymers. Performance of virgin and modified polymers was evaluated by measuring hexane, toluene, and methyl-ethyl-ketone (MEK) adsorption capacity, adsorption/desorption kinetics, and desorption efficiency. Hyper-cross-linked polymers have up to 128% higher adsorption capacity than virgin polymers at P/P 0  = 0.05 due to micropore volume increases up to 330%. Improvements are most pronounced with the DCE cross-linker. Hyper-cross-linking has minimal impact on hexane adsorption kinetics, but adsorption rates for toluene and MEK decrease by 6-41%. Desorption rates decreased (3-36%) for all materials after hyper-cross-linking, with larger decreases for DCE hyper-cross-linked polymers due to smaller average pore widths. For room temperature desorption, 20-220% more adsorbate remains in hyper-cross-linked polymers after regeneration compared to virgin materials. DCE hyper-cross-linked polymers have 13-92% more residual adsorbate than DCH counterparts. Higher temperatures were required for DCE hyper-cross-linked polymers to completely desorb VOCs compared to the DCH hyper-cross-linked and virgin counterparts. Results show that the one-step hyper-cross-linking method for modifying styrenic polymers improves adsorption capacity because of added micropores, but decreases adsorption/desorption kinetics and desorption efficiency for large VOCs due to a decrease in average pore width. Copyright © 2018 Elsevier B.V. All rights reserved.

The mechanism of collagen cross-linking in diabetes: a puzzle nearing resolution.

Science.gov (United States)

Monnier, V M; Glomb, M; Elgawish, A; Sell, D R

1996-07-01

Considerable interest has been focused in recent years on the mechanism of collagen cross-linking by high glucose in vitro and in vivo. Experiments in both diabetic humans and in animals have shown that over time collagen becomes less soluble, less digestible by collagenase, more stable to heat-induced denaturation, and more glycated. In addition, collagen becomes more modified by advanced products of the Maillard reaction, i.e., immunoreactive advanced glycation end products and the glycoxidation markers carboxymethyllysine and pentosidine. Mechanistic studies have shown that collagen cross-linking in vitro can be uncoupled from glycation by the use of antioxidants and chelating agents. Experiments in the authors' laboratory revealed that approximately 50% of carboxymethyllysine formed in vitro originates from pathways other than oxidation of Amadori products, i.e., most likely the oxidation of Schiff base-linked glucose. In addition, the increase in thermal stability of rat tail tendons exposed to high glucose in vitro or in vivo was found to strongly depend on H2O2 formation. The final missing piece of the puzzle is that of the structure of the major cross-link. We speculate that it is a nonfluorescent nonultraviolet active cross-link between two lysine residues, which includes a fragmentation product of glucose linked in a nonreducible bond labile to both strong acids and bases.

Interstrand cross-links arising from strand breaks at true abasic sites in duplex DNA

Science.gov (United States)

Yang, Zhiyu; Price, Nathan E.; Johnson, Kevin M.

2017-01-01

Abstract Interstrand cross-links are exceptionally bioactive DNA lesions. Endogenous generation of interstrand cross-links in genomic DNA may contribute to aging, neurodegeneration, and cancer. Abasic (Ap) sites are common lesions in genomic DNA that readily undergo spontaneous and amine-catalyzed strand cleavage reactions that generate a 2,3-didehydro-2,3-dideoxyribose sugar remnant (3’ddR5p) at the 3’-terminus of the strand break. Interestingly, this strand scission process leaves an electrophilic α,β-unsaturated aldehyde residue embedded within the resulting nicked duplex. Here we present evidence that 3’ddR5p derivatives generated by spermine-catalyzed strand cleavage at Ap sites in duplex DNA can react with adenine residues on the opposing strand to generate a complex lesion consisting of an interstrand cross-link adjacent to a strand break. The cross-link blocks DNA replication by ϕ29 DNA polymerase, a highly processive polymerase enzyme that couples synthesis with strand displacement. This suggests that 3’ddR5p-derived cross-links have the potential to block critical cellular DNA transactions that require strand separation. LC-MS/MS methods developed herein provide powerful tools for studying the occurrence and properties of these cross-links in biochemical and biological systems. PMID:28531327

Peptide Based Targeted Therapeutic Radiopharmaceuticals: A Focus on the Synthesis of Radiolabelled Nanobodies

International Nuclear Information System (INIS)

Impens, N.; Campsteyn, A.; Aerts, A.; Baatout, S.; Devoogdt, N.; Caveliers, V.; Xavier, C.; Lahoutte, T.

2009-01-01

In 1993, the Vrije Universiteit Brussel (Brussels, Belgium) discovered in the blood of camelidae antibodies consisting of only a heavy chain. Due to the lack of the light chain only the variable part of the heavy chain is important for antigen binding. This variable part of these heavy-chain-only antibodies is a good candidate as a targeted therapeutic radiopharmaceutical and was called a nanobody, having a molecular weight of about 15 kDa. Its dimensions are included in between the small peptides like derived from e.g. somatostatin, and the classical monoclonal antibodies. This makes that some characteristics like the physical behaviour, the chemical stability, the penetration in tumour and in healthy tissues, and the blood clearance lie in between the characteristics of the small peptides and the monoclonal antibodies, therefore taking advantage of both extremes. Nanobodies have been humanised to decrease the immunogenic response. The building blocks of molecules such as the octreotide, nanobodies and monoclonal antibodies are amino acids linked via peptide bonds. The modification reactions are therefore all based on the same 'peptide chemistry'. The functional groups on the present amino acids will determine the possible reactions. In order to link a radionuclide to the nanobodies, we opted to use bifunctional ligands containing DOTA, because this is a suitable chelating agent for the diagnostic radionuclide Ga-68, and for therapeutic radionuclides such as Lu-177 and Y-90, covering short and long range β-particle emitters suitable for attacking a wide range of tumour sizes. The ratio of bifunctional ligand to nanobody can be varied by carefully selecting the functional groups of the peptide involved in the reaction with the bifunctional ligand, avoiding the complementarity determining region (CDR), i.e. the part of the molecule binding to the antigen. This is a first way to predetermine the amount of radionuclides that can be linked to the peptide, or the

Controlled swollen and drug release from urea-cross-linked polyether/siloxane hybrids

International Nuclear Information System (INIS)

Santilli, Celso V.; Lopes, Leandro; Pulcinelli, Sandra H.; Chiavacci, Leila A.; Oliveira, Anselmo G.

2009-01-01

From a simple synthesis method we produced transparent ureasil cross-linked polyether (poly(ethylene oxide), PEO, or poly (propylene oxide), PPO) networks, whose designed inter cross-linking distance and tunable swell ability was assessed by small angle X-ray scattering on the D11A-SAXS1 beamline of the LNLS, we demonstrated that the controlled drug release from swell able hydrophilic ureasil-PEO materials can be sustained during some days, while from the unswell able ureasil-PPO ones, during some weeks. This outstanding feature conjugated with the bio medically safe formulation of the ureasil cross-linked polyether/siloxane hybrid widen their scope of application to include the domain of soft and implantable drug delivery devices. (author)

Synthesis and Characterization of Chemically Cross-Linked Acrylic Acid/Gelatin Hydrogels: Effect of pH and Composition on Swelling and Drug Release

Directory of Open Access Journals (Sweden)

Syed Majid Hanif Bukhari

2015-01-01

Full Text Available This present work was aimed at synthesizing pH-sensitive cross-linked AA/Gelatin hydrogels by free radical polymerization. Ammonium persulfate and ethylene glycol dimethacrylate (EGDMA were used as initiator and as cross-linking agent, respectively. Different feed ratios of acrylic acid, gelatin, and EGDMA were used to investigate the effect of monomer, polymer, and degree of cross-linking on swelling and release pattern of the model drug. The swelling behavior of the hydrogel samples was studied in 0.05 M USP phosphate buffer solutions of various pH values pH 1.2, pH 5.5, pH 6.5, and pH 7.5. The prepared samples were evaluated for porosity and sol-gel fraction analysis. Pheniramine maleate used for allergy treatment was loaded as model drug in selected samples. The release study of the drug was investigated in 0.05 M USP phosphate buffer of varying pH values (1.2, 5.5, and 7.5 for 12 hrs. The release data was fitted to various kinetic models to study the release mechanism. Hydrogels were characterized by Fourier transformed infrared (FTIR spectroscopy which confirmed formation of structure. Surface morphology of unloaded and loaded samples was studied by surface electron microscopy (SEM, which confirmed the distribution of model drug in the gel network.

CD24 cross-linking induces apoptosis in, and inhibits migration of, MCF-7 breast cancer cells

International Nuclear Information System (INIS)

Kim, Jong Bin; Bae, Ji-Yeon; Jee, Hyeon-Gun; Noh, Dong-Young; Ko, Eunyoung; Han, Wonshik; Lee, Jeong Eon; Lee, Kyung-Min; Shin, Incheol; Kim, Sangmin; Lee, Jong Won; Cho, Jihyoung

2008-01-01

The biological effects of CD24 (FL-80) cross-linking on breast cancer cells have not yet been established. We examined the impact of CD24 cross-linking on human breast cancer cell line MCF-7. MCF-7 and MDA-MB-231 cells were treated with anti-rabbit polyclonal IgG or anti-human CD24 rabbit polyclonal antibodies to induce cross-linking, and then growth was studied. Changes in cell characteristics such as cell cycle modulation, cell death, survival in three-dimensional cultures, adhesion, and migration ability were assayed after CD24 cross-linking in MCF-7. Expression of CD24 was analyzed by flow cytometry in MDA-MB-231 and MCF-7 cells where 2% and 66% expression frequencies were observed, respectively. CD24 cross-linking resulted in time-dependent proliferation reduction in MCF-7 cells, but no reduction in MDA-MB-231 cells. MCF-7 cell survival was reduced by 15% in three-dimensional culture after CD24 cross-linking. Increased MCF-7 cell apoptosis was observed after CD24 cross-linking, but no cell cycle arrest was observed in that condition. The migration capacity of MCF-7 cells was diminished by 30% after CD24 cross-linking. Our results showed that CD24 cross-linking induced apoptosis and inhibited migration in MCF-7 breast cancer cells. We conclude that CD24 may be considered as a novel therapeutic target for breast cancer

The A-chain of insulin contacts the insert domain of the insulin receptor. Photo-cross-linking and mutagenesis of a diabetes-related crevice.

Science.gov (United States)

Huang, Kun; Chan, Shu Jin; Hua, Qing-xin; Chu, Ying-Chi; Wang, Run-ying; Klaproth, Birgit; Jia, Wenhua; Whittaker, Jonathan; De Meyts, Pierre; Nakagawa, Satoe H; Steiner, Donald F; Katsoyannis, Panayotis G; Weiss, Michael A

2007-11-30

The contribution of the insulin A-chain to receptor binding is investigated by photo-cross-linking and nonstandard mutagenesis. Studies focus on the role of Val(A3), which projects within a crevice between the A- and B-chains. Engineered receptor alpha-subunits containing specific protease sites ("midi-receptors") are employed to map the site of photo-cross-linking by an analog containing a photoactivable A3 side chain (para-azido-Phe (Pap)). The probe cross-links to a C-terminal peptide (residues 703-719 of the receptor A isoform, KTFEDYLHNVVFVPRPS) containing side chains critical for hormone binding (underlined); the corresponding segment of the holoreceptor was shown previously to cross-link to a Pap(B25)-insulin analog. Because Pap is larger than Val and so may protrude beyond the A3-associated crevice, we investigated analogs containing A3 substitutions comparable in size to Val as follows: Thr, allo-Thr, and alpha-aminobutyric acid (Aba). Substitutions were introduced within an engineered monomer. Whereas previous studies of smaller substitutions (Gly(A3) and Ser(A3)) encountered nonlocal conformational perturbations, NMR structures of the present analogs are similar to wild-type insulin; the variant side chains are accommodated within a native-like crevice with minimal distortion. Receptor binding activities of Aba(A3) and allo-Thr(A3) analogs are reduced at least 10-fold; the activity of Thr(A3)-DKP-insulin is reduced 5-fold. The hormone-receptor interface is presumably destabilized either by a packing defect (Aba(A3)) or by altered polarity (allo-Thr(A3) and Thr(A3)). Our results provide evidence that Val(A3), a site of mutation causing diabetes mellitus, contacts the insert domain-derived tail of the alpha-subunit in a hormone-receptor complex.

Jumping Hurdles: Peptides Able To Overcome Biological Barriers.

Science.gov (United States)

Sánchez-Navarro, Macarena; Teixidó, Meritxell; Giralt, Ernest

2017-08-15

The cell membrane, the gastrointestinal tract, and the blood-brain barrier (BBB) are good examples of biological barriers that define and protect cells and organs. They impose different levels of restriction, but they also share common features. For instance, they all display a high lipophilic character. For this reason, hydrophilic compounds, like peptides, proteins, or nucleic acids have long been considered as unable to bypass them. However, the discovery of cell-penetrating peptides (CPPs) opened a vast field of research. Nowadays, CPPs, homing peptides, and blood-brain barrier peptide shuttles (BBB-shuttles) are good examples of peptides able to target and to cross various biological barriers. CPPs are a group of peptides able to interact with the plasma membrane and enter the cell. They display some common characteristics like positively charged residues, mainly arginines, and amphipathicity. In this field, our group has been focused on the development of proline rich CPPs and in the analysis of the importance of secondary amphipathicity in the internalization process. Proline has a privileged structure being the only amino acid with a secondary amine and a cyclic side chain. These features constrain its structure and hamper the formation of H-bonds. Taking advantage of this privileged structure, three different families of proline-rich peptides have been developed, namely, a proline-rich dendrimer, the sweet arrow peptide (SAP), and a group of foldamers based on γ-peptides. The structure and the mechanism of internalization of all of them has been evaluated and analyzed. BBB-shuttles are peptides able to cross the BBB and to carry with them compounds that cannot reach the brain parenchyma unaided. These peptides take advantage of the natural transport mechanisms present at the BBB, which are divided in active and passive transport mechanisms. On the one hand, we have developed BBB-shuttles that cross the BBB by a passive transport mechanism, like

Swelling of cross-linked polymers: interpretations and misinterpretations

Czech Academy of Sciences Publication Activity Database

Dušek, Karel; Dušková-Smrčková, Miroslava

2017-01-01

Roč. 254, 20 August (2017), s. 102 ISSN 0065-7727. [ACS National Meeting & Exposition /254./. 20.08.2017-24.08.2017, Washington] Institutional support: RVO:61389013 Keywords : swelling * cross-linked polymer Subject RIV: CD - Macromolecular Chemistry OBOR OECD: Polymer science

Cross Country MetroLink Segment I Business Plan

Science.gov (United States)

1997-12-02

In the St. Louis, Missouri metropolitan area, the East-West Gateway Coordinating : Council decided the route for the first MetroLink extension in the Cross-County : Corridor in September 1997. The next phase, reflected in this paper is develop, : dur...

Hyper-cross-linked, hybrid membranes via interfacial polymerization

NARCIS (Netherlands)

Raaijmakers, Michiel

2015-01-01

Hyper-cross-linked, hybrid membranes consist of covalent networks of alternating organic and inorganic, or biological groups. This thesis reports on the preparation of such hybrid networks via interfacial polymerization. The structure-property relationships of the hybrid networks depend strongly on

Bypass of a psoralen DNA interstrand cross-link by DNA polymerases beta, iota, and kappa in vitro

Science.gov (United States)

Smith, Leigh A.; Makarova, Alena V.; Samson, Laura; Thiesen, Katherine E.; Dhar, Alok; Bessho, Tadayoshi

2012-01-01

Repair of DNA inter-strand cross-links in mammalian cells involves several biochemically distinctive processes, including the release of one of the cross-linked strands and translesion DNA synthesis (TLS). In this report, we investigated in vitro TLS activity of psoralen DNA inter-strand cross-link by three DNA repair polymerases, DNA polymerase beta, kappa and iota. DNA polymerase beta is capable of bypassing a psoralen cross-link with a low efficiency. Cell extracts prepared from DNA polymerase beta knockout mouse embryonic fibroblast showed a reduced bypass activity of the psoralen cross-link and purified DNA polymerase beta restored the bypass activity. In addition, DNA polymerase iota mis-incorporated thymine across the psoralen cross-link and DNA polymerase kappa extended these mis-paired primer ends, suggesting that DNA polymerase iota may serve as an inserter and DNA polymerase kappa may play a role as an extender in the repair of psoralen DNA inter-strand cross-links. The results demonstrated here indicate that multiple DNA polymerases could participate in TLS steps in mammalian DNA inter-strand cross-link repair. PMID:23106263

Analysis of protein-nucleic acid interactions by photochemical cross-linking and mass spectrometry

DEFF Research Database (Denmark)

Steen, Hanno; Jensen, Ole Nørregaard

2002-01-01

. Mass spectrometry (MS) has emerged as a sensitive and efficient analytical technique for determination of such cross-linking sites in proteins. The present review of the field describes a number of MS-based approaches for the characterization of cross-linked protein-nucleic acid complexes...

Automated solid-phase peptide synthesis to obtain therapeutic peptides

Directory of Open Access Journals (Sweden)

Veronika Mäde

2014-05-01

Full Text Available The great versatility and the inherent high affinities of peptides for their respective targets have led to tremendous progress for therapeutic applications in the last years. In order to increase the drugability of these frequently unstable and rapidly cleared molecules, chemical modifications are of great interest. Automated solid-phase peptide synthesis (SPPS offers a suitable technology to produce chemically engineered peptides. This review concentrates on the application of SPPS by Fmoc/t-Bu protecting-group strategy, which is most commonly used. Critical issues and suggestions for the synthesis are covered. The development of automated methods from conventional to essentially improved microwave-assisted instruments is discussed. In order to improve pharmacokinetic properties of peptides, lipidation and PEGylation are described as covalent conjugation methods, which can be applied by a combination of automated and manual synthesis approaches. The synthesis and application of SPPS is described for neuropeptide Y receptor analogs as an example for bioactive hormones. The applied strategies represent innovative and potent methods for the development of novel peptide drug candidates that can be manufactured with optimized automated synthesis technologies.

Nanomechanics of layer-by-layer polyelectrolyte complexes: a manifestation of ionic cross-links and fixed charges.

Science.gov (United States)

Han, Biao; Chery, Daphney R; Yin, Jie; Lu, X Lucas; Lee, Daeyeon; Han, Lin

2016-01-28

This study investigates the roles of two distinct features of ionically cross-linked polyelectrolyte networks - ionic cross-links and fixed charges - in determining their nanomechanical properties. The layer-by-layer assembled poly(allylamine hydrochloride)/poly(acrylic acid) (PAH/PAA) network is used as the model material. The densities of ionic cross-links and fixed charges are modulated through solution pH and ionic strength (IS), and the swelling ratio, elastic and viscoelastic properties are quantified via an array of atomic force microscopy (AFM)-based nanomechanical tools. The roles of ionic cross-links are underscored by the distinctive elastic and viscoelastic nanomechanical characters observed here. First, as ionic cross-links are highly sensitive to solution conditions, the instantaneous modulus, E0, exhibits orders-of-magnitude changes upon pH- and IS-governed swelling, distinctive from the rubber elasticity prediction based on permanent covalent cross-links. Second, ionic cross-links can break and self-re-form, and this mechanism dominates force relaxation of PAH/PAA under a constant indentation depth. In most states, the degree of relaxation is >90%, independent of ionic cross-link density. The importance of fixed charges is highlighted by the unexpectedly more elastic nature of the network despite low ionic cross-link density at pH 2.0, IS 0.01 M. Here, the complex is a net charged, loosely cross-linked, where the degree of relaxation is attenuated to ≈50% due to increased elastic contribution arising from fixed charge-induced Donnan osmotic pressure. In addition, this study develops a new method for quantifying the thickness of highly swollen polymer hydrogel films. It also underscores important technical considerations when performing nanomechanical tests on highly rate-dependent polymer hydrogel networks. These results provide new insights into the nanomechanical characters of ionic polyelectrolyte complexes, and lay the ground for further

Whey protein isolate with improved film properties through cross-linking catalyzed by small laccase from Streptomyces coelicolor.

Science.gov (United States)

Quan, Wei; Zhang, Chong; Zheng, Meixia; Lu, Zhaoxin; Lu, Fengxia

2018-08-01

The effects of small laccase (SLAC) from Streptomyces coelicolor on the properties of whey protein isolate (WPI) films were studied. WPI was catalyze by SLAC without phenolic acid assistance. Particle size distribution results showed that some complexes with higher relative molecular weight formed in WPI samples treated with SLAC. The content of α-helixes decreased while those of β-sheets and random coils increased following SLAC treatment according to circular dichroism results. Fourier transform infrared spectral analysis suggested that some conformational changes occurred in WPI following SLAC treatment. Analysis of WPI films prepared by casting after SLAC treatment indicated that their film properties were all improved, including mechanical properties, solubility, water vapor, oxygen and carbon dioxide barrier properties, film color, light transmission, transparency and thermal properties. Compared with that of the control film, some obvious differences in the morphology of the WPI films were observed following SLAC treatment. This report demonstrates that laccase can directly catalyze protein cross-linking, which may be useful to improve the performance of protein films. In this study, SLAC was applied to WPI edible film during the film-making process. The results showed that SLAC can catalyze WPI cross-linking without phenolic acid assistance, and WPI film properties were improved after SLAC treatment. © 2018 Society of Chemical Industry. © 2018 Society of Chemical Industry.

Vitamin E-diffused highly cross-linked UHMWPE particles induce less osteolysis compared to highly cross-linked virgin UHMWPE particles in vivo

DEFF Research Database (Denmark)

Bichara, David A; Malchau, Erik; Sillesen, Nanna H

2014-01-01

when compared to virgin gamma irradiated cross-linked UHMWPE. Groups received equal amount of particulate debris overlaying the calvarium for 10 days. Calvarial bone was examined using high resolution micro-CT and histomorphometric analyses. There was a statistically significant difference between...

Molecular contacts for chlorosome envelope proteins revealed by cross-linking studies with chlorosomes from Chlorobium tepidum

DEFF Research Database (Denmark)

Li, Hui; Frigaard, Niels-Ulrik; Bryant, Donald A

2006-01-01

type and mutants lacking a single chlorosome protein were cross-linked with the zero-length cross-linker 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide (EDC) and analyzed by gel electrophoresis. Similar cross-linking products were observed when the time and temperature were varied or when EDC...... was replaced with glutaraldehyde. Specific interactions between chlorosome proteins in cross-linked products were identified by immunoblotting with polyclonal antibodies raised against recombinant chlorosome proteins. We confirmed these interactions by demonstrating that these products were missing...... in appropriate mutants. Confirming the location of CsmA in the paracrystalline baseplate, cross-linking showed that CsmA forms dimers, trimers, and homomultimers as large as dodecamers and that CsmA directly interacts with the Fenna-Matthews-Olson protein. Cross-linking further suggests that the precursor form...

Improvement on Physical Properties of Pullulan Films by Novel Cross-Linking Strategy.

Science.gov (United States)

Chen, Chieh-Ting; Chen, Kuan-I; Chiang, Hsin-Han; Chen, Yu-Kuo; Cheng, Kuan-Chen

2017-01-01

Pullulan based films possess several advantages, including high transparency, low toxicity, good biodegradability, good mechanical properties, and low oxygen permeability, are preferable for food packaging. The application of pullulan films on food packaging, however, has inherent disadvantage of high water solubility. In this study, glutaraldehyde and glycerol were used as the cross-linking reagent and the plasticizer respectively to improve water resistance and physical properties of the pullulan films. Effects of cross-linking degree on physical properties, including water absorptions, swelling behaviors, water vapor permeability and tensile strengths of films were evaluated. FTIR results demonstrated that the pullulan films were successfully cross-linked by glutaraldehyde. The tensile strength of pullulan films could be enhanced significantly (P permeability. © 2016 Institute of Food Technologists®.

The theory and art of corneal cross-linking.

Science.gov (United States)

McQuaid, Rebecca; Cummings, Arthur B; Mrochen, Michael

2013-08-01

Before the discovery of corneal cross-linking (CXL), patients with keratoconus would have had to undergo corneal transplantation, or wear rigid gas permeable lenses (RGPs) that would temporarily flatten the cone, thereby improving the vision. The RGP contact lens (CL) would not however alter the corneal stability and if the keratoconus was progressive, the continued steepening of the cone would occur under the RGP CL. To date, the Siena Eye has been the largest study to investigate long term effects of standard CXL. Three hundred and sixty-three eyes were treated and monitored over 4 years, producing reliable long-term results proving long-term stability of the cornea by halting the progression of keratoconus, and proving the safety of the procedure. Traditionally, CXL requires epithelial removal prior to corneal soakage of a dextran-based 0.1% riboflavin solution, followed by exposure of ultraviolet-A (UV-A) light for 30 min with an intensity of 3 mW/cm2. A series of in vitro investigations on human and porcine corneas examined the best treatment parameters for standard CXL, such as riboflavin concentration, intensity, wavelength of UV-A light, and duration of treatment. Photochemically, CXL is achieved by the generation of chemical bonds within the corneal stroma through localized photopolymerization, strengthening the cornea whilst minimizing exposure to the surrounding structures of the eye. In vitro studies have shown that CXL has an effect on the biomechanical properties of the cornea, with an increased corneal rigidity of approximately 70%. This is a result of the creation of new chemical bonds within the stroma.

Interstrand cross-links arising from strand breaks at true abasic sites in duplex DNA.

Science.gov (United States)

Yang, Zhiyu; Price, Nathan E; Johnson, Kevin M; Wang, Yinsheng; Gates, Kent S

2017-06-20

Interstrand cross-links are exceptionally bioactive DNA lesions. Endogenous generation of interstrand cross-links in genomic DNA may contribute to aging, neurodegeneration, and cancer. Abasic (Ap) sites are common lesions in genomic DNA that readily undergo spontaneous and amine-catalyzed strand cleavage reactions that generate a 2,3-didehydro-2,3-dideoxyribose sugar remnant (3'ddR5p) at the 3'-terminus of the strand break. Interestingly, this strand scission process leaves an electrophilic α,β-unsaturated aldehyde residue embedded within the resulting nicked duplex. Here we present evidence that 3'ddR5p derivatives generated by spermine-catalyzed strand cleavage at Ap sites in duplex DNA can react with adenine residues on the opposing strand to generate a complex lesion consisting of an interstrand cross-link adjacent to a strand break. The cross-link blocks DNA replication by ϕ29 DNA polymerase, a highly processive polymerase enzyme that couples synthesis with strand displacement. This suggests that 3'ddR5p-derived cross-links have the potential to block critical cellular DNA transactions that require strand separation. LC-MS/MS methods developed herein provide powerful tools for studying the occurrence and properties of these cross-links in biochemical and biological systems. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Structural and Functional Characterization of an Ancient Bacterial Transglutaminase Sheds Light on the Minimal Requirements for Protein Cross-Linking.

Science.gov (United States)

Fernandes, Catarina G; Plácido, Diana; Lousa, Diana; Brito, José A; Isidro, Anabela; Soares, Cláudio M; Pohl, Jan; Carrondo, Maria A; Archer, Margarida; Henriques, Adriano O

2015-09-22

Transglutaminases are best known for their ability to catalyze protein cross-linking reactions that impart chemical and physical resilience to cellular structures. Here, we report the crystal structure and characterization of Tgl, a transglutaminase from the bacterium Bacillus subtilis. Tgl is produced during sporulation and cross-links the surface of the highly resilient spore. Tgl-like proteins are found only in spore-forming bacteria of the Bacillus and Clostridia classes, indicating an ancient origin. Tgl is a single-domain protein, produced in active form, and the smallest transglutaminase characterized to date. We show that Tgl is structurally similar to bacterial cell wall endopeptidases and has an NlpC/P60 catalytic core, thought to represent the ancestral unit of the cysteine protease fold. We show that Tgl functions through a unique partially redundant catalytic dyad formed by Cys116 and Glu187 or Glu115. Strikingly, the catalytic Cys is insulated within a hydrophobic tunnel that traverses the molecule from side to side. The lack of similarity of Tgl to other transglutaminases together with its small size suggests that an NlpC/P60 catalytic core and insulation of the active site during catalysis may be essential requirements for protein cross-linking.

Cross-linked polyethylene does not reduce wear in total knee arthroplasty.

Science.gov (United States)

Lasurt-Bachs, S; Torner, P; Maculé, F; Prats, E; Menéndez-García, F; Ríos-Guillermo, J; Torrents, A

To compare two different types of inserts: Ultra-high molecular weight polyethylene (UHMWPE) and cross-linked polyethylene with a quantitative and qualitative study of polyethylene wear particles in synovial fluid 3 years after total knee arthroplasty. A prospective, randomized, controlled cohort study with blinded evaluation was carried out on 25 patients undergoing staged bilateral total knee replacement, 6 months apart. Knee arthrocentesis was performed on 12 patients 3 years after surgery, and the polyethylene particles were analyzed. No significant differences were found in the number of particles generated by the two different types of inserts at 3 years from total knee arthroplasty (3,000×: x¯ cross-linked=849.7; x¯ UHMWPE=796.9; P=.63; 20,000×: x¯ cross-linked=66.3; x¯ UHMWPE=73.1; P=.76). Likewise, no differences in the probability of finding elongated (χ 2 =0.19; P=.66) or rounded (χ 2 =1.44; P=.23) particles in both types of inserts were observed. However, the probability of finding fibrillar particles is 3.08 times greater in UHMWPE. Cross-linked polyethylene does not significantly reduce the generation of polyethylene particles in patients with total knee arthroplasty, 3 years after the surgical procedure. Copyright © 2018 SECOT. Publicado por Elsevier España, S.L.U. All rights reserved.

Collagen and elastin cross-linking is altered during aberrant late lung development associated with hyperoxia.

Science.gov (United States)

Mižíková, Ivana; Ruiz-Camp, Jordi; Steenbock, Heiko; Madurga, Alicia; Vadász, István; Herold, Susanne; Mayer, Konstantin; Seeger, Werner; Brinckmann, Jürgen; Morty, Rory E

2015-06-01

Maturation of the lung extracellular matrix (ECM) plays an important role in the formation of alveolar gas exchange units. A key step in ECM maturation is cross-linking of collagen and elastin, which imparts stability and functionality to the ECM. During aberrant late lung development in bronchopulmonary dysplasia (BPD) patients and animal models of BPD, alveolarization is blocked, and the function of ECM cross-linking enzymes is deregulated, suggesting that perturbed ECM cross-linking may impact alveolarization. In a hyperoxia (85% O2)-based mouse model of BPD, blunted alveolarization was accompanied by alterations to lung collagen and elastin levels and cross-linking. Total collagen levels were increased (by 63%). The abundance of dihydroxylysinonorleucine collagen cross-links and the dihydroxylysinonorleucine-to-hydroxylysinonorleucine ratio were increased by 11 and 18%, respectively, suggestive of a profibrotic state. In contrast, insoluble elastin levels and the abundance of the elastin cross-links desmosine and isodesmosine in insoluble elastin were decreased by 35, 30, and 21%, respectively. The lung collagen-to-elastin ratio was threefold increased. Treatment of hyperoxia-exposed newborn mice with the lysyl oxidase inhibitor β-aminopropionitrile partially restored normal collagen levels, normalized the dihydroxylysinonorleucine-to-hydroxylysinonorleucine ratio, partially normalized desmosine and isodesmosine cross-links in insoluble elastin, and partially restored elastin foci structure in the developing septa. However, β-aminopropionitrile administration concomitant with hyperoxia exposure did not improve alveolarization, evident from unchanged alveolar surface area and alveoli number, and worsened septal thickening (increased by 12%). These data demonstrate that collagen and elastin cross-linking are perturbed during the arrested alveolarization of developing mouse lungs exposed to hyperoxia. Copyright © 2015 the American Physiological Society.

Lactoferrin binding to transglutaminase cross-linked casein micelles

NARCIS (Netherlands)

Anema, S.G.; de Kruif, C.G.|info:eu-repo/dai/nl/073609609

2012-01-01

Casein micelles in skim milk were either untreated (untreated milk) or were cross-linked using transglutaminase (TGA-milk). Added lactoferrin (LF) bound to the casein micelles and followed Langmuir adsorption isotherms. The adsorption level was the same in both milks and decreased the micellar zeta

Collagen cross-linking: insights on the evolution of metazoan extracellular matrix.

Science.gov (United States)

Rodriguez-Pascual, Fernando; Slatter, David Anthony

2016-11-23

Collagens constitute a large family of extracellular matrix (ECM) proteins that play a fundamental role in supporting the structure of various tissues in multicellular animals. The mechanical strength of fibrillar collagens is highly dependent on the formation of covalent cross-links between individual fibrils, a process initiated by the enzymatic action of members of the lysyl oxidase (LOX) family. Fibrillar collagens are present in a wide variety of animals, therefore often being associated with metazoan evolution, where the emergence of an ancestral collagen chain has been proposed to lead to the formation of different clades. While LOX-generated collagen cross-linking metabolites have been detected in different metazoan families, there is limited information about when and how collagen acquired this particular modification. By analyzing telopeptide and helical sequences, we identified highly conserved, potential cross-linking sites throughout the metazoan tree of life. Based on this analysis, we propose that they have importantly contributed to the formation and further expansion of fibrillar collagens.

Photoinduced electron transfer through peptide-based self-assembled monolayers chemisorbed on gold electrodes: directing the flow-in and flow-out of electrons through peptide helices.

Science.gov (United States)

Venanzi, Mariano; Gatto, Emanuela; Caruso, Mario; Porchetta, Alessandro; Formaggio, Fernando; Toniolo, Claudio

2014-08-21

Photoinduced electron transfer (PET) experiments have been carried out on peptide self-assembled monolayers (SAM) chemisorbed on a gold substrate. The oligopeptide building block was exclusively formed by C(α)-tetrasubstituted α-aminoisobutyric residues to attain a helical conformation despite the shortness of the peptide chain. Furthermore, it was functionalized at the C-terminus by a pyrene choromophore to enhance the UV photon capture cross-section of the compound and by a lipoic group at the N-terminus for linking to gold substrates. Electron transfer across the peptide SAM has been studied by photocurrent generation experiments in an electrochemical cell employing a gold substrate modified by chemisorption of a peptide SAM as a working electrode and by steady-state and time-resolved fluorescence experiments in solution and on a gold-coated glass. The results show that the electronic flow through the peptide bridge is strongly asymmetric; i.e., PET from the C-terminus to gold is highly favored with respect to PET in the opposite direction. This effect arises from the polarity of the Au-S linkage (Au(δ+)-S(δ-), junction effect) and from the electrostatic field generated by the peptide helix.

Cross-modal links among vision, audition, and touch in complex environments.

Science.gov (United States)

Ferris, Thomas K; Sarter, Nadine B

2008-02-01

This study sought to determine whether performance effects of cross-modal spatial links that were observed in earlier laboratory studies scale to more complex environments and need to be considered in multimodal interface design. It also revisits the unresolved issue of cross-modal cuing asymmetries. Previous laboratory studies employing simple cues, tasks, and/or targets have demonstrated that the efficiency of processing visual, auditory, and tactile stimuli is affected by the modality, lateralization, and timing of surrounding cues. Very few studies have investigated these cross-modal constraints in the context of more complex environments to determine whether they scale and how complexity affects the nature of cross-modal cuing asymmetries. Amicroworld simulation of battlefield operations with a complex task set and meaningful visual, auditory, and tactile stimuli was used to investigate cuing effects for all cross-modal pairings. Significant asymmetric performance effects of cross-modal spatial links were observed. Auditory cues shortened response latencies for collocated visual targets but visual cues did not do the same for collocated auditory targets. Responses to contralateral (rather than ipsilateral) targets were faster for tactually cued auditory targets and each visual-tactile cue-target combination, suggesting an inhibition-of-return effect. The spatial relationships between multimodal cues and targets significantly affect target response times in complex environments. The performance effects of cross-modal links and the observed cross-modal cuing asymmetries need to be examined in more detail and considered in future interface design. The findings from this study have implications for the design of multimodal and adaptive interfaces and for supporting attention management in complex, data-rich domains.

Cross-linked PAN-based thin-film composite membranes for non-aqueous nanofiltration

KAUST Repository

Pérez-Manríquez, Liliana

2015-01-01

A new approach on the development of cross-linked PAN based thin film composite (TFC) membranes for non-aqueous application is presented in this work. Polypropylene backed neat PAN membranes fabricated by phase inversion process were cross-linked with hydrazine to get excellent solvent stability toward dimethylformamide (DMF). By interfacial polymerization a selective polyamide active layer was coated over the cross-linked PAN using N,N′-diamino piperazine (DAP) and trimesoyl chloride (TMC) as monomers. Permeation and molecular weight cut off (MWCO) experiments using various dyes were done to evaluate the performance of the membranes. Membranes developed by such method show excellent solvent stability toward DMF with a permeance of 1.7 L/m2 h bar and a molecular weight cut-off of less than 600 Da.

Antibiofouling hybrid dendritic Boltorn/star PEG thiol-ene cross-linked networks.

Science.gov (United States)

Bartels, Jeremy W; Imbesi, Philip M; Finlay, John A; Fidge, Christopher; Ma, Jun; Seppala, Jonathan E; Nystrom, Andreas M; Mackay, Michael E; Callow, James A; Callow, Maureen E; Wooley, Karen L

2011-06-01

probed using atomic force microscopy (AFM), where it was observed that the surface of the amphiphilic films became increasingly rough with increasing PEG wt%. On the basis of the physicochemical data from the diverse sample matrix, a focused compositional profile was then investigated further to determine the antifouling performance of the cross-linked Boltorn-PEG-PETMP networks. For these studies, a low, constant PETMP concentration of 16 wt% was maintained with variation in the PEG wt% (0-35 wt%). Antifouling and fouling-release activities were tested against the marine alga Ulva. Spore settlement densities were low on these films, compared to that on standards of polydimethylsiloxane and glass. © 2011 American Chemical Society

Two-photon induced collagen cross-linking in bioartificial cardiac tissue

Science.gov (United States)

Kuetemeyer, Kai; Kensah, George; Heidrich, Marko; Meyer, Heiko; Martin, Ulrich; Gruh, Ina; Heisterkamp, Alexander

2011-08-01

Cardiac tissue engineering is a promising strategy for regenerative therapies to overcome the shortage of donor organs for transplantation. Besides contractile function, the stiffness of tissue engineered constructs is crucial to generate transplantable tissue surrogates with sufficient mechanical stability to withstand the high pressure present in the heart. Although several collagen cross-linking techniques have proven to be efficient in stabilizing biomaterials, they cannot be applied to cardiac tissue engineering, as cell death occurs in the treated area. Here, we present a novel method using femtosecond (fs) laser pulses to increase the stiffness of collagen-based tissue constructs without impairing cell viability. Raster scanning of the fs laser beam over riboflavin-treated tissue induced collagen cross-linking by two-photon photosensitized singlet oxygen production. One day post-irradiation, stress-strain measurements revealed increased tissue stiffness by around 40% being dependent on the fibroblast content in the tissue. At the same time, cells remained viable and fully functional as demonstrated by fluorescence imaging of cardiomyocyte mitochondrial activity and preservation of active contraction force. Our results indicate that two-photon induced collagen cross-linking has great potential for studying and improving artificially engineered tissue for regenerative therapies.

Capacity of cognitive radio under imperfect secondary and cross link channel state information

KAUST Repository

Sboui, Lokman

2011-09-01

In this paper, we study the ergodic capacity of secondary user channel in a spectrum sharing scenario in which the secondary transmitter is instantaneously aware of estimated versions of the cross link (between the secondary transmitter and the primary receiver) and the secondary link Channel State Information (CSI). The secondary link optimal power profile along with the ergodic capacity are derived for a class of fading channels, under an average power constraint and an instantaneous interference outage constraint. We also show that our framework is rather general as it encompasses several previously studied spectrum sharing settings as special cases. In order to gain some insights on the capacity behavior, numerical results are shown for independent Rayleigh fading channels where it is found for instance, that at low SNR regime, only the secondary channel estimation matters and that the cross link CSI has no effect on the ergodic capacity; whereas at high SNR regime, the capacity is rather driven by the cross link CSI. © 2011 IEEE.

Oxidative cross-linking of casein by horseradish peroxidase and its ...

African Journals Online (AJOL)

STORAGESEVER

2009-10-19

Oct 19, 2009 ... The cross-linking of casein was demonstrated by capillary zone electrophoresis analysis. .... linking reaction was started by addition of 1.0 ml 3% (w/v) H2O2 and .... by Design Expert Software (Version 7.0), keeping one variable at its ... The emulsion was immediately transferred into a 250 ml capa-.

Use of cross-linked carboxymethyl cellulose for soft-tissue augmentation: preliminary clinical studies

Directory of Open Access Journals (Sweden)

Mauro Leonardis

2010-11-01

Full Text Available Mauro Leonardis1, Andrea Palange2, Rodrigo FV Dornelles3, Felipe Hund41Department of Plastic Surgery, Salvator Mundi International Hospital, Roma, Italy; 2Department of Aesthetic Medicine, Fisiobios, Roma, Italy; 3Department of Plastic Surgery, Núcleo de Plástica Avançada, São Paulo, SP, Brazil; 4Department of Plastic Surgery, Consultorio de Cirurgia Plastica, Criciuma, SC, BrazilPurpose: The continual search for new products for soft-tissue augmentation has in recent years led to the introduction of long lasting alternatives to hyaluronic acids and collagen that are composed of other polymers able to improve clinical persistence over time. This is the first report in which sodium carboxymethyl cellulose (CMC has been chemically treated by the cross-linking process and thus used as a hydrogel for soft-tissue augmentation through injection with thin needles. The study evaluates, from a clinical point of view, the behavior of cross-linked carboxymethyl cellulose hydrogel used in the aesthetic field and its side effects so as to check the safety and performance of the polymer following intradermal injections.Patients and methods: This work shows the preliminary results of an ongoing clinical study conducted between 2006 and 2009, performed on 84 healthy volunteers (62 females, 22 males aged between 18 and 72 years, for the treatment of 168 nasolabial folds, 45 perioral wrinkles, and 39 lip volume.Results: Study results show an excellent correction of facial defects. Tolerance and aesthetic quality of the correction obtained indicate considerable safety features and absence of side effects. From a clinical point of view, hydrogel is gradually absorbed into the injection site without migration issues.Conclusion: Cross-linked CMC hydrogel proves to be an ideal agent for soft tissue augmentation with regard to safety and ease of application. It did not cause infection, extrusion, migration, or adverse reactions in the patients who have been

Development of a method for environmentally friendly chemical peptide synthesis in water using water-dispersible amino acid nanoparticles

Directory of Open Access Journals (Sweden)

Fukumori Yoshinobu

2011-08-01

Full Text Available Abstract Due to the vast importance of peptides in biological processes, there is an escalating need for synthetic peptides to be used in a wide variety of applications. However, the consumption of organic solvent is extremely large in chemical peptide syntheses because of the multiple condensation steps in organic solvents. That is, the current synthesis method is not environmentally friendly. From the viewpoint of green sustainable chemistry, we focused on developing an organic solvent-free synthetic method using water, an environmentally friendly solvent. Here we described in-water synthesis technology using water-dispersible protected amino acids.

CHARACTERIZATION OF SORBENT PRODUCED THROUGH IMMOBILIZATION OF HUMIC ACID ON CHITOSAN USING GLUTARALDEHYDE AS CROSS-LINKING AGENT AND Pb(II ION AS ACTIVE SITE PROTECTOR

Directory of Open Access Journals (Sweden)

Uripto Trisno Santoso

2010-12-01

Full Text Available Sorbent produced through immobilization of humic acid (HA on chitosan using glutaraldehyde as cross-linking agent and Pb(II ions as active site protector has been characterized. Active sorption site of HA was protected by reacting HA with Pb(II ion, and the protected-HA was then activated by glutaraldehyde, crosslinked onto chitosan, and deprotected by 0.1 M disodium ethylenediamine tetra-acetic acid (Na2EDTA. The protected-crosslinking method enhanced the content of immobilized-HA and its chemical stability. Based on the FTIR spectra, crosslinking of HA on chitosan probably occurred through a chemical reaction. The sorption capacity of sorbent still remains unchanged after the second regeneration, but some of HA start to be soluble. The latter shows that cross-linking reaction between HA and chitosan is through formation an unstable product. The effectiveness of sorbent regeneration can also be identified by the XRD pattern.

Conservation of RNA sequence and cross-linking ability in ribosomes from a higher eukaryote: photochemical cross-linking of the anticodon of P site bound tRNA to the penultimate cytidine of the UACACACG sequence in Artemia salina 18S rRNA

International Nuclear Information System (INIS)

Ciesiolka, J.; Nurse, K.; Klein, J.; Ofengand, J.

1985-01-01

The complex of Artemia salina ribosomes and Escherichia coli acetylvalyl-tRNA could be cross-linked by irradiation with near-UV light. Cross-linking required the presence of the codon GUU, GUA being ineffective. The acetylvalyl group could be released from the cross-linked tRNA by treatment with puromycin, demonstrating that cross-linking had occurred at the P site. This was true both for pGUU- and also for poly(U2,G)-dependent cross-linking. All of the cross-linking was to the 18S rRNA of the small ribosomal subunit. Photolysis of the cross-link at 254 nm occurred with the same kinetics as that for the known cyclobutane dimer between this tRNA and Escherichia coli 16S rRNA. T1 RNase digestion of the cross-linked tRNA yielded an oligonucleotide larger in molecular weight than any from un-cross-linked rRNA or tRNA or from a prephotolyzed complex. Extended electrophoresis showed this material to consist of two oligomers of similar mobility, a faster one-third component and a slower two-thirds component. Each oligomer yielded two components on 254-nm photolysis. The slower band from each was the tRNA T1 oligomer CACCUCCCUVACAAGp, which includes the anticodon. The faster band was the rRNA 9-mer UACACACCGp and its derivative UACACACUG. Unexpectedly, the dephosphorylated and slower moving 9-mer was derived from the faster moving dimer. Deamination of the penultimate C to U is probably due to cyclobutane dimer formation and was evidence for that nucleotide being the site of cross-linking. Direct confirmation of the cross-linking site was obtained by Z-gel analysis

Cross-linking for microbial keratitis

Directory of Open Access Journals (Sweden)

Jayesh Vazirani

2013-01-01

Full Text Available The success of collagen cross-linking as a clinical modality to modify the clinical course in keratoconus seems to have fueled the search for alternative applications for this treatment. Current clinical data on its efficacy is limited and laboratory data seems to indicate that it performs poorly against resistant strains of bacteria and against slow growing organisms. However, the biological plausibility of crosslinking and the lack of effective strategies in managing infections with these organisms continue to focus attention on this potential treatment. Well-conducted experimental and clinical studies with controls are required to answer the questions of its efficacy in future.

Insights into the swelling process and drug release mechanisms from cross-linked pectin/high amylose starch matrices

Directory of Open Access Journals (Sweden)

Fernanda M. Carbinatto

2014-02-01

Full Text Available Cross-linked pectin/high amylose mixtures were evaluated as a new excipient for matrix tablets formulations, since the mixing of polymers and cross-linking reaction represent rational tools to reach materials with modulated and specific properties that meet specific therapeutic needs. Objective: In this work the influence of polymer ratio and cross-linking process on the swelling and the mechanism driving the drug release from swellable matrix tablets prepared with this excipient was investigated. Methods: Cross-linked samples were characterized by their micromeritic properties (size and shape, density, angle of repose and flow rate and liquid uptake ability. Matrix tablets were evaluated according their physical properties and the drug release rates and mechanisms were also investigated. Results: Cross-linked samples demonstrated size homogeneity and irregular shape, with liquid uptake ability insensible to pH. Cross-linking process of samples allowed the control of drug release rates and the drug release mechanism was influenced by both polymer ratio and cross-linking process. The drug release of samples with minor proportion of pectin was driven by an anomalous transport and the increase of the pectin proportion contributed to the erosion of the matrix. Conclusion: The cross-linked mixtures of high amylose and pectin showed a suitable excipient for slowing the drug release rates.

Impaired Cleavage of Preproinsulin Signal Peptide Linked to Autosomal-Dominant Diabetes

Science.gov (United States)

Liu, Ming; Lara-Lemus, Roberto; Shan, Shu-ou; Wright, Jordan; Haataja, Leena; Barbetti, Fabrizio; Guo, Huan; Larkin, Dennis; Arvan, Peter

2012-01-01

Recently, missense mutations upstream of preproinsulin’s signal peptide (SP) cleavage site were reported to cause mutant INS gene-induced diabetes of youth (MIDY). Our objective was to understand the molecular pathogenesis using metabolic labeling and assays of proinsulin export and insulin and C-peptide production to examine the earliest events of insulin biosynthesis, highlighting molecular mechanisms underlying β-cell failure plus a novel strategy that might ameliorate the MIDY syndrome. We find that whereas preproinsulin-A(SP23)S is efficiently cleaved, producing authentic proinsulin and insulin, preproinsulin-A(SP24)D is inefficiently cleaved at an improper site, producing two subpopulations of molecules. Both show impaired oxidative folding and are retained in the endoplasmic reticulum (ER). Preproinsulin-A(SP24)D also blocks ER exit of coexpressed wild-type proinsulin, accounting for its dominant-negative behavior. Upon increased expression of ER–oxidoreductin-1, preproinsulin-A(SP24)D remains blocked but oxidative folding of wild-type proinsulin improves, accelerating its ER export and increasing wild-type insulin production. We conclude that the efficiency of SP cleavage is linked to the oxidation of (pre)proinsulin. In turn, impaired (pre)proinsulin oxidation affects ER export of the mutant as well as that of coexpressed wild-type proinsulin. Improving oxidative folding of wild-type proinsulin may provide a feasible way to rescue insulin production in patients with MIDY. PMID:22357960

Dehydration of an ethanol/water azeotrope through alginate-DNA membranes cross-linked with metal ions by pervaporation.

Science.gov (United States)

Uragami, Tadashi; Banno, Masashi; Miyata, Takashi

2015-12-10

To obtain high dehydration membranes for an ethanol/water azeotrope, dried blend membranes prepared from mixtures of sodium alginate (Alg-Na) and sodium deoxyribonucleate (DNA-Na) were cross-linked by immersing in a methanol solution of CaCl2 or MaCl2. In the dehydration of an ethanol/water azeotropic mixture by pervaporation, the effects of immersion time in methanol solution of CaCl2 or MaCl2 on the permeation rate and water/ethanol selectivity through Alg-DNA/Ca(2+) and Alg-DNA/Mg(2+) cross-linked membranes were investigated. Alg-DNA/Mg(2+) cross-linked membrane immersed for 12h in methanol solution of MaCl2 exhibited the highest water/ethanol selectivity. This results from depressed swelling of the membranes by formation of a cross-linked structure. However, excess immersion in solution containing cross-linker led to an increase in the hydrophobicity of cross-linked membrane. Therefore, the water/ethanol selectivity of Alg-DNA/Mg(2+) cross-linked membranes with an excess immersion in cross-linking solution was lowered. The relationship between the structure of Alg-DNA/Ca(2+) and Alg-DNA/Mg(2+) cross-linked membranes and their permeation and separation characteristics during pervaporation of an ethanol/water azeotropic mixture is discussed in detail. Copyright © 2015 Elsevier Ltd. All rights reserved.

SECONDARY CYTOTOXICITY OF CROSS-LINKED DERMAL SHEEP COLLAGENS DURING REPEATED EXPOSURE TO HUMAN FIBROBLASTS

NARCIS (Netherlands)

VANLUYN, MJA; VANWACHEM, PB; DAMINK, LHHO; DIJKSTRA, PJ; FEIJEN, J; NIEUWENHUIS, P

1992-01-01

We investigated commercially available dermal sheep collagen either cross-linked with hexamethylenedlisocyanate, or cross-linked with glutaraldehyde. In previous in vitro studies we could discriminate primary, i.e. extractable, and secondary cytotoxicity, due to cell-biomaterial interactions, i.e.

Preparation and Characterization of Epoxy Resin Cross-Linked with High Wood Pyrolysis Bio-Oil Substitution by Acetone Pretreatment

Directory of Open Access Journals (Sweden)

Yi Liu

2017-03-01

Full Text Available The use of cost effective solvents may be necessary to store wood pyrolysis bio-oil in order to stabilize and control its viscosity, but this part of the production system has not been explored. Conversely, any rise in viscosity during storage, that would occur without a solvent, will add variance to the production system and render it cost ineffective. The purpose of this study was to modify bio-oil with a common solvent and then react the bio-oil with an epoxy for bonding of wood without any loss in properties. The acetone pretreatment of the bio-oil/epoxy mixture was found to improve the cross-linking potential and substitution rate based on its mechanical, chemical, and thermal properties. Specifically, the bio-oil was blended with epoxy resin at weight ratios ranging from 2:1 to 1:5 and were then cured. A higher bio-oil substitution rate was found to lower the shear bond strength of the bio-oil/epoxy resins. However, when an acetone pretreatment was used, it was possible to replace the bio-oil by as much as 50% while satisfying usage requirements. Extraction of the bio-oil/epoxy mixture with four different solvents demonstrated an improvement in cross-linking after acetone pretreatment. ATR-FTIR analysis confirmed that the polymer achieved a higher cross-linked structure. DSC and TGA curves showed improved thermal stability with the addition of the acetone pretreatment. UV-Vis characterization showed that some functional groups of the bio-oil to epoxy system were unreacted. Finally, when the resin mixture was utilized to bond wood, the acetone pretreatment coupled with precise tuning of the bio-oil:epoxy ratio was an effective method to control cross-linking while ensuring acceptable bond strength.

Kinetics of enzyme-catalyzed cross-linking of feruloylated arabinan from sugar beet

DEFF Research Database (Denmark)

Abang Zaidel, Dayang Norulfairuz; Arnous, Anis; Holck, Jesper

2011-01-01

the kinetics of HRP catalyzed cross-linking of FA esterified to α-(1,5)-linked arabinans are affected by the length of the arabinan chains carrying the feruloyl substitutions. The kinetics of the HRP-catalyzed cross-linking of four sets of arabinan samples from sugar beet pulp, having different molecular...... weights and hence different degrees of polymerization, were monitored by the disappearance of FA absorbance at 316 nm. MALDI-TOF/TOF-MS analysis confirmed that the sugar beet arabinans were feruloyl-substituted, and HPLC analysis verified that the amounts of diFAs increased when FA levels decreased...

Eco-friendly and biocompatible cross-linked carboxymethylcellulose hydrogels as adsorbents for the removal of organic dye pollutants for environmental applications.

Science.gov (United States)

Capanema, Nádia S V; Mansur, Alexandra A P; Mansur, Herman S; de Jesus, Anderson C; Carvalho, Sandhra M; Chagas, Poliane; de Oliveira, Luiz C

2017-08-28

In this study, new eco-friendly hydrogel adsorbents were synthesized based on carboxymethylcellulose (CMC, degree of substitution [DS] = 0.7) chemically cross-linked with citric acid (CA) using a green process in aqueous solution and applied for the adsorption of methylene blue (MB). Spectroscopic analyses demonstrated the mechanism of cross-linking through the reaction of hydroxyl functional groups from CMC with CA. These CMC hydrogels showed very distinct morphological features dependent on the extension of cross-linking and their nanomechanical properties were drastically increased by approximately 300% after cross-linking with 20% CA (e.g. elastic moduli from 80 ± 15 to 270 ± 50 MPa). Moreover, they were biocompatible using an in vitro cell viability assay in contact with human osteosarcoma-derived cells (SAOS) for 24 h. These CMC-based hydrogels exhibited adsorption efficiency above 90% (24 h) and maximum removal capacity of MB from 5 to 25 mg g -1 depending on the dye concentration (from 100 to 500 mg L -1 ), which was used as the model cationic organic pollutant. The adsorption of process of MB was well-fit to the pseudo-second-order kinetics model. The desorption of MB by immersion in KCl solution (3 mol L -1 , 24 h) showed a typical recovery efficiency of over 60% with conceivable reuse of these CMC-based hydrogels. Conversely, CMC hydrogels repelled methyl orange dye used as model anionic pollutant, proving the mechanism of adsorption by the formation of charged polyelectrolyte/dye complexes.

Synthesis of Cross-Linked Polymeric Micelle pH Nanosensors

DEFF Research Database (Denmark)

Ek, Pramod Kumar; Jølck, Rasmus Irming; Andresen, Thomas Lars

2015-01-01

The design flexibility that polymeric micelles offer in the fabrication of optical nanosensors for ratiometric pH measurements is investigated. pH nanosensors based on polymeric micelles are synthesized either by a mixed-micellization approach or by a postmicelle modification strategy. In the mixed......-micellization approach, self-assembly of functionalized unimers followed by shell cross-linking by copper-catalyzed azide-alkyne cycloaddition (CuAAC) results in stabilized cRGD-functionalized micelle pH nanosensors. In the postmicelle modification strategy, simultaneous cross-linking and fluorophore conjugation...... at the micelle shell using CuAAC results in a stabilized micelle pH nanosensor. Compared to the postmicelle modification strategy, the mixed-micellization approach increases the control of the overall composition of the nanosensors.Both approaches provide stable nanosensors with similar pKa profiles and thereby...

Preparation of protein- and cell-resistant surfaces by hyperthermal hydrogen induced cross-linking of poly(ethylene oxide).

Science.gov (United States)

Bonduelle, Colin V; Lau, Woon M; Gillies, Elizabeth R

2011-05-01

The functionalization of surfaces with poly(ethylene oxide) (PEO) is an effective means of imparting resistance to the adsorption of proteins and the attachment and growth of cells, properties that are critical for many biomedical applications. In this work, a new hyperthermal hydrogen induced cross-linking (HHIC) method was explored as a simple one-step approach for attaching PEO to surfaces through the selective cleavage of C-H bonds and subsequent cross-linking of the resulting carbon radicals. In order to study the effects of the process on the polymer, PEO-coated silicon wafers were prepared and the effects of different treatment times were investigated. Subsequently, using an optimized treatment time and a modified butyl polymer with increased affinity for PEO, the technique was applied to butyl rubber surfaces. All of the treated surfaces exhibited significantly reduced protein adsorption and cell growth relative to control surfaces and compared favorably with surfaces that were functionalized with PEO using conventional chemical methods. Thus HHIC is a simple and effective means of attaching PEO to non-functional polymer surfaces.

NMR spectroscopic studies of a TAT-derived model peptide in imidazolium-based ILs: influence on chemical shifts and the cis/trans equilibrium state.

Science.gov (United States)

Wiedemann, Christoph; Ohlenschläger, Oliver; Mrestani-Klaus, Carmen; Bordusa, Frank

2017-09-13

NMR spectroscopy was used to study systematically the impact of imidazolium-based ionic liquid (IL) solutions on a TAT-derived model peptide containing Xaa-Pro peptide bonds. The selected IL anions cover a wide range of the Hofmeister series of ions. Based on highly resolved one- and two-dimensional NMR spectra individual 1 H and 13 C peptide chemical shift differences were analysed and a classification of IL anions according to the Hofmeister series was derived. The observed chemical shift changes indicate significant interactions between the peptide and the ILs. In addition, we examined the impact of different ILs towards the cis/trans equilibrium state of the Xaa-Pro peptide bonds. In this context, the IL cations appear to be of exceptional importance for inducing an alteration of the native cis/trans equilibrium state of Xaa-Pro bonds in favour of the trans-isomers.

Cell protein cross-linking by erbstatin and related compounds | Center for Cancer Research

Science.gov (United States)

The scheme depicts a possible mechanism of cross-linking by erbstatin and related analogues. A mechanism of action is proposed which involves initial oxidation to reactive quinone intermediates that subsequently cross-link protein nucleophiles via multiple 1,4-Michael-type additions. Similar alkylation of protein by protein-tyrosine kinase inhibitors, such as herbimycin A, has

A photoactivatable probe for the Na+/H+ exchanger cross-links a 66-kDa renal brush border membrane protein

International Nuclear Information System (INIS)

Ross, W.; Bertrand, W.; Morrison, A.

1990-01-01

Earlier studies on LLC-PK1 cells have demonstrated two pharmacologically distinct Na+/H+ exchangers in renal epithelia. In addition, the cDNA clone for the human Na+/H+ antiporter which is growth factor activatable has been isolated and expressed. We report here the synthesis of an amiloride analogue that can be photoactivated and labeled with 125I. This analogue covalently cross-links a 66-kDa protein of bovine renal brush border membranes. A rabbit polyclonal antibody that was directed against a 20-amino acid peptide of the cytoplasmic domain of its human Na+/H+ antiporter also gives a positive Western against 66-kDa protein of bovine brush border membranes. Thus, the photoactive probe may be helpful in the isolation and purification of the brush border Na+/H+ exchanger

Total synthesis of cytochrome b562 by native chemical ligation using a removable auxiliary

Science.gov (United States)

Low, Donald W.; Hill, Michael G.; Carrasco, Michael R.; Kent, Stephen B. H.; Botti, Paolo

2001-01-01

We have completed the total chemical synthesis of cytochrome b562 and an axial ligand analogue, [SeMet7]cyt b562, by thioester-mediated chemical ligation of unprotected peptide segments. A novel auxiliary-mediated native chemical ligation that enables peptide ligation to be applied to protein sequences lacking cysteine was used. A cleavable thiol-containing auxiliary group, 1-phenyl-2-mercaptoethyl, was added to the α-amino group of one peptide segment to facilitate amide bond-forming ligation. The amine-linked 1-phenyl-2-mercaptoethyl auxiliary was stable to anhydrous hydrogen fluoride used to cleave and deprotect peptides after solid-phase peptide synthesis. Following native chemical ligation with a thioester-containing segment, the auxiliary group was cleanly removed from the newly formed amide bond by treatment with anhydrous hydrogen fluoride, yielding a full-length unmodified polypeptide product. The resulting polypeptide was reconstituted with heme and folded to form the functional protein molecule. Synthetic wild-type cyt b562 exhibited spectroscopic and electrochemical properties identical to the recombinant protein, whereas the engineered [SeMet7]cyt b562 analogue protein was spectroscopically and functionally distinct, with a reduction potential shifted by ≈45 mV. The use of the 1-phenyl-2-mercaptoethyl removable auxiliary reported here will greatly expand the applicability of total protein synthesis by native chemical ligation of unprotected peptide segments. PMID:11390992

Cross-Linking Mast Cell Specific Gangliosides Stimulates the Release of Newly Formed Lipid Mediators and Newly Synthesized Cytokines

Directory of Open Access Journals (Sweden)

Edismauro Garcia Freitas Filho

2016-01-01

Full Text Available Mast cells are immunoregulatory cells that participate in inflammatory processes. Cross-linking mast cell specific GD1b derived gangliosides by mAbAA4 results in partial activation of mast cells without the release of preformed mediators. The present study examines the release of newly formed and newly synthesized mediators following ganglioside cross-linking. Cross-linking the gangliosides with mAbAA4 released the newly formed lipid mediators, prostaglandins D2 and E2, without release of leukotrienes B4 and C4. The effect of cross-linking these gangliosides on the activation of enzymes in the arachidonate cascade was then investigated. Ganglioside cross-linking resulted in phosphorylation of cytosolic phospholipase A2 and increased expression of cyclooxygenase-2. Translocation of 5-lipoxygenase from the cytosol to the nucleus was not induced by ganglioside cross-linking. Cross-linking of GD1b derived gangliosides also resulted in the release of the newly synthesized mediators, interleukin-4, interleukin-6, and TNF-α. The effect of cross-linking the gangliosides on the MAP kinase pathway was then investigated. Cross-linking the gangliosides induced the phosphorylation of ERK1/2, JNK1/2, and p38 as well as activating both NFκB and NFAT in a Syk-dependent manner. Therefore, cross-linking the mast cell specific GD1b derived gangliosides results in the activation of signaling pathways that culminate with the release of newly formed and newly synthesized mediators.

Covalently cross-linked polyetheretherketone proton exchange membrane for DMFC

CSIR Research Space (South Africa)

Luo, H

2009-05-01

Full Text Available -7 cm2/s) and good electrochemical stability. The results suggested that cross-linked polyetheretherketone membrane is particularly promising to be used as proton exchange membrane for the direct methanol fuel cell application....

Lipid-peptide-polymer conjugates and nanoparticles thereof

Science.gov (United States)

Xu, Ting; Dong, He; Shu, Jessica

2015-06-02

The present invention provides a conjugate having a peptide with from about 10 to about 100 amino acids, wherein the peptide adopts a helical structure. The conjugate also includes a first polymer covalently linked to the peptide, and a hydrophobic moiety covalently linked to the N-terminus of the peptide, wherein the hydrophobic moiety comprises a second polymer or a lipid moiety. The present invention also provides helix bundles form by self-assembling the conjugates, and particles formed by self-assembling the helix bundles. Methods of preparing the helix bundles and particles are also provided.

Chemical deamidation: a common pitfall in large-scale N-linked glycoproteomic mass spectrometry-based analyses

DEFF Research Database (Denmark)

Palmisano, Giuseppe; Melo-Braga, Marcella Nunes; Engholm-Keller, Kasper

2012-01-01

for false positives. The confusion arises since the protein N-glycosidase F (PNGase F) reaction used to separate N-glycans from formerly glycosylated peptides catalyses the cleavage and deamidates the asparagine residue. This is typically viewed as beneficial since it acts to highlight the modification site......-linked consensus sites based on common N-linked glycoproteomics strategies without proper control experiments. Beside showing the spontaneous deamidation we provide alternative methods for validation that should be used in such experiments....

The theory and art of corneal cross-linking

Directory of Open Access Journals (Sweden)

Rebecca McQuaid

2013-01-01

Full Text Available Before the discovery of corneal cross-linking (CXL, patients with keratoconus would have had to undergo corneal transplantation, or wear rigid gas permeable lenses (RGPs that would temporarily flatten the cone, thereby improving the vision. The RGP contact lens (CL would not however alter the corneal stability and if the keratoconus was progressive, the continued steepening of the cone would occur under the RGP CL. To date, the Siena Eye has been the largest study to investigate long term effects of standard CXL. Three hundred and sixty-three eyes were treated and monitored over 4 years, producing reliable long-term results proving long-term stability of the cornea by halting the progression of keratoconus, and proving the safety of the procedure. Traditionally, CXL requires epithelial removal prior to corneal soakage of a dextran-based 0.1% riboflavin solution, followed by exposure of ultraviolet-A (UV-A light for 30 min with an intensity of 3 mW/cm2. A series of in vitro investigations on human and porcine corneas examined the best treatment parameters for standard CXL, such as riboflavin concentration, intensity, wavelength of UV-A light, and duration of treatment. Photochemically, CXL is achieved by the generation of chemical bonds within the corneal stroma through localized photopolymerization, strengthening the cornea whilst minimizing exposure to the surrounding structures of the eye. In vitro studies have shown that CXL has an effect on the biomechanical properties of the cornea, with an increased corneal rigidity of approximately 70%. This is a result of the creation of new chemical bonds within the stroma.

Tidbits for the synthesis of bis(2-sulfanylethyl)amido (SEA) polystyrene resin, SEA peptides and peptide thioesters.

Science.gov (United States)

Ollivier, Nathalie; Raibaut, Laurent; Blanpain, Annick; Desmet, Rémi; Dheur, Julien; Mhidia, Reda; Boll, Emmanuelle; Drobecq, Hervé; Pira, Silvain L; Melnyk, Oleg

2014-02-01

Protein total chemical synthesis enables the atom-by-atom control of the protein structure and therefore has a great potential for studying protein function. Native chemical ligation of C-terminal peptide thioesters with N-terminal cysteinyl peptides and related methodologies are central to the field of protein total synthesis. Consequently, methods enabling the facile synthesis of peptide thioesters using Fmoc-SPPS are of great value. Herein, we provide a detailed protocol for the preparation of bis(2-sulfanylethyl)amino polystyrene resin as a starting point for the synthesis of C-terminal bis(2-sulfanylethyl)amido peptides and of peptide thioesters derived from 3-mercaptopropionic acid. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.

The Preparation and Properties of Thermo-reversibly Cross-linked Rubber Via Diels-Alder Chemistry

NARCIS (Netherlands)

Polgar, Lorenzo Massimo; van Duin, Martin; Picchioni, Francesco

2016-01-01

A method for using Diels Alder thermo-reversible chemistry as cross-linking tool for rubber products is demonstrated. In this work, a commercial ethylene-propylene rubber, grafted with maleic anhydride, is thermo-reversibly cross-linked in two steps. The pending anhydride moieties are first modified

Membrane-Based Separation of Phenol/Water Mixtures Using Ionically and Covalently Cross-Linked Ethylene-Methacrylic Acid Copolymers

Directory of Open Access Journals (Sweden)

Alexander Mixa

2008-01-01

Full Text Available Membrane-based separation of phenol/water mixtures with concentrations of phenol between 3 wt% and 8 wt% in the feed has been performed with nonmodified as well as cross-linked ethylene-methacrylic acid (E-MAA copolymers with different amounts of methacrylic acid. As cross-linking agents, aluminium acetyl acetonate, which leads to ionically cross-linked membranes, and 2,3,5,6-tetramethyl-1,4-phenylene diamine and glycerine digycidether, leading to covalently cross-linked membranes, have been used. Generally, it was found that with increasing phenol content in the feed, the total flux is increasing whereas the enrichment factor is decreasing. Using nonmodified membranes with higher methacrylic acid monomer content in the polymer, lower fluxes and higher enrichment factors were observed. Investigation of different cross-linked membranes showed that with high phenol concentration in the feed, ionic cross-linking seems to be very promising. Furthermore, variation of feed temperature shows that ionically cross-linked membranes reached higher fluxes as well as higher enrichment factors at elevated temperatures. The temperature-dependent data were fitted based on an Arrhenius-type equation, and activation energies for the permeation of phenol and water through the membrane were calculated.

Effects of alginate hydrogel cross-linking density on mechanical and biological behaviors for tissue engineering.

Science.gov (United States)

Jang, Jinah; Seol, Young-Joon; Kim, Hyeon Ji; Kundu, Joydip; Kim, Sung Won; Cho, Dong-Woo

2014-09-01

An effective cross-linking of alginate gel was made through reaction with calcium carbonate (CaCO3). We used human chondrocytes as a model cell to study the effects of cross-linking density. Three different pore size ranges of cross-linked alginate hydrogels were fabricated. The morphological, mechanical, and rheological properties of various alginate hydrogels were characterized and responses of biosynthesis of cells encapsulated in each gel to the variation in cross-linking density were investigated. Desired outer shape of structure was maintained when the alginate solution was cross-linked with the applied method. The properties of alginate hydrogel could be tailored through applying various concentrations of CaCO3. The rate of synthesized GAGs and collagens was significantly higher in human chondrocytes encapsulated in the smaller pore structure than that in the larger pore structure. The expression of chondrogenic markers, including collagen type II and aggrecan, was enhanced in the smaller pore structure. It was found that proper structural morphology is a critical factor to enhance the performance and tissue regeneration. Copyright © 2014 Elsevier Ltd. All rights reserved.

Immobilization of cross-linked tannase enzyme on multiwalled carbon nanotubes and its catalytic behavior.

Science.gov (United States)

Ong, Chong-Boon; Annuar, Mohamad S M

2018-02-07

Immobilization of cross-linked tannase on pristine multiwalled carbon nanotubes (MWCNT) was successfully performed. Cross-linking of tannase molecules was made through glutaraldehyde. The immobilized tannase exhibited significantly improved pH, thermal, and recycling stability. The optimal pH for both free and immobilized tannase was observed at pH 5.0 with optimal operating temperature at 30°C. Moreover, immobilized enzyme retained greater biocatalytic activities upon 10 repeated uses compared to free enzyme in solution. Immobilization of tannase was accomplished by strong hydrophobic interaction most likely between hydrophobic amino acid moieties of the glutaraldehyde-cross-linked tannase to the MWCNT.

Alkali reversal of psoralen cross-link for the targeted delivery of psoralen monoadduct lesion

International Nuclear Information System (INIS)

Yeung, A.T.; Dinehart, W.J.; Jones, B.K.

1988-01-01

Psoralen intercalates into double-stranded DNA and photoreacts mainly with thymines to form monoadducts and interstrand cross-links. The authors used an oligonucleotide model to demonstrate a novel mechanism: the reversal of psoralen cross-links by base-catalyzed rearrangement at 90 0 C (BCR). The BCR reaction is more efficient than the photoreversal reaction. They show that the BCR occurs predominantly on the furan side of a psoralen cross-link. The cleavage does not result in the breaking of the DNA backbone, and the thymine based freed from the cross-link by the cleavage reaction appears to be unmodified. Similarly, BCR of the furan-side monoadduct of psoralen removed the psoralen molecule and regenerated the unaltered native oligonucleotide. The pyrone-side psoralen monoadduct is relatively resistant to BCR. One can use BCR to perform efficient oligonucleotide-directed, site-specific delivery of a psoralen monoadduct. As a demonstration of this approach, they have hybridized a 19 base long oligonucleotide vehicle containing a furan-side psoralen monoadduct to a 56 base long complementary oligonucleotide target strand and formed a specific cross-link at the target site with 365-nm UV. Subsequent BCR released the oligonucleotide vehicle and deposited the psoralen at the target site

Alzheimer's disease against peptides products of enzymatic cleavage APP protein: Biological, pathobiological and physico-chemical properties of fibrillating peptides.

Science.gov (United States)

Marszałek, Małgorzata

2017-05-17

Various peptides products of enzymatic cleavage of key for Alzheimer's disease Amyloid Precursor Protein (APP) are well known, but still are matter of scientific debate. The Aβ type products are especially challenging for experimental and medical research. This paper outlines several, still poorly known, biological and medical processes such as peptides biology, i.e., formation, biodistribution, translocation, transport and finally removal from brain compartments and body fluids like Intracellular Fluid (ICF), Cerebrospinal Fluid (CSF), Interstitial Fluid (ISF), blood serum or urine. In addition, the following studies concerning AD patients might prove challenging and simultaneously promising: peptides translocation through Blood-Brain - Barrier (BBB) and Blood-Cerebrospinal Fluid Barrier (BCSFB) and their removal from the brain according to a new concept of glymphatic system; - diagnostic difficulties that stem from physico-chemical properties and the nature of proteins or fibrillating peptides itself like low concentration, short half-live and from experimental-technical problems as well like high adsorption or low solubility of Aβ, tau or amylin. The study of diagnostic parameters is very important, as it may better reflect early changes before the disease develops; one such parameter is the Aβ42/Aβ40 ratio, or the ratio with the total tau concentration combination and other new biomarkers like Aβ1-38; other factors include oxidative stress and inflammation process proteins, complement factor H, alpha-2-macroglobulin, or clusterin. The study of various forms of pathological amyloid deposits that emerge in different but specific brain regions AD patients seems to be crucial as well. The composition of the first initial pathological, pre-fibrillating monomers of fibrillating peptides and their role in AD development and disease progression have been described as well. They are even more challenging for science and simultaneously might be more promising in

PET imaging with copper-64 as a tool for real-time in vivo investigations of the necessity for cross-linking of polymeric micelles in nanomedicine.

Science.gov (United States)

Jensen, Andreas I; Binderup, Tina; Ek, Pramod Kumar; Grandjean, Constance E; Rasmussen, Palle H; Kjaer, Andreas; Andresen, Thomas L

2017-06-30

Polymeric micelles in nanomedicine are often cross-linked to prevent disintegration in vivo. This typically requires clinically problematic chemicals or laborious procedures. In addition, cross-linking may interfere with advanced release strategies. Despite this, it is often not investigated whether cross-linking is necessary for efficient drug delivery. We used positron emission tomography (PET) imaging with 64 Cu to demonstrate general methodology for real-time in vivo investigations of micelle stability. Triblock copolymers with 4-methylcoumarin cores of ABC-type (PEG-PHEMA-PCMA) were functionalized in the handle region (PHEMA) with CB-TE2A chelators. Polymeric micelles were formed by dialysis and one half was core cross-linked (CL) by UV light and the other half was not (nonCL). Both CL and nonCL were radiolabeled with 64 Cu and compared in vivo in tumor-bearing mice, with free 64 Cu as control. Accumulation in relevant organs was quantified by region of interest analysis on PET images and ex vivo counting. It was observed that CL and nonCL showed limited differences in biodistribution from each other, whereas both differed markedly from control (free 64 Cu). This demonstrated that 4-methylcoumarin core micelles may form micelles that are stable in circulation even without cross-linking. The methodology presented here where individual unimers are radiolabeled is applicable to a wide range of polymeric micelle types. Copyright © 2017 John Wiley & Sons, Ltd.

An atomistic model for cross-linked HNBR elastomers used in seals

Science.gov (United States)

Molinari, Nicola; Sutton, Adrian; Stevens, John; Mostofi, Arash

2015-03-01

Hydrogenated nitrile butadiene rubber (HNBR) is one of the most common elastomeric materials used for seals in the oil and gas industry. These seals sometimes suffer ``explosive decompression,'' a costly problem in which gases permeate a seal at the elevated temperatures and pressures pertaining in oil and gas wells, leading to rupture when the seal is brought back to the surface. The experimental evidence that HNBR and its unsaturated parent NBR have markedly different swelling properties suggests that cross-linking may occur during hydrogenation of NBR to produce HNBR. We have developed a code compatible with the LAMMPS molecular dynamics package to generate fully atomistic HNBR configurations by hydrogenating initial NBR structures. This can be done with any desired degree of cross-linking. The code uses a model of atomic interactions based on the OPLS-AA force-field. We present calculations of the dependence of a number of bulk properties on the degree of cross-linking. Using our atomistic representations of HNBR and NBR, we hope to develop a better molecular understanding of the mechanisms that result in explosive decompression.

Simulations of tensile failure in glassy polymers: effect of cross-link density

International Nuclear Information System (INIS)

Panico, M; Narayanan, S; Brinson, L C

2010-01-01

Molecular dynamics simulations are adopted to investigate the failure mechanisms of glassy polymers, particularly with respect to increasing density of cross-links. In our simulations thermosetting polymers, which are cross-linked, exhibit an embrittlement compared with uncross-linked thermoplastics in a similar fashion to several experimental investigations (Levita et al 1991 J. Mater. Sci. 26 2348; Sambasivam et al 1997 J. Appl. Polym. Sci. 65 1001; Iijima et al 1992 Eur. Polym. J. 28 573). We perform a detailed analysis of this phenomenon and propose an interpretation based on the predominance of chain scission process over disentanglement in thermosetting polymers. We also elucidate the brittle fracture response of the thermosetting polymers

Phosphopeptide occupancy and photoaffinity cross-linking of the v-Src SH2 domain attenuates tyrosine kinase activity.

Science.gov (United States)

Garcia, P; Shoelson, S E; Drew, J S; Miller, W T

1994-12-02

Phosphorylation of c-Src at carboxyl-terminal Tyr-527 suppresses tyrosine kinase activity and transforming potential, presumably by facilitating the intramolecular interaction of the C terminus of Src with its SH2 domain. In addition, it has been shown previously that occupancy of the c-Src SH2 domain with a phosphopeptide stimulates c-Src kinase catalytic activity. We have performed analogous studies with v-Src, the transforming protein from Rous sarcoma virus, which has extensive homology with c-Src. v-Src lacks an autoregulatory phosphorylation site, and its kinase domain is constitutively active. Phosphopeptides corresponding to the sequences surrounding c-Src Tyr-527 and a Tyr-Glu-Glu-Ile motif from the hamster polyoma virus middle T antigen inhibit tyrosine kinase activity of baculovirus-expressed v-Src 2- and 4-fold, respectively. To determine the mechanism of this regulation, the Tyr-527 phosphopeptide was substituted with the photoactive amino acid p-benzoylphenylalanine at the adjacent positions (N- and C-terminal) to phosphotyrosine. These peptides photoinactivate the v-Src tyrosine kinase 5-fold in a time- and concentration-dependent manner. Furthermore, the peptides cross-link an isolated Src SH2 domain with similar rates and specificity. These data indicate that occupancy of the v-Src SH2 domain induces a conformational change that is transmitted to the kinase domain and attenuates tyrosine kinase activity.

Location of DNA-protein cross-links in mammalian cell nuclei

International Nuclear Information System (INIS)

Oleinick, N.L.

1985-01-01

DNA-protein cross-links (DPCs) occur in 1-3% of the bulk DNA of unirradiated cells, and dose-dependent increases in DPCs with γ- or UV-radiation can be detected by filter-binding. DPCs may contribute to cell lethality, since their formation is prevented by radical scavengers. Since the environment of DNA varies within eukaryotic nuclei, we have probed the composition and sub-nuclear location of DPCs. Both before and after irradiation, the major proteins cross-linked to DNA have molecular weights similar to known proteins of the nuclear matrix. The DNA cross-linked to protein is enriched in sequences which hybridize to mRNA or rRNA transcripts; such sequences are also found preferentially in preparations of nuclear matrix. When histone-depleted, matrix-associated DNA is separated from the DNA of the supercoiled ''loops'' by digestion with EcoRI and assayed for DPCs by filter binding, the frequency of DPCs is greater in the matrix. During repair of DPCs, protein-associated DNA becomes depleted in actively transcribing DNA, followed by reconstitution of the active-gene-enriched nuclear matrix. These data are consistent with known properties of the matrix and suggest the hypothesis that in intact cells, radiation-induced DPCs are primarily a product of matrix-associated DNA sequences and matrix protein

Effects of glycosylation on the conformation and dynamics of O-linked glycoproteins: Carbon-13 NMR studies of ovine submaxillary mucin

International Nuclear Information System (INIS)

Gerken, T.A.; Butenhof, K.J.; Shogren, R.

1989-01-01

Carbon-13 NMR spectroscopic studies of native and sequentially deglycosylated ovine submaxillary mucin (OSM) have been performed to examine the effects of glycosylation on the conformation and dynamics of the peptide core of O-linked glycoproteins. OSM is a large nonglobular glycoprotein in which nearly one-third of the amino acid residues are Ser and Thr which are glycosylated by the α-Neu-NAc(2-6)α-Ga1NAc- disaccharide. The β-carbon resonances of glycosylated Ser and Thr residues in intact and asialo mucin display considerable chemical shift heterogeneity which, upon the complete removal of carbohydrate, coalesces to single sharp resonances. This chemical shift heterogeneity is due to peptide sequence variability and is proposed to reflect the presence of sequence-dependent conformations of the peptide core. These different conformations are thought to be determined by steric interactions of the Ga1NAc residue with adjacent peptide residues. The absence of chemical shift heterogeneity in apo mucin is taken to indicate a loss in the peptide-carbohydrate steric interactions, consistent with a more relaxed random coiled structure. On the basis of the 13 C relaxation behavior the dynamics of the α-carbons appear to be unique to each amino acid type and glycosylation state. These results are consistent with the changes in molecular dimensions determined by light-scattering techniques for the same series of modified mucins. Taken together, these results further demonstrate that mucins possess a highly expanded conformation that is dominated by steric interactions between the peptide core and the O-linked Ga1NAc residue

Genipin Cross-Linked Glucose Oxidase and Catalase Multi-enzyme for Gluconic Acid Synthesis.

Science.gov (United States)

Cui, Caixia; Chen, Haibin; Chen, Biqiang; Tan, Tianwei

2017-02-01

In this work, glucose oxidase (GOD) and catalase (CAT) were used simultaneously to produce gluconic acid from glucose. In order to reduce the distance between the two enzymes, and therefore improve efficiency, GOD and CAT were cross-linked together using genipin. Improvements in gluconic acid production were due to quick removal of harmful intermediate hydrogen peroxide by CAT. GOD activity was significantly affected by the proportion of CAT in the system, with GOD activity in the cross-linked multi-enzyme (CLME) being 10 times higher than that in an un-cross-linked GOD/CAT mixture. The glucose conversion rate after 15 h using 15 % glucose was also 10 % higher using the CLME than was measured using a GOD/CAT mixture.

Cross-linking of streptomycin to the 16S ribosomal RNA of Escherichia coli

International Nuclear Information System (INIS)

Gravel, M.; Melancon, P.; Barkier-Gingras, L.

1987-01-01

[ 3 H]Dihydrostreptomycin was cross-linked to the 30S ribosomal subunit from Escherichia coli with the bifunctional reagent nitrogen mustard. The cross-linking primarily involved the 16S RNA. To localize the site of cross-linking of streptomycin to the 16S RNA, the authors hybridized RNA labeled with streptomycin to restriction fragments of the 16S RNA gene. Labeled RNA hybridized to DNA fragments corresponding to bases 892-917 and bases 1394-1415. These two segments of the ribosomal RNA must by juxtaposed in the ribosome, since there is a single binding site for streptomycin. This region has been implicated both in the decoding site and in the binding of initiation factor IF-3, indicating its functional importance

Higher number of pentosidine cross-links induced by ribose does not alter tissue stiffness of cancellous bone

Energy Technology Data Exchange (ETDEWEB)

Willems, Nop M.B.K., E-mail: n.willems@acta.nl [Dept. of Orthodontics, Academic Centre for Dentistry Amsterdam (ACTA), University of Amsterdam and VU University, Gustav Mahlerlaan 3004, 1081 LA Amsterdam (Netherlands); Dept. of Oral Cell Biology and Functional Anatomy, MOVE Research Institute, Academic Centre for Dentistry Amsterdam (ACTA), University of Amsterdam and VU University, Gustav Mahlerlaan 3004, 1081 LA Amsterdam (Netherlands); Langenbach, Geerling E.J. [Dept. of Oral Cell Biology and Functional Anatomy, MOVE Research Institute, Academic Centre for Dentistry Amsterdam (ACTA), University of Amsterdam and VU University, Gustav Mahlerlaan 3004, 1081 LA Amsterdam (Netherlands); Stoop, Reinout [Dept. of Metabolic Health Research, TNO, P.O. Box 2215, 2301 CE Leiden (Netherlands); Toonder, Jaap M.J. den [Dept. of Mechanical Engineering, Eindhoven University of Technology, P.O. Box 513, 5600 MB Eindhoven (Netherlands); Mulder, Lars [Dept. of Biomedical Engineering, Eindhoven University of Technology, P.O. Box 513, 5600 MB Eindhoven (Netherlands); Zentner, Andrej [Dept. of Orthodontics, Academic Centre for Dentistry Amsterdam (ACTA), University of Amsterdam and VU University, Gustav Mahlerlaan 3004, 1081 LA Amsterdam (Netherlands); Everts, Vincent [Dept. of Oral Cell Biology and Functional Anatomy, MOVE Research Institute, Academic Centre for Dentistry Amsterdam (ACTA), University of Amsterdam and VU University, Gustav Mahlerlaan 3004, 1081 LA Amsterdam (Netherlands)

2014-09-01

The role of mature collagen cross-links, pentosidine (Pen) cross-links in particular, in the micromechanical properties of cancellous bone is unknown. The aim of this study was to examine nonenzymatic glycation effects on tissue stiffness of demineralized and non-demineralized cancellous bone. A total of 60 bone samples were derived from mandibular condyles of six pigs, and assigned to either control or experimental groups. Experimental handling included incubation in phosphate buffered saline alone or with 0.2 M ribose at 37 °C for 15 days and, in some of the samples, subsequent complete demineralization of the sample surface using 8% EDTA. Before and after experimental handling, bone microarchitecture and tissue mineral density were examined by means of microcomputed tomography. After experimental handling, the collagen content and the number of Pen, hydroxylysylpyridinoline (HP), and lysylpyridinoline (LP) cross-links were estimated using HPLC, and tissue stiffness was assessed by means of nanoindentation. Ribose treatment caused an up to 300-fold increase in the number of Pen cross-links compared to nonribose-incubated controls, but did not affect the number of HP and LP cross-links. This increase in the number of Pen cross-links had no influence on tissue stiffness of both demineralized and nondemineralized bone samples. These findings suggest that Pen cross-links do not play a significant role in bone tissue stiffness. - Highlights: • The assessment of effects of glycation in bone using HPLC, microCT, and nanoindentation • Ribose incubation: 300‐fold increase in the number of pentosidine cross-links • 300‐fold increase in the number of pentosidine cross-links: no changes in bone tissue stiffness.

The Fanconi anemia pathway promotes replication-dependent DNA interstrand cross-link repair.

Science.gov (United States)

Knipscheer, Puck; Räschle, Markus; Smogorzewska, Agata; Enoiu, Milica; Ho, The Vinh; Schärer, Orlando D; Elledge, Stephen J; Walter, Johannes C

2009-12-18

Fanconi anemia is a human cancer predisposition syndrome caused by mutations in 13 Fanc genes. The disorder is characterized by genomic instability and cellular hypersensitivity to chemicals that generate DNA interstrand cross-links (ICLs). A central event in the activation of the Fanconi anemia pathway is the mono-ubiquitylation of the FANCI-FANCD2 complex, but how this complex confers ICL resistance remains enigmatic. Using a cell-free system, we showed that FANCI-FANCD2 is required for replication-coupled ICL repair in S phase. Removal of FANCD2 from extracts inhibits both nucleolytic incisions near the ICL and translesion DNA synthesis past the lesion. Reversal of these defects requires ubiquitylated FANCI-FANCD2. Our results show that multiple steps of the essential S-phase ICL repair mechanism fail when the Fanconi anemia pathway is compromised.

Glutamic Acid Selective Chemical Cleavage of Peptide Bonds.

Science.gov (United States)

Nalbone, Joseph M; Lahankar, Neelam; Buissereth, Lyssa; Raj, Monika

2016-03-04

Site-specific hydrolysis of peptide bonds at glutamic acid under neutral aqueous conditions is reported. The method relies on the activation of the backbone amide chain at glutamic acid by the formation of a pyroglutamyl (pGlu) imide moiety. This activation increases the susceptibility of a peptide bond toward hydrolysis. The method is highly specific and demonstrates broad substrate scope including cleavage of various bioactive peptides with unnatural amino acid residues, which are unsuitable substrates for enzymatic hydrolysis.

Low-friction arthroplasty of the hip using alumina ceramic and cross-linked polyethylene. A 17-year follow-up report.

Science.gov (United States)

Wroblewski, B M; Siney, P D; Fleming, P A

2005-09-01

We report the results of our continued review of 11 total hip arthroplasties using 22.225 mm alumina ceramic femoral heads on a Charnley flanged stem, articulating with chemically cross-linked polyethylene. There was an initial bedding-in of up to 0.41 mm at the articular surface in the first two years. This had not progressed further, at a minimum follow-up of 15 years. Radiographically no femoral or acetabular component showed loosening or osteolysis.

Collagen Cross-Linking: Current Status and Future Directions

Directory of Open Access Journals (Sweden)

Marine Hovakimyan

2012-01-01

Full Text Available Collagen cross-linking (CXL using UVA light and riboflavin (vitamin B2 was introduced as a clinical application to stabilize the cornea by inducing cross-links within and between collagen fibers. CXL has been investigated extensively and has been shown clinically to arrest the progression of keratoconic or post-LASIK ectasia. With its minimal cost, simplicity, and proven positive clinical outcome, CXL can be regarded as a useful approach to reduce the number of penetrating keratoplasties performed. Small case series have also indicated that CXL is beneficial in corneal edema by reducing stromal swelling behavior and in keratitis by inhibiting pathogen growth. Despite these encouraging results, CXL remains a relatively new method that is potentially associated with complications. Aspects such as side effects and recurrence rates have still to be elucidated. In light of the growing interest in CXL, our paper summarizes present knowledge about this promising approach. We have intentionally endeavored to include the more relevant studies from the recent literature to provide an overview of the current status of CXL.

Cross-linking e segmento de anel corneano intraestromal

Directory of Open Access Journals (Sweden)

Adimara da Candelaria Renesto

2011-02-01

Full Text Available O cross-linking corneano é um procedimento usado para a estabilização mecânica e aumento da rigidez corneana em pacientes com ceratocone (reduzindo a possibilidade de progressão, e também em processos inflamatórios de afinamento corneano. Os segmentos de anéis corneanos intraestromais têm como princípio o aplanamento central da córnea. Inicialmente utilizados para correção de baixa miopia, a principal indicação atual é em pacientes com ceratocone, para melhorar a acuidade visual não corrigida, a acuidade visual corrigida e permitir uma melhor tolerância ao uso de lentes de contato como também retardar a necessidade de um transplante de córnea. O objetivo deste artigo é revisar algumas publicações relacionadas ao cross-linking corneano e à inserção do segmento de anel intraestromal, apresentando suas indicações, resultados e complicações relatadas até o momento.

Cathepsin-Mediated Cleavage of Peptides from Peptide Amphiphiles Leads to Enhanced Intracellular Peptide Accumulation

Energy Technology Data Exchange (ETDEWEB)

Acar, Handan [Institute; Department; Samaeekia, Ravand [Institute; Department; Schnorenberg, Mathew R. [Institute; Department; Medical; Sasmal, Dibyendu K. [Institute; Huang, Jun [Institute; Tirrell, Matthew V. [Institute; Institute; LaBelle, James L. [Department

2017-08-24

Peptides synthesized in the likeness of their native interaction domain(s) are natural choices to target protein protein interactions (PPIs) due to their fidelity of orthostatic contact points between binding partners. Despite therapeutic promise, intracellular delivery of biofunctional peptides at concentrations necessary for efficacy remains a formidable challenge. Peptide amphiphiles (PAs) provide a facile method of intracellular delivery and stabilization of bioactive peptides. PAs consisting of biofunctional peptide headgroups linked to hydrophobic alkyl lipid-like tails prevent peptide hydrolysis and proteolysis in circulation, and PA monomers are internalized via endocytosis. However, endocytotic sequestration and steric hindrance from the lipid tail are two major mechanisms that limit PA efficacy to target intracellular PPIs. To address these problems, we have constructed a PA platform consisting of cathepsin-B cleavable PAs in which a selective p53-based inhibitory peptide is cleaved from its lipid tail within endosomes, allowing for intracellular peptide accumulation and extracellular recycling of the lipid moiety. We monitor for cleavage and follow individual PA components in real time using a resonance energy transfer (FRET)-based tracking system. Using this platform, components in real time using a Forster we provide a better understanding and quantification of cellular internalization, trafficking, and endosomal cleavage of PAs and of the ultimate fates of each component.

Sodium tripolyphosphate cross-linked chitosan based sensor for enhacing sensing properties towards acetone

Science.gov (United States)

Nasution, T. I.; Asrosa, R.; Nainggolan, I.; Balyan, M.; Indah, R.; Wahyudi, A.

2018-02-01

In this report, sensing properties of sodium tripolyphosphate (TPP) cross-linked chitosan based sensor has been successfully enhanced towards acetone. Chitosan solutions were cross-linked with sodium TPP in variation of 0.1%, 0.5%, 1% and 1.5% w/v, respectively. The sensors were fabricated in film form using an electrochemical deposition method. The sensing properties of the sensors were observed by exposing the pure chitosan and sodium TPP cross-linked chitosan sensors towards acetone concentrations of 5, 10, 50, 100 and 200 ppm. The measurement results revealed that the maximum response in output voltage value of pure chitosan sensor was 0.35 V while sodium TPP crosslinked chitosan sensors were above 0.35 V towards 5 ppm acetone concentration. When the sensors were exposed towards acetone concentration of 200 ppm, the maximum response of pure chitosan was 0.45 V while sodium TPP crosslinked chitosan sensors were above 0.45 V. Amongst the variation of sodium TPP, the maximum response of 1% sodium TPP was the highest since the maximum response was 0.4 V and 0.6 V towards 5 ppm and 200 ppm acetone concentration, respectively. While the maximum responses of other sodium TPP concentrations were under 0.4 V and 0.6 V towards 5 ppm and 200 ppm acetone concentration. Moreover, 1% sodium TPP cross-linked chitosan based sensor showed good reproducibility and outstanding lifetime. Therefore, 1% sodium TPP cross-linked chitosan based sensor has exhibited remarkable sensing properties as a novel acetone sensor.

Low-Temperature Cross-Linking of PEDOT:PSS Films Using Divinylsulfone.

Science.gov (United States)

Mantione, Daniele; Del Agua, Isabel; Schaafsma, Wandert; ElMahmoudy, Mohammed; Uguz, Ilke; Sanchez-Sanchez, Ana; Sardon, Haritz; Castro, Begoña; Malliaras, George G; Mecerreyes, David

2017-05-31

Recent interest in bioelectronics has prompted the exploration of properties of conducting polymer films at the interface with biological milieus. Poly(3,4-ethylenedioxythiophene) doped with poly(styrenesulfonate) (PEDOT:PSS) from a commercially available source has been used as a model system for these studies. Different cross-linking schemes have been used to stabilize films of this material against delamination and redispersion, but the cost is a decrease in the electrical conductivity and/or additional heat treatment. Here we introduce divinylsulfone (DVS) as a new cross-linker for PEDOT:PSS. Thanks to the higher reactiveness of the vinyl groups of DVS, the cross-linking can be performed at room temperature. In addition, DVS does not reduce electronic conductivity of PEDOT:PSS but rather increases it by acting as a secondary dopant. Cell culture studies show that PEDOT:PSS:DVS films are cytocompatible and support neuroregeneration. As an example, we showed that this material improved the transconductance value and stability of an organic electrochemical transistor (OECT) device. These results open the way for the utilization of DVS as an effective cross-linker for PEDOT:PSS in bioelectronics applications.

Scleral lens tolerance after corneal cross-linking for keratoconus

NARCIS (Netherlands)

Visser, Esther Simone; Soeters, Nienke; Tahzib, Nayyirih G.

2015-01-01

Purpose. Subjective and objective evaluation of scleral lens tolerance and fitting before and after corneal cross-linking (CXL) for progressive keratoconus. Methods. In this prospective cohort, evaluations were made of 18 unilateral eyes in patients who underwent CXL and had been wearing scleral

Mechanocatalytic polymerization and cross-linking in a polymeric matrix

NARCIS (Netherlands)

Jakobs, R.T.M.; Ma, Shuang; Sijbesma, R.P.

2013-01-01

A latent olefin metathesis catalyst, bearing two polymeric NHC ligands, was embedded in a semicrystalline polymer matrix containing cyclic olefins. The catalyst was activated by straining the solid material under compression, resulting in polymerization and cross-linking reactions of the monomers in

Functionalisation of cross-linked polyethylenimine for the removal of ...

African Journals Online (AJOL)

... and describe the experimental data. The thermodynamic study of the adsorption process indicated high activation energies (55.91 kJ mol-1) which confirms chemisorption as a mechanism of interaction between As and PCPEI. Keywords: Adsorption; arsenic; phosphonated cross-linked polyethylenimine, functionalisation ...

Folic acid-targeted disulfide-based cross-linking micelle for enhanced drug encapsulation stability and site-specific drug delivery against tumors

Directory of Open Access Journals (Sweden)

Zhang Y

2016-03-01

Full Text Available Yumin Zhang,1,* Junhui Zhou,2,* Cuihong Yang,1 Weiwei Wang,3 Liping Chu,1 Fan Huang,1 Qiang Liu,1 Liandong Deng,2 Deling Kong,3 Jianfeng Liu,1 Jinjian Liu1 1Tianjin Key Laboratory of Radiation Medicine and Molecular Nuclear Medicine, Institute of Radiation Medicine, Chinese Academy of Medical Science and Peking Union Medical College, 2Department of Polymer Science and Technology, School of Chemical Engineering and Technology, Tianjin University, 3Tianjin Key Laboratory of Biomaterial Research, Institute of Biomedical Engineering, Chinese Academy of Medical Science and Peking Union Medical College, Tianjin, People’s Republic of China *These authors contributed equally in this work Abstract: Although the shortcomings of small molecular antitumor drugs were efficiently improved by being entrapped into nanosized vehicles, premature drug release and insufficient tumor targeting demand innovative approaches that boost the stability and tumor responsiveness of drug-loaded nanocarriers. Here, we show the use of the core cross-linking method to generate a micelle with enhanced drug encapsulation ability and sensitivity of drug release in tumor. This kind of micelle could increase curcumin (Cur delivery to HeLa cells in vitro and improve tumor accumulation in vivo. We designed and synthesized the core cross-linked micelle (CCM with polyethylene glycol and folic acid-polyethylene glycol as the hydrophilic units, pyridyldisulfide as the cross-linkable and hydrophobic unit, and disulfide bond as the cross-linker. CCM showed spherical shape with a diameter of 91.2 nm by the characterization of dynamic light scattering and transmission electron microscope. Attributed to the core cross-linking, drug-loaded CCM displayed higher Nile Red or Cur-encapsulated stability and better sensitivity to glutathione than noncross-linked micelle (NCM. Cellular uptake and in vitro antitumor studies proved the enhanced endocytosis and better cytotoxicity of CCM-Cur against

Analysis of glycation induced protein cross-linking inhibitory effects of some antidiabetic plants and spices.

Science.gov (United States)

Perera, Handunge Kumudu Irani; Handuwalage, Charith Sandaruwan

2015-06-09

Protein cross-linking which occurs towards the latter part of protein glycation is implicated in the development of chronic diabetic complications. Glycation induced protein cross-linking inhibitory effects of nine antidiabetic plants and three spices were evaluated in this study using a novel, simple, electrophoresis based method. Methanol extracts of thirteen plants including nine antidiabetic plants and three spices were used. Lysozyme and fructose were incubated at 37 °C in the presence or absence of different concentrations of plant extracts up to 31 days. Standard glycation inhibitor aminoguanidine and other appropriate controls were included. A recently established sodium dodecyl polyacrylamide gel electrophoresis (SDS-PAGE) method was used to detect the products of protein cross-linking in the incubation mixtures. High molecular weight protein products representing the dimer, trimer and tetramer of lysozyme were detected in the presence of fructose. Among the nine antidiabetic plants, seven showed glycation induced protein cross-linking inhibitory effects namely Ficus racemosa (FR) stem bark, Gymnema sylvestre (GS) leaves, Musa paradisiaca (MP) yam, Phyllanthus debilis (PD) whole plant, Phyllanthus emblica (PE) fruit, Pterocarpus marsupium (PM) latex and Tinospora cordifolia (TC) leaves. Inhibition observed with Coccinia grandis (CG) leaves and Strychnos potatorum (SP) seeds were much low. Leaves of Gymnema lactiferum (GL), the plant without known antidiabetic effects showed the lowest inhibition. All three spices namely Coriandrum sativum (CS) seeds, Cinnamomum zeylanicum (CZ) bark and Syzygium aromaticum (SA) flower buds showed cross-link inhibitory effects with higher effects in CS and SA. PD, PE, PM, CS and SA showed almost complete inhibition on the formation of cross-linking with 25 μg/ml extracts. Methanol extracts of PD, PE, PM, CS and SA have shown promising inhibitory effects on glycation induced protein cross-linking.

Wear of PEEK-OPTIMA® and PEEK-OPTIMA®-Wear Performance articulating against highly cross-linked polyethylene.

Science.gov (United States)

East, Rebecca H; Briscoe, Adam; Unsworth, Anthony

2015-03-01

The idea of all polymer artificial joints, particularly for the knee and finger, has been raised several times in the past 20 years. This is partly because of weight but also to reduce stress shielding in the bone when stiffer materials such as metals or ceramics are used. With this in mind, pin-on-plate studies of various polyetheretherketone preparations against highly cross-linked polyethylene were conducted to investigate the possibility of using such a combination in the design of a new generation of artificial joints. PEEK-OPTIMA(®) (no fibre) against highly cross-linked polyethylene gave very low wear factors of 0.0384 × 10(-6) mm(3)/N m for the polyetheretherketone pins and -0.025 × 10(-6) mm(3)/N m for the highly cross-linked polyethylene plates. The carbon-fibre-reinforced polyetheretherketone (PEEK-OPTIMA(®)-Wear Performance) also produced very low wear rates in the polyetheretherketone pins but produced very high wear in the highly cross-linked polyethylene, as might have been predicted since the carbon fibres are quite abrasive. When the fibres were predominantly tangential to the sliding plane, the mean wear factor was 0.052 × 10(-6) mm(3)/N m for the pins and 49.3 × 10(-6) mm(3)/N m for the highly cross-linked polyethylene plates; a half of that when the fibres ran axially in the pins (0.138 × 10(-6) mm(3)/N m for the pins and 97.5 × 10(-6) mm/ N m for the cross-linked polyethylene plates). PEEK-OPTIMA(®) against highly cross-linked polyethylene merits further investigation. © IMechE 2015.

Synthesis and characterization of a biocompatible chitosan-based hydrogel cross-linked via 'click' chemistry for controlled drug release.

Science.gov (United States)

Guaresti, O; García-Astrain, C; Palomares, T; Alonso-Varona, A; Eceiza, A; Gabilondo, N

2017-09-01

A chemically cross-linked chitosan-based hydrogel was successfully synthesized through Diels-Alder (DA) reaction and characterized. The final product was obtained after different steps; on the one hand, furan-modified chitosan (Cs-Fu) was synthesized by the reaction of furfural with the free amino groups of chitosan. On the other hand, highlighting the novelty of the present research, maleimide-functionalized chitosan (Cs-AMI) was prepared by the reaction of a maleimide-modified aminoacid with the amino groups of chitosan through amide coupling. The two complementary chitosan derivatives were cross-linked to the final hydrogel network. Both modification reactions were confirmed by FTIR and 1 H NMR, obtaining a degree of substitution (DS) of 31% and 26% for Cs-Fu and Cs-AMI, respectively. The as-designed hydrogel was analyzed in terms of microstructure, swelling capacity and rheological behaviour. The hydrogel showed pH-sensitivity, biocompatibility and inhibitory bacterial activity, promising features for biomedical applications, particularly for targeted-drug delivery. Copyright © 2017 Elsevier B.V. All rights reserved.

Urinary osteocalcin and serum pro-C-type natriuretic peptide predict linear catch-up growth in infants

DEFF Research Database (Denmark)

Kilpeläinen, Leena; Ivaska, Kaisa K; Kuiri-Hänninen, Tanja

2012-01-01

of this longitudinal study was to determine the extent to which postnatal levels of circulating cartilage (serum pro-C-type natriuretic peptide [S-proCNP]) and urinary bone metabolic markers (urinary osteocalcin [MidOC] and two forms of C-terminal cross-linked telopeptide of type I collagen [U-α-CTX-I and U...

Entropic benefit of a cross-link in protein association.

Science.gov (United States)

Zaman, Muhammad H; Berry, R Stephen; Sosnick, Tobin R

2002-08-01

We introduce a method to estimate the loss of configurational entropy upon insertion of a cross-link to a dimeric system. First, a clear distinction is established between the loss of entropy upon tethering and binding, two quantities that are often considered to be equivalent. By comparing the probability distribution of the center-to-center distances for untethered and cross-linked versions, we are able to calculate the loss of translational entropy upon cross-linking. The distribution function for the untethered helices is calculated from the probability that a given helix is closer to its partner than to all other helices, the "Nearest Neighbor" method. This method requires no assumptions about the nature of the solvent, and hence resolves difficulties normally associated with calculations for systems in liquids. Analysis of the restriction of angular freedom upon tethering indicates that the loss of rotational entropy is negligible. The method is applied in the context of the folding of a ten turn helical coiled coil with the tether modeled as a Gaussian chain or a flexible amino acid chain. After correcting for loop closure entropy in the docked state, we estimate the introduction of a six-residue tether in the coiled coil results in an effective concentration of the chain to be about 4 or 100 mM, depending upon whether the helices are denatured or pre-folded prior to their association. Thus, tethering results in significant stabilization for systems with millimolar or stronger dissociation constants. Copyright 2002 Wiley-Liss, Inc.

Micropattern printing of adhesion, spreading, and migration peptides on poly(tetrafluoroethylene) films to promote endothelialization.

Science.gov (United States)

Gauvreau, Virginie; Laroche, Gaétan

2005-01-01

We report here the development of an original multistep micropatterning technique for printing peptides on surfaces, based on the ink-jet printer technology. Contrary to most micropatterning methods used nowadays, this technique is advantageous because it allows displaying 2D-arrays of multiple biomolecules. Moreover, this low cost procedure allies the advantages of computer-aided design with high flexibility and reproducibility. A Hewlett-Packard printer was modified to print peptide solutions, and Adobe Illustrator was used as the graphic-editing software to design high-resolution checkerboard-like micropatterns. In a first step, PTFE films were treated with ammonia plasma to introduce amino groups on the surface. These chemical functionalities were reacted with heterobifunctional cross-linker sulfo-succinimidyl 4-(N-maleimidomethyl)cycloexane-1-carboxylate (S-SMCC) to allow the subsequent surface covalent conjugation of various cysteine-modified peptides to the polymer substrate. These peptidic molecules containing RGD and WQPPRARI sequences were selected for their adhesive, spreading, and migrational properties toward endothelial cells. On one hand, our data demonstrated that the initial cell adhesion does not depend on the chemical structure and combination of the peptides covalently bonded either through conventional conjugation or micropatterning. On the other hand, spreading and migration of endothelial cells is clearly enhanced while coconjugating the GRGDS peptide in conjunction with WQPPRARI. This behavior is further improved by micropatterning these peptides on specific areas of the polymer surface.

Helleborus purpurascens—Amino Acid and Peptide Analysis Linked to the Chemical and Antiproliferative Properties of the Extracted Compounds

Directory of Open Access Journals (Sweden)

Adina-Elena Segneanu

2015-12-01

Full Text Available There is a strong drive worldwide to discover and exploit the therapeutic potential of a large variety of plants. In this work, an alcoholic extract of Helleborus purpurascens (family Ranunculaceae was investigated for the identification of amino acids and peptides with putative antiproliferative effects. In our work, a separation strategy was developed using solvents of different polarity in order to obtain active compounds. Biochemical components were characterized through spectroscopic (mass spectroscopy and chromatographic techniques (RP-HPLC and GC-MS. The biological activity of the obtained fractions was investigated in terms of their antiproliferative effects on HeLa cells. Through this study, we report an efficient separation of bioactive compounds (amino acids and peptides from a plant extract dependent on solvent polarity, affording fractions with unaffected antiproliferative activities. Moreover, the two biologically tested fractions exerted a major antiproliferative effect, thereby suggesting potential anticancer therapeutic activity.

Helleborus purpurascens-Amino Acid and Peptide Analysis Linked to the Chemical and Antiproliferative Properties of the Extracted Compounds.

Science.gov (United States)

Segneanu, Adina-Elena; Grozescu, Ioan; Cziple, Florentina; Berki, Daniel; Damian, Daniel; Niculite, Cristina Mariana; Florea, Alexandru; Leabu, Mircea

2015-12-11

There is a strong drive worldwide to discover and exploit the therapeutic potential of a large variety of plants. In this work, an alcoholic extract of Helleborus purpurascens (family Ranunculaceae) was investigated for the identification of amino acids and peptides with putative antiproliferative effects. In our work, a separation strategy was developed using solvents of different polarity in order to obtain active compounds. Biochemical components were characterized through spectroscopic (mass spectroscopy) and chromatographic techniques (RP-HPLC and GC-MS). The biological activity of the obtained fractions was investigated in terms of their antiproliferative effects on HeLa cells. Through this study, we report an efficient separation of bioactive compounds (amino acids and peptides) from a plant extract dependent on solvent polarity, affording fractions with unaffected antiproliferative activities. Moreover, the two biologically tested fractions exerted a major antiproliferative effect, thereby suggesting potential anticancer therapeutic activity.

Effect of pore size and cross-linking of a novel collagen-elastin dermal substitute on wound healing.

Science.gov (United States)

Boekema, Bouke K H L; Vlig, Marcel; Olde Damink, Leon; Middelkoop, Esther; Eummelen, Lizette; Bühren, Anne V; Ulrich, Magda M W

2014-02-01

Collagen-elastin (CE) scaffolds are frequently used for dermal replacement in the treatment of full-thickness skin defects such as burn wounds. But little is known about the optimal pore size and level of cross-linking. Different formulations of dermal substitutes with unidirectional pores were tested in porcine full-thickness wounds in combination with autologous split skin mesh grafts (SSG). Effect on wound healing was evaluated both macro- and microscopically. CE scaffolds with a pore size of 80 or 100 μm resulted in good wound healing after one-stage grafting. Application of scaffolds with a larger average pore size (120 μm) resulted in more myofibroblasts and more foreign body giant cells (FBGC). Moderate crosslinking impaired wound healing as it resulted in more wound contraction, more FBGC and increased epidermal thickness compared to no cross-linking. In addition, take rate and redness were negatively affected compared to SSG only. Vascularization and the number of myofibroblasts were not affected by cross-linking. Surprisingly, stability of cross-linked scaffolds was not increased in the wound environment, in contrast to in vitro results. Cross-linking reduced the proliferation of fibroblasts in vitro, which might explain the reduced clinical outcome. The non-cross-linked CE substitute with unidirectional pores allowed one-stage grafting of SSG, resulting in good wound healing. In addition, only a very mild foreign body reaction was observed. Cross-linking of CE scaffolds negatively affected wound healing on several important parameters. The optimal non-cross-linked CE substitute is a promising candidate for future clinical evaluation.

Use of Activated Carbon in Packaging to Attenuate Formaldehyde-Induced and Formic Acid-Induced Degradation and Reduce Gelatin Cross-Linking in Solid Dosage Forms.

Science.gov (United States)

Colgan, Stephen T; Zelesky, Todd C; Chen, Raymond; Likar, Michael D; MacDonald, Bruce C; Hawkins, Joel M; Carroll, Sophia C; Johnson, Gail M; Space, J Sean; Jensen, James F; DeMatteo, Vincent A

2016-07-01

Formaldehyde and formic acid are reactive impurities found in commonly used excipients and can be responsible for limiting drug product shelf-life. Described here is the use of activated carbon in drug product packaging to attenuate formaldehyde-induced and formic acid-induced drug degradation in tablets and cross-linking in hard gelatin capsules. Several pharmaceutical products with known or potential vulnerabilities to formaldehyde-induced or formic acid-induced degradation or gelatin cross-linking were subjected to accelerated stability challenges in the presence and absence of activated carbon. The effects of time and storage conditions were determined. For all of the products studied, activated carbon attenuated drug degradation or gelatin cross-linking. This novel use of activated carbon in pharmaceutical packaging may be useful for enhancing the chemical stability of drug products or the dissolution stability of gelatin-containing dosage forms and may allow for the 1) extension of a drug product's shelf-life when the limiting attribute is a degradation product induced by a reactive impurity, 2) marketing of a drug product in hotter and more humid climatic zones than currently supported without the use of activated carbon, and 3) enhanced dissolution stability of products that are vulnerable to gelatin cross-linking. Copyright © 2016 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

Enzyme-linked immunosorbent assay using a virus type-specific peptide based on a subdomain of envelope protein e(rns) for serologic diagnosis of pestivirus infections in swine

NARCIS (Netherlands)

Langedijk, J.P.; Middel, W.G.; Meloen, R.H.; Kramps, J.A.; Smit, de J.A.

2001-01-01

Peptides deduced from the C-terminal end (residues 191 to 227) of pestivirus envelope protein Erns were used to develop enzyme-linked immunosorbent assays (ELISAs) to measure specifically antibodies against different types of pestiviruses. The choice of the peptide was based on the modular structure

Cross-protective peptide vaccine against influenza A viruses developed in HLA-A*2402 human immunity model.

Directory of Open Access Journals (Sweden)

Toru Ichihashi

Full Text Available BACKGROUND: The virus-specific cytotoxic T lymphocyte (CTL induction is an important target for the development of a broadly protective human influenza vaccine, since most CTL epitopes are found on internal viral proteins and relatively conserved. In this study, the possibility of developing a strain/subtype-independent human influenza vaccine was explored by taking a bioinformatics approach to establish an immunogenic HLA-A24 restricted CTL epitope screening system in HLA-transgenic mice. METHODOLOGY/PRINCIPAL FINDINGS: HLA-A24 restricted CTL epitope peptides derived from internal proteins of the H5N1 highly pathogenic avian influenza A virus were predicted by CTL epitope peptide prediction programs. Of 35 predicted peptides, six peptides exhibited remarkable cytotoxic activity in vivo. More than half of the mice which were subcutaneously vaccinated with the three most immunogenic and highly conserved epitopes among three different influenza A virus subtypes (H1N1, H3N2 and H5N1 survived lethal influenza virus challenge during both effector and memory CTL phases. Furthermore, mice that were intranasally vaccinated with these peptides remained free of clinical signs after lethal virus challenge during the effector phase. CONCLUSIONS/SIGNIFICANCE: This CTL epitope peptide selection system can be used as an effective tool for the development of a cross-protective human influenza vaccine. Furthermore this vaccine strategy can be applicable to the development of all intracellular pathogens vaccines to induce epitope-specific CTL that effectively eliminate infected cells.

Rheological Behavior, Granule Size Distribution and Differential Scanning Calorimetry of Cross-Linked Banana (Musa paradisiaca) Starch.

Science.gov (United States)

Núñez-Santiago, María C.; Maristany-Cáceres, Amira J.; Suárez, Francisco J. García; Bello-Pérez, Arturo

2008-07-01

Rheological behavior at 60 °C, granule size distribution and Differential Scanning Calorimetry (DSC) tests were employed to study the effect of diverse reaction conditions: adipic acid concentration, pH and temperature during cross-linking of banana (Musa paradisiaca) starch. These properties were determined in native banana starch pastes for the purpose of comparison. Rheological behavior from pastes of cross-linked starch at 60 °C did not show hysteresis, probably due the cross-linkage of starch that avoided disruption of granules, elsewhere, native starch showed hysteresis in a thixotropic loop. All pastes exhibited non-Newtonian shear thinning behavior. In all cases, size distribution showed a decrease in the median diameter in cross-linked starches. This condition produces a decrease in swelling capacity of cross-linked starch. The median diameter decreased with an increase of acid adipic concentration; however, an increase of pH and Temperature produced an increase in this variable. Finally, an increase in gelatinization temperature and entalphy (ΔH) were observed as an effect of cross-linkage. An increase in acid adipic concentration produced an increase in Tonset and a decrease in ΔH. pH and temperature. The cross-linked of banana starch produced granules more resistant during the pasting procedure.

An electron microscopic study of the photochemical cross-linking of DNA in guinea pig epidermis by psoralen derivatives

International Nuclear Information System (INIS)

Cech, T.; Pathak, M.A.; Biswas, R.K.

1979-01-01

Albino guinea pigs were treated with psoralen derivatives plus 320-400 nm ultraviolet radiation, and DNA was extracted from their epidermis. The DNA was assayed for the presence of interstrand cross-links by standard denaturation-renaturation assays and by a new technique, electron microscopy of the DNA under totally denaturing conditions. The latter method allows individual cross-links to be directly observed and counted. When either 4,5',8-trimethylpsoralen or 8-methoxypsoralen was applied topically to the skin (8-20 μg/cm 2 ) or administered orally (10-12 mg/kg body weight), followed by exposure to 320-400 nm ultraviolet radiation, most of the epidermal DNA was found to contain a high frequency of cross-links. For example, oral or topical trimethylpsoralen treatment gave an average of one cross-link per 250 nucleotide pairs or about 3 . 10 5 cross-links per guinea pig chromosome. When the dose of either drug was decreased 20-fold to the level used in the clinical treatment of psoriasis, however, no cross-links could be detected in the epidermal DNA. The electron microscopic assay is sensitive enough that one can put an upper limit of 1 cross-link per 10 6 nucleotide pairs (80 cross-links per chromosome) for the low dose studies. The significance of these findings to the understanding of the effectiveness of psoralens in psoriasis therapy is discussed. (Auth.)

Application of polymers cross-linked by electron beam irradiation to electric wire industry

International Nuclear Information System (INIS)

Oda, Eisuke

1976-01-01

Applications of the polymers cross-linked by electron beam irradiation to electric wire industry as an example of dully developed utilization are reviewed. The report is divided into five parts, namely 1) radiation sources and irradiation processes, 2) development of crosslinking materials, 3) accumulation of electric charge and accumulation of heat, 4) examples of application, and 5) future prospect. Such a phenomenon as discharge destruction pattern (Lichtenberg figure) must be solved, when cable insulation materials are cross-linked by electron beam irradiation. The measures for preventing the discharge destruction are required, especially when the layers of polyethylene insulation for high voltage cables are irradiated. The accumulation of heat causes the troubles in foaming, degeneration and wire running of high potential cables, when the layers of insulation are thick. Effective promoters for cross-linking must be studied to reduce the radiation dose. The irradiators capable of irradiating wires uniformly are desirable. Electron beam accelerators will be used, as far as the radiation dose of 10 or more Mrad is required for cross-linking irradiation. If the dose of one tenth or less of the above value is required, gamma-ray sources (RI) are rather easily applicable than focused strong beam. The utilization of spent nuclear fuel is desirable. (Iwakiri, K.)

Reorientational motion of a cross-link junction in a poly(dimethylsiloxane) network measured by time-resolved fluorescence depolarization

International Nuclear Information System (INIS)

Stein, A.D.; Hoffman, D.A.; Frank, C.W.; Fayer, M.D.

1992-01-01

The reorientational dynamics of a cross-link junction in poly(dimethylsiloxane) networks, measured by the fluorescence anisotropy decay of a chromophore tagged to the cross-link, have been investigated over a range of temperatures from T g +75 to T g +150. The probe chromophore, 1-dimethylamino-5-sulfonylnaphthalene amide (dansyl amide), is pendant to a trifunctional silane that acts as a cross-linking molecule. In cyclohexanol, the fluorescence anisotropy decay is in agreement with Debye--Stokes--Einstein hydrodynamic theory (rotational diffusion) demonstrating that the cross-linker can be used as a probe of orientational relaxation. The fluorescence anisotropy decays at a rapid rate in an end-linked poly(dimethyl siloxane) network reflecting fast reorientational motion of the cross-link junction. This reorientation appears diffusive and has a temperature dependence in accord with the Williams--Landel--Ferry equation. A model is proposed that suggests that reorientation and translational motion of the cross-link occur simultaneously and are both coupled to fluctuations of the polymer chain ends

Damage and fatigue in cross-linked rubbers

Science.gov (United States)

Melnikov, Alexei

Damage and fatigue of elastomers have not been fundamentally understood because of the complex nature of these materials. All currently existing models are completely phenomenological. Therefore two problems have been investigated in this research to address those fundamental issues. The first problem was creating an innovative concept with a mathematical modeling, which would be able to describe the damage using molecular characteristics of elastomers. The second problem is developing new approaches to study fatigue, and especially impact fatigue of elastomers. The following results have been obtained in this research. A theoretical model of damage has been developed which involves the basic molecular characteristics of cross-linked elastomers and takes into account the effects of viscoelasticity and stress-induced crystallization. This model was found very reliable and successful in description of numerous quasi-static simple extension experiments for monotonous and repeating loadings. It also roughly predicts in molecular terms the failure of elastomers with various degrees of cross-linking. Quasi-impact fatigue tests with different geometry of an indenter have also been performed. Some microscopic features of rubber damage have been investigated using optical microscopy and SEM. In particular, the accumulation of a completely de-vulcanized, liquid-like substance was observed under intense, multi-cycle impacts. All the findings discovered in quasi-impact experiments are consistent with the damage model predictions.

Iodinated derivatives of vasoactive intestinal peptide (VIP), PHI and PHM: purification, chemical characterization and biological activity

International Nuclear Information System (INIS)

McMaster, D.; Suzuki, Y.; Rorstad, O.; Lederis, K.

1987-01-01

The iodination of vasoactive intestinal peptide (VIP) was studied, using a variety of enzymatic and chemical iodination methods. Reversed phase high performance liquid chromatography (HPLC) was used to purify the reaction products. The lactoperoxidase-glucose oxidase method gave excellent results in terms of reproducibility, iodine incorporation, and yield of the non-oxidized products [Tyr(I)10]VIP and [Tyr(I)22]VIP, and was used to prepare both 125 I and 127 I labelled derivatives. In both cases, direct application to HPLC and a single column system were used. Although the oxidized peptides [Tyr(I)10,Met(O)17]VIP and [Tyr(I)22,Met(O)17]VIP could be generated to varying degrees directly by iodination of VIP, these were most conveniently prepared by iodination of [Met(O)17]VIP. Iodinated derivatives of the homologous peptides PHI and PHM were likewise prepared by rapid, one-step HPLC procedures. The site and degree of iodination were determined by HPLC peptide mapping of tryptic digests and amino acid analyses, and in the case of [Tyr(I)10]VIP also by sequencing. The vasorelaxant activities of the iodinated peptides in bovine cerebral artery preparations did not differ significantly from those of the corresponding noniodinated peptides, with the exception of [Tyr(I)10,Met(O)17]VIP and [Tyr(I)22,Met(O)17]VIP which, unlike [Met(O)17]VIP itself, had slightly lower potency than VIP

Enzymatic cross-linking of human recombinant elastin (HELP) as biomimetic approach in vascular tissue engineering.

Science.gov (United States)

Bozzini, Sabrina; Giuliano, Liliana; Altomare, Lina; Petrini, Paola; Bandiera, Antonella; Conconi, Maria Teresa; Farè, Silvia; Tanzi, Maria Cristina

2011-12-01

The use of polymers naturally occurring in the extracellular matrix (ECM) is a promising strategy in regenerative medicine. If compared to natural ECM proteins, proteins obtained by recombinant DNA technology have intrinsic advantages including reproducible macromolecular composition, sequence and molecular mass, and overcoming the potential pathogens transmission related to polymers of animal origin. Among ECM-mimicking materials, the family of recombinant elastin-like polymers is proposed for drug delivery applications and for the repair of damaged elastic tissues. This work aims to evaluate the potentiality of a recombinant human elastin-like polypeptide (HELP) as a base material of cross-linked matrices for regenerative medicine. The cross-linking of HELP was accomplished by the insertion of cross-linking sites, glutamine and lysine, in the recombinant polymer and generating ε-(γ-glutamyl) lysine links through the enzyme transglutaminase. The cross-linking efficacy was estimated by infrared spectroscopy. Freeze-dried cross-linked matrices showed swelling ratios in deionized water (≈2500%) with good structural stability up to 24 h. Mechanical compression tests, performed at 37°C in wet conditions, in a frequency sweep mode, indicated a storage modulus of 2/3 kPa, with no significant changes when increasing number of cycles or frequency. These results demonstrate the possibility to obtain mechanically resistant hydrogels via enzymatic crosslinking of HELP. Cytotoxicity tests of cross-linked HELP were performed with human umbilical vein endothelial cells, by use of transwell filter chambers for 1-7 days, or with its extracts in the opportune culture medium for 24 h. In both cases no cytotoxic effects were observed in comparison with the control cultures. On the whole, the results suggest the potentiality of this genetically engineered HELP for regenerative medicine applications, particularly for vascular tissue regeneration.

Two DD-carboxypeptidases from Mycobacterium smegmatis affect cell surface properties through regulation of peptidoglycan cross-linking and glycopeptidolipids.

Science.gov (United States)

Pandey, Satya Deo; Pal, Shilpa; Kumar N, Ganesh; Bansal, Ankita; Mallick, Sathi; Ghosh, Anindya S

2018-05-07

During the peptidoglycan (PG) maturation of mycobacteria, the glycan strands are interlinked by both 3-3 (between two meso-DAP) and 4-3 cross-links (between D-ala and meso-DAP), though there is a predominance (60-80%) of 3-3 cross-links. The DD-CPases act on pentapeptides to generate tetrapeptides that are used by LD-transpeptidases as substrates to form 3-3 cross-links. Therefore, DD-CPases play a crucial role in mycobacterial PG cross-link formation. However, the physiology of DD-CPases in mycobacteria is relatively unexplored. Here, we deleted two DD-CPase genes, msmeg_2433 , and msmeg_2432 , both individually and in combination, from Mycobacterium smegmatis mc 2 155. Though the single DD-CPase deletions had no significant impact on the mycobacterial physiology, many interesting functional alterations were observed in the double deletion mutant, viz. , a predominance in PG cross-link formation was shifted from 3-3 cross-links to 4-3, cell surface glycopeptidolipid (GPL) expression was reduced and susceptibility towards β-lactams and anti-tubercular agents was enhanced. Moreover, the existence of the double mutant within murine macrophages was better as compared to the parent. Interestingly, the complementation with any one of the DD-CPase genes could restore the wild-type phenotype. In a nutshell, we infer that the altered ratio of 4-3: 3-3 PG cross-links might have influenced the expression of surface GPLs, colony morphology, biofilm formation,, drug susceptibility and subsistence of the cells within macrophages. Importance The glycan strands in mycobacterial peptidoglycan (PG) are interlinked by both 3-3 and 4-3 cross-links. The DD-CPases generate tetrapeptides by acting on the pentapeptides, and LD-transpeptidases use tetrapeptides as substrates to form 3-3 cross-links. Here, we showed that simultaneous deletions of two DD-CPases alter the nature of PG cross-linking from 3-3 cross-links to 4-3 cross-links. The deletions subsequently decrease the expression

The properties of water in swollen cross-linked polystyrene sulfo acids

Science.gov (United States)

Gagarin, A. N.; Tokmachev, M. G.; Kovaleva, S. S.; Ferapontov, N. B.

2008-11-01

The properties of water in polystyrene sulfo acid gels with various cross-linking degrees were studied by optical volumetry and dynamic desorption porosimetry. The isotherms of water desorption obtained by dynamic desorption porosimetry coincided with isopiestic isotherms, which allowed this method to be recommended for the determination of the amount of water in polymer gels. Joint optical volumetry and dynamic desorption porosimetry studies showed that the interphase boundary in the cross-liked hydrophilic polymer-water system did not coincide with the visible gel boundary, because gels were two-phase systems, which contained water of two types, “free” and “bound.” The influence of the degree of polymer cross-linking on the amounts and properties of water of the two types was studied. It was shown that constants of water distribution in the polymer could be calculated from the dynamic desorption porosimetry data.

Characterization of solid UV cross-linked PEGDA for biological applications

KAUST Repository

Castro, David; Ingram, Patrick; Kodzius, Rimantas; Conchouso Gonzalez, David; Yoon, Euisik; Foulds, Ian G.

2013-01-01

This paper reports on solid UV cross-linked Poly(ethylene)-glycol-diacrylate (PEGDA) as a material for microfluidic devices for biological applications. We have evaluated biocompatibility of PEGDA through two separate means: 1) by examining cell

Novel thermally cross-linked polyimide membranes for ethanol dehydration via pervaporation

KAUST Repository

Xu, Sheng

2015-12-01

© 2015 Elsevier B.V. In this work, two novel carboxyl-containing polyimides, 2,2\\'-bis(3,4-dicarboxyphenyl) hexafluoropropane dianhydride-4,4\\'-diaminodiphenylmethane/3,5-diaminobenzoic acid (6FDA-MDA/DABA, FMD) and 3,3\\',4,4\\'-benzophenone tetracarboxylic dianhydride-4,4\\'-diaminodiphenylmethane/3,5-diaminobenzoic acid (BTDA-MDA/DABA, BMD), are synthesized via chemical and thermal imidization methods, respectively, and employed as pervaporation membranes for ethanol dehydration. Chemical structures of the two polyimides are examined by FTIR and TGA to confirm the successful synthesis. A post thermal treatment of the polyimide membranes with the temperature range of 250 to 400. °C is applied, and its effects on the membrane morphology and separation performance are studied and characterized by FTIR, TGA, WXRD, solubility and sorption test. It is believed that the thermal treatment of the carboxyl-containing polyimide membrane at a relative low temperature only leads to the physical annealing, while it may cause the decarboxylation-induced cross-linking at a higher temperature. In addition, the operation temperature in pervaporation is also varied and shown to be an important factor to affect the final membrane performance. Performance benchmarking shows that the developed polyimide membranes both have superior pervaporation performance to most other flat-sheet dense membranes. This work is believed to shed useful insights on polyimide membranes for pervaporation applications.

Novel thermally cross-linked polyimide membranes for ethanol dehydration via pervaporation

KAUST Repository

Xu, Sheng; Wang, Yan

2015-01-01

© 2015 Elsevier B.V. In this work, two novel carboxyl-containing polyimides, 2,2'-bis(3,4-dicarboxyphenyl) hexafluoropropane dianhydride-4,4'-diaminodiphenylmethane/3,5-diaminobenzoic acid (6FDA-MDA/DABA, FMD) and 3,3',4,4'-benzophenone tetracarboxylic dianhydride-4,4'-diaminodiphenylmethane/3,5-diaminobenzoic acid (BTDA-MDA/DABA, BMD), are synthesized via chemical and thermal imidization methods, respectively, and employed as pervaporation membranes for ethanol dehydration. Chemical structures of the two polyimides are examined by FTIR and TGA to confirm the successful synthesis. A post thermal treatment of the polyimide membranes with the temperature range of 250 to 400. °C is applied, and its effects on the membrane morphology and separation performance are studied and characterized by FTIR, TGA, WXRD, solubility and sorption test. It is believed that the thermal treatment of the carboxyl-containing polyimide membrane at a relative low temperature only leads to the physical annealing, while it may cause the decarboxylation-induced cross-linking at a higher temperature. In addition, the operation temperature in pervaporation is also varied and shown to be an important factor to affect the final membrane performance. Performance benchmarking shows that the developed polyimide membranes both have superior pervaporation performance to most other flat-sheet dense membranes. This work is believed to shed useful insights on polyimide membranes for pervaporation applications.

Protein cross-linking by chlorinated polyamines and transglutamylation stabilizes neutrophil extracellular traps.

Science.gov (United States)

Csomós, Krisztián; Kristóf, Endre; Jakob, Bernadett; Csomós, István; Kovács, György; Rotem, Omri; Hodrea, Judit; Bagoly, Zsuzsa; Muszbek, Laszlo; Balajthy, Zoltán; Csősz, Éva; Fésüs, László

2016-08-11

Neutrophil extracellular trap (NET) ejected from activated dying neutrophils is a highly ordered structure of DNA and selected proteins capable to eliminate pathogenic microorganisms. Biochemical determinants of the non-randomly formed stable NETs have not been revealed so far. Studying the formation of human NETs we have observed that polyamines were incorporated into the NET. Inhibition of myeloperoxidase, which is essential for NET formation and can generate reactive chlorinated polyamines through hypochlorous acid, decreased polyamine incorporation. Addition of exogenous primary amines that similarly to polyamines inhibit reactions catalyzed by the protein cross-linker transglutaminases (TGases) has similar effect. Proteomic analysis of the highly reproducible pattern of NET components revealed cross-linking of NET proteins through chlorinated polyamines and ɛ(γ-glutamyl)lysine as well as bis-γ-glutamyl polyamine bonds catalyzed by the TGases detected in neutrophils. Competitive inhibition of protein cross-linking by monoamines disturbed the cross-linking pattern of NET proteins, which resulted in the loss of the ordered structure of the NET and significantly reduced capacity to trap bacteria. Our findings provide explanation of how NETs are formed in a reproducible and ordered manner to efficiently neutralize microorganisms at the first defense line of the innate immune system.

Vitamin E diffused highly cross-linked polyethylene in total hip arthroplasty at five years

DEFF Research Database (Denmark)

Nebergall, Audrey K; Greene, M. E.; Laursen, M B

2017-01-01

Aims: The objective of this five-year prospective, blinded, randomised controlled trial (RCT) was to compare femoral head penetration into a Vitamin E diffused highly cross-linked polyethylene (HXLPE) liner with penetration into a medium cross-linked polyethylene control liner using......, ArComXL. This is the longest-term RCT comparing the wear performance and clinical outcome of Vitamin E diffused HXLPE with a previous generation of medium cross-linked polyethylene....... radiostereometric analysis. Patients and Methods: Patients scheduled for total hip arthroplasty (THA) were randomised to receive either the study E1 (32 patients) or the control ArComXL polyethylene (35 patients). The median age (range) of the overall cohort was 66 years (40 to 76). Results: The five-year median...

Biocompatibility of two model elastin-like recombinamer-based hydrogels formed through physical or chemical cross-linking for various applications in tissue engineering and regenerative medicine.

Science.gov (United States)

Ibáñez-Fonseca, Arturo; Ramos, Teresa L; González de Torre, Israel; Sánchez-Abarca, Luis Ignacio; Muntión, Sandra; Arias, Francisco Javier; Del Cañizo, María Consuelo; Alonso, Matilde; Sánchez-Guijo, Fermín; Rodríguez-Cabello, José Carlos

2018-03-01

Biocompatibility studies, especially innate immunity induction, in vitro and in vivo cytotoxicity, and fibrosis, are often lacking for many novel biomaterials including recombinant protein-based ones, such as elastin-like recombinamers (ELRs), and has not been extensively explored in the scientific literature, in contrast to traditional biomaterials. Herein, we present the results from a set of experiments designed to elucidate the preliminary biocompatibility of 2 types of ELRs that are able to form extracellular matrix-like hydrogels through either physical or chemical cross-linking both of which are intended for different applications in tissue engineering and regenerative medicine. Initially, we present in vitro cytocompatibility results obtained upon culturing human umbilical vein endothelial cells on ELR substrates, showing optimal proliferation up to 9 days. Regarding in vivo cytocompatibility, luciferase-expressing hMSCs were viable for at least 4 weeks in terms of bioluminescence emission when embedded in ELR hydrogels and injected subcutaneously into immunosuppressed mice. Furthermore, both types of ELR-based hydrogels were injected subcutaneously in immunocompetent mice and serum TNFα, IL-1β, IL-4, IL-6, and IL-10 concentrations were measured by enzyme-linked immunosorbent assay, confirming the lack of inflammatory response, as also observed upon macroscopic and histological evaluation. All these findings suggest that both types of ELRs possess broad biocompatibility, thus making them very promising for tissue engineering and regenerative medicine-related applications. Copyright © 2017 John Wiley & Sons, Ltd.

Sensitivity of ab Initio vs Empirical Methods in Computing Structural Effects on NMR Chemical Shifts for the Example of Peptides.

Science.gov (United States)

Sumowski, Chris Vanessa; Hanni, Matti; Schweizer, Sabine; Ochsenfeld, Christian

2014-01-14

The structural sensitivity of NMR chemical shifts as computed by quantum chemical methods is compared to a variety of empirical approaches for the example of a prototypical peptide, the 38-residue kaliotoxin KTX comprising 573 atoms. Despite the simplicity of empirical chemical shift prediction programs, the agreement with experimental results is rather good, underlining their usefulness. However, we show in our present work that they are highly insensitive to structural changes, which renders their use for validating predicted structures questionable. In contrast, quantum chemical methods show the expected high sensitivity to structural and electronic changes. This appears to be independent of the quantum chemical approach or the inclusion of solvent effects. For the latter, explicit solvent simulations with increasing number of snapshots were performed for two conformers of an eight amino acid sequence. In conclusion, the empirical approaches neither provide the expected magnitude nor the patterns of NMR chemical shifts determined by the clearly more costly ab initio methods upon structural changes. This restricts the use of empirical prediction programs in studies where peptide and protein structures are utilized for the NMR chemical shift evaluation such as in NMR refinement processes, structural model verifications, or calculations of NMR nuclear spin relaxation rates.

Defective DNA cross-link removal in Chinese hamster cell mutants hypersensitive to bifunctional alkylating agents

International Nuclear Information System (INIS)

Hoy, C.A.; Thompson, L.H.; Mooney, C.L.; Salazar, E.P.

1985-01-01

DNA repair-deficient mutants from five genetic complementation groups isolated previously from Chinese hamster cells were assayed for survival after exposure to the bifunctional alkylating agents mitomycin C or diepoxybutane. Groups 1, 3, and 5 exhibited 1.6- to 3-fold hypersensitivity compared to the wild-type cells, whereas Groups 2 and 4 exhibited extraordinary hypersensitivity. Mutants from Groups 1 and 2 were exposed to 22 other bifunctional alkylating agents in a rapid assay that compared cytotoxicity of the mutants to the wild-type parental strain, AA8. With all but two of the compounds, the Group 2 mutant (UV4) was 15- to 60-fold more sensitive than AA8 or the Group 1 mutant (UV5). UV4 showed only 6-fold hypersensitivity to quinacrine mustard. Alkaline elution measurements showed that this compound produced few DNA interstrand cross-links but numerous strand breaks. Therefore, the extreme hypersensitivity of mutants from Groups 2 and 4 appeared specific for compounds the main cytotoxic lesions of which were DNA cross-links. Mutant UV5 was only 1- to 4-fold hypersensitive to all the compounds. Although the initial number of cross-links was similar for the three cell lines, the efficiency of removal of cross-links was lowest in UV4 and intermediate in UV5. These results suggest that the different levels of sensitivity are specifically related to different efficiencies of DNA cross-link removal. The phenotype of hypersensitivity to both UV radiation and cross-link damage exhibited by the mutants in Groups 2 and 4 appears to differ from those of the known human DNA repair syndromes

Peptides, polypeptides and peptide-polymer hybrids as nucleic acid carriers.

Science.gov (United States)

Ahmed, Marya

2017-10-24

Cell penetrating peptides (CPPs), and protein transduction domains (PTDs) of viruses and other natural proteins serve as a template for the development of efficient peptide based gene delivery vectors. PTDs are sequences of acidic or basic amphipathic amino acids, with superior membrane trespassing efficacies. Gene delivery vectors derived from these natural, cationic and cationic amphipathic peptides, however, offer little flexibility in tailoring the physicochemical properties of single chain peptide based systems. Owing to significant advances in the field of peptide chemistry, synthetic mimics of natural peptides are often prepared and have been evaluated for their gene expression, as a function of amino acid functionalities, architecture and net cationic content of peptide chains. Moreover, chimeric single polypeptide chains are prepared by a combination of multiple small natural or synthetic peptides, which imparts distinct physiological properties to peptide based gene delivery therapeutics. In order to obtain multivalency and improve the gene delivery efficacies of low molecular weight cationic peptides, bioactive peptides are often incorporated into a polymeric architecture to obtain novel 'polymer-peptide hybrids' with improved gene delivery efficacies. Peptide modified polymers prepared by physical or chemical modifications exhibit enhanced endosomal escape, stimuli responsive degradation and targeting efficacies, as a function of physicochemical and biological activities of peptides attached onto a polymeric scaffold. The focus of this review is to provide comprehensive and step-wise progress in major natural and synthetic peptides, chimeric polypeptides, and peptide-polymer hybrids for nucleic acid delivery applications.

Temperature dependence of creep compliance of highly cross-linked epoxy: A molecular simulation study

International Nuclear Information System (INIS)

Khabaz, Fardin; Khare, Ketan S.; Khare, Rajesh

2014-01-01

We have used molecular dynamics (MD) simulations to study the effect of temperature on the creep compliance of neat cross-linked epoxy. Experimental studies of mechanical behavior of cross-linked epoxy in literature commonly report creep compliance values, whereas molecular simulations of these systems have primarily focused on the Young’s modulus. In this work, in order to obtain a more direct comparison between experiments and simulations, atomistically detailed models of the cross-linked epoxy are used to study their creep compliance as a function of temperature using MD simulations. The creep tests are performed by applying a constant tensile stress and monitoring the resulting strain in the system. Our results show that simulated values of creep compliance increase with an increase in both time and temperature. We believe that such calculations of the creep compliance, along with the use of time temperature superposition, hold great promise in connecting the molecular insight obtained from molecular simulation at small length- and time-scales with the experimental behavior of such materials. To the best of our knowledge, this work is the first reported effort that investigates the creep compliance behavior of cross-linked epoxy using MD simulations

Sequence specificity for uridylylation of the viral peptide linked to the genome (VPg) of enteroviruses.

Science.gov (United States)

Schein, Catherine H; Ye, Mengyi; Paul, Aniko V; Oberste, M Steven; Chapman, Nora; van der Heden van Noort, Gerbrand J; Filippov, Dmitri V; Choi, Kyung H

2015-10-01

Enteroviruses (EV) uridylylate a peptide, VPg, as the first step in their replication. VPgpUpU, found free in infected cells, serves as the primer for RNA elongation. The abilities of four polymerases (3D(pol)), from EV-species A-C, to uridylylate VPgs that varied by up to 60% of their residues were compared. Each 3D(pol) was able to uridylylate all five VPgs using polyA RNA as template, while showing specificity for its own genome encoded peptide. All 3D(pol) uridylylated a consensus VPg representing the physical chemical properties of 31 different VPgs. Thus the residues required for uridylylation and the enzymatic mechanism must be similar in diverse EV. As VPg-binding sites differ in co-crystal structures, the reaction is probably done by a second 3D(pol) molecule. The conservation of polymerase residues whose mutation reduces uridylylation but not RNA elongation is compared. Copyright © 2015 Elsevier Inc. All rights reserved.

Reorientational motion of a cross-link junction in a poly(dimethylsiloxane) network measured by time-resolved fluorescence depolarization

Energy Technology Data Exchange (ETDEWEB)

Stein, A.D. (Department of Chemistry, Stanford University, Stanford, California 94305 (United States)); Hoffman, D.A. (Department of Materials Science and Engineering, Stanford University, Stanford, California 94305 (United States)); Frank, C.W. (Department of Chemical Engineering, Stanford University, Stanford, California 94305 (United States)); Fayer, M.D. (Department of Chemistry, Stanford University, Stanford, California 94305 (United States))

1992-02-15

The reorientational dynamics of a cross-link junction in poly(dimethylsiloxane) networks, measured by the fluorescence anisotropy decay of a chromophore tagged to the cross-link, have been investigated over a range of temperatures from {ital T}{sub {ital g}}+75 to {ital T}{sub {ital g}}+150. The probe chromophore, 1-dimethylamino-5-sulfonylnaphthalene amide (dansyl amide), is pendant to a trifunctional silane that acts as a cross-linking molecule. In cyclohexanol, the fluorescence anisotropy decay is in agreement with Debye--Stokes--Einstein hydrodynamic theory (rotational diffusion) demonstrating that the cross-linker can be used as a probe of orientational relaxation. The fluorescence anisotropy decays at a rapid rate in an end-linked poly(dimethyl siloxane) network reflecting fast reorientational motion of the cross-link junction. This reorientation appears diffusive and has a temperature dependence in accord with the Williams--Landel--Ferry equation. A model is proposed that suggests that reorientation and translational motion of the cross-link occur simultaneously and are both coupled to fluctuations of the polymer chain ends.

Sharp kink of DNA at psoralen-cross-link site deduced from crystal structure of psoralen-thymine monoadduct

International Nuclear Information System (INIS)

Kim, S.H.; Peckler, S.; Graves, B.; Kanne, D.; Rapoport, H.; Hearst, J.E.

1983-01-01

Light-induced cross-linking of double-stranded nucleic acids by psoralens has been exploited to locate, in vivo or in vitro, those double-helical regions of DNA or RNA that can accommodate any structural changes caused by the psoralen cross-links. To determine three-dimensional structural parameters of the cross-link, we have solved the crystal structure of the psoralen-thymine monoadduct formed in photoreaction between calf thymus DNA and 8-methoxypsoralen (8MOP). There are eight possible configurations for psoralen-thymine monoadducts and 64 for diadducts. We describe here the structural details of a psoralen-thymine monoadduct obtained in a biological environment and the consequences of the photo-cross-link between 8MOP and double-helical DNA

Wood Sawdust/Natural Rubber Ecocomposites Cross-Linked by Electron Beam Irradiation

Directory of Open Access Journals (Sweden)

Elena Manaila

2016-06-01

Full Text Available The obtaining and characterization of some polymeric eco-composites based on wood sawdust and natural rubber is presented. The natural rubber was cross-linked using the electron beam irradiation. The irradiation doses were of 75, 150, 300 and 600 kGy and the concentrations of wood sawdust were of 10 and 20 phr, respectively. As a result of wood sawdust adding, the physical and mechanical properties such as hardness, modulus at 100% elongation and tensile strength, showed significant improvements. The presence of wood sawdust fibers has a reinforcing effect on natural rubber, similar or better than of mineral fillers. An increase in the irradiation dose leads to the increasing of cross-link density, which is reflected in the improvement of hardness, modulus at 100% elongation and tensile strength of blends. The cross-linking rates, appreciated using the Flory-Rehner equation, have increased with the amount of wood sawdust in blends and with the irradiation dose. Even if the gel fraction values have varied irregularly with the amount of wood sawdust and irradiation dose it was over 90% for all blends, except for the samples without wood sawdust irradiated with 75 kGy. The water uptake increased with increasing of fiber content and decreased with the irradiation dose.

Glutaraldehyde Cross-Linking of TendonMechanical Effects at the Level of the Tendon Fascicle and Fibril

DEFF Research Database (Denmark)

Hansen, P.; Svensson, R.B.; Aagaard, P.

2009-01-01

were examined by atomic force microscopy. Peak forces increased from 1379 to 2622 pN while an extended Hertz fit of force-indentation data showed a 24 fold increase in Young's modulus on indentation. The effect of glutaraldehyde cross-linking on the tensile properties of a single collagen fibril......Conclusive insight into the microscopic principles that govern the strength of tendon and related connective tissues is lacking and the importance of collagen cross-linking has not been firmly established. The combined application of whole-tissue mechanical testing and atomic force spectroscopy...... allowed for a detailed characterization of the effect of cross-linking in rat-tail tendon. The cross-link inducing agent glutaraldehyde augmented the tensile strength of tendon fascicles. Stress at failure increased from 8 MPa to 39 MPa. The mechanical effects of glutaraldehyde at the tendon fibril level...

beta-Scission of C-3 (beta-carbon) alkoxyl radicals on peptides and proteins

DEFF Research Database (Denmark)

Headlam, H A; Mortimer, A; Easton, C J

2000-01-01

Exposure of proteins to radicals in the presence of O(2) brings about multiple changes in the target molecules. These alterations include oxidation of side chains, fragmentation, cross-linking, changes in hydrophobicity and conformation, altered susceptibility to proteolytic enzymes, and formation...... of methanal (formaldehyde). This product has been quantified with a number of oxidized peptides and proteins, and can account for up to 64% of the initial attacking radicals with some Ala peptides. When quantified together with the hydroperoxide precursors, these species account for up to 80% of the initial...... radicals, confirming that this is a major process. Methanal causes cell toxicity and DNA damage and is an animal carcinogen and a genotoxic agent in human cells. Thus, the formation and subsequent reaction of alkoxyl radicals formed at the C-3 position on aliphatic amino acid side chains on peptides...

Effectiveness of trimethylopropane trimethacrylate for the electron-beam-irradiation-induced cross-linking of polylactic acid

Energy Technology Data Exchange (ETDEWEB)

Ng, Hon-Meng [Department of Chemical Engineering, Faculty of Engineering and Science, Universiti Tunku Abdul Rahman, Jalan Genting Kelang, 53300 Setapak, Kuala Lumpur (Malaysia); Bee, Soo-Tueen, E-mail: beest@utar.edu.my [Department of Chemical Engineering, Faculty of Engineering and Science, Universiti Tunku Abdul Rahman, Jalan Genting Kelang, 53300 Setapak, Kuala Lumpur (Malaysia); Ratnam, C.T. [Radiation Processing Technology Division, Malaysian Nuclear Agency, Bangi, 43000 Kajang, Selangor (Malaysia); Sin, Lee Tin; Phang, Yee-Yao; Tee, Tiam-Ting [Department of Chemical Engineering, Faculty of Engineering and Science, Universiti Tunku Abdul Rahman, Jalan Genting Kelang, 53300 Setapak, Kuala Lumpur (Malaysia); Rahmat, A.R. [Department of Polymer Engineering, Faculty of Chemical Engineering, Universiti Teknologi Malaysia, 81310 UTM Skudai, Johor (Malaysia)

2014-01-15

Highlights: • Investigation of trimethylopropane trimethacrylate (TMPTMA) on electron beam irradiated PLA. • Irradiated PLA blends were weakened by incorporation of high amount of TMPTMA. • TMPTMA interacts with polymer free radicals to build crosslinking network. -- Abstract: The purpose of this research was to investigate the effects of various loading levels of trimethylopropane trimethacrylate (TMPTMA) on the properties of polylactic acid (PLA) cross-linked via electron-beam irradiation. PLA was compounded with 3–5 wt.% of TMPTMA to induce cross-linking upon subjection to electron-beam irradiation doses of 25–250 kGy. The physical properties of the PLA samples were characterised by means of X-ray diffraction, gel fraction and scanning electron microscopy analyses on fractured surfaces after tensile tests. The presence of TMPTMA in PLA was found to effectively increase the crystallite size and gel fraction. However, higher loading levels of TMPTMA could compromise the properties of the PLA/TMPTMA samples, indicating that a larger amount of monomer free radicals might promote degradation within the substantially cross-linked amorphous phase. Irradiation-induced cross-linking in the samples could improve the cross-linking density while decreasing the elongation and interfering with the crystallisation. These effects are caused by the intensive irradiation-induced chain scission that is responsible for the deterioration of the mechanical and crystalline properties of the samples.

Effectiveness of trimethylopropane trimethacrylate for the electron-beam-irradiation-induced cross-linking of polylactic acid

International Nuclear Information System (INIS)

Ng, Hon-Meng; Bee, Soo-Tueen; Ratnam, C.T.; Sin, Lee Tin; Phang, Yee-Yao; Tee, Tiam-Ting; Rahmat, A.R.

2014-01-01

Highlights: • Investigation of trimethylopropane trimethacrylate (TMPTMA) on electron beam irradiated PLA. • Irradiated PLA blends were weakened by incorporation of high amount of TMPTMA. • TMPTMA interacts with polymer free radicals to build crosslinking network. -- Abstract: The purpose of this research was to investigate the effects of various loading levels of trimethylopropane trimethacrylate (TMPTMA) on the properties of polylactic acid (PLA) cross-linked via electron-beam irradiation. PLA was compounded with 3–5 wt.% of TMPTMA to induce cross-linking upon subjection to electron-beam irradiation doses of 25–250 kGy. The physical properties of the PLA samples were characterised by means of X-ray diffraction, gel fraction and scanning electron microscopy analyses on fractured surfaces after tensile tests. The presence of TMPTMA in PLA was found to effectively increase the crystallite size and gel fraction. However, higher loading levels of TMPTMA could compromise the properties of the PLA/TMPTMA samples, indicating that a larger amount of monomer free radicals might promote degradation within the substantially cross-linked amorphous phase. Irradiation-induced cross-linking in the samples could improve the cross-linking density while decreasing the elongation and interfering with the crystallisation. These effects are caused by the intensive irradiation-induced chain scission that is responsible for the deterioration of the mechanical and crystalline properties of the samples

Chemo-enzymatic peptide synthesis : bioprocess engineering aspects

NARCIS (Netherlands)

Vossenberg, P.

2012-01-01

Peptides, in particular oligopeptides, play an important role in the fields of health care, nutrition and cosmetics. Chemical synthesis is currently the most mature technique for the synthesis of peptides that range in length from 5 to 80 amino acids. Chemical synthesis is, however,

Genipin Cross-Linked Polymeric Alginate-Chitosan Microcapsules for Oral Delivery: In-Vitro Analysis

Directory of Open Access Journals (Sweden)

Hongmei Chen

2009-01-01

Full Text Available We have previously reported the preparation of the genipin cross-linked alginate-chitosan (GCAC microcapsules composed of an alginate core with a genipin cross-linked chitosan membrane. This paper is the further investigation on their structural and physical characteristics. Results showed that the GCAC microcapsules had a smooth and dense surface and a networked interior. Cross-linking by genipin substantially reduced swelling and physical disintegration of microcapsules induced by nongelling ions and calcium sequestrants. Strong resistance to mechanical shear forces and enzymatic degradation was observed. Furthermore, the GCAC membranes were permeable to bovine serum albumin and maintained a molecular weight cutoff at 70 KD, analogous to the widely studied alginate-chitosan, and alginate-poly-L-lysine-alginate microcapsules. The release features and the tolerance of the GCAC microcapsules in the stimulated gastrointestinal environment were also investigated. This GCAC microcapsule formulation offers significant potential as a delivery vehicle for many biomedical applications.

21 CFR 177.2710 - Styrene-divinylbenzene resins, cross-linked.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Styrene-divinylbenzene resins, cross-linked. 177.2710 Section 177.2710 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN CONSUMPTION (CONTINUED) INDIRECT FOOD ADDITIVES: POLYMERS Substances for Use Only as Components of Articles Intended...

Bifunctional alkylating agent-mediated MGMT-DNA cross-linking and its proteolytic cleavage in 16HBE cells

International Nuclear Information System (INIS)

Cheng, Jin; Ye, Feng; Dan, Guorong; Zhao, Yuanpeng; Wang, Bin; Zhao, Jiqing; Sai, Yan; Zou, Zhongmin

2016-01-01

Nitrogen mustard (NM), a bifunctional alkylating agent (BAA), contains two alkyl arms and can act as a cross-linking bridge between DNA and protein to form a DNA-protein cross-link (DPC). O 6 -methylguanine–DNA methyltransferase (MGMT), a DNA repair enzyme for alkyl adducts removal, is found to enhance cell sensitivity to BAAs and to promote damage, possibly due to its stable covalent cross-linking with DNA mediated by BAAs. To investigate MGMT-DNA cross-link (mDPC) formation and its possible dual roles in NM exposure, human bronchial epithelial cell line 16HBE was subjected to different concentrations of HN2, a kind of NM, and we found mDPC was induced by HN2 in a concentration-dependent manner, but the mRNA and total protein of MGMT were suppressed. As early as 1 h after HN2 treatment, high mDPC was achieved and the level maintained for up to 24 h. Quick total DPC (tDPC) and γ-H2AX accumulation were observed. To evaluate the effect of newly predicted protease DVC1 on DPC cleavage, we applied siRNA of MGMT and DVC1, MG132 (proteasome inhibitor), and NMS-873 (p97 inhibitor) and found that proteolysis plays a role. DVC1 was proven to be more important in the cleavage of mDPC than tDPC in a p97-dependent manner. HN2 exposure induced DVC1 upregulation, which was at least partially contributed to MGMT cleavage by proteolysis because HN2-induced mDPC level and DNA damage was closely related with DVC1 expression. Homologous recombination (HR) was also activated. Our findings demonstrated that MGMT might turn into a DNA damage promoter by forming DPC when exposed to HN2. Proteolysis, especially DVC1, plays a crucial role in mDPC repair. - Highlights: • Nitrogen mustard-induced MGMT-DNA cross-linking was detected in a living cell. • Concentration- and time-dependent manners of MGMT-DNA cross-linking were revealed. • Proteolysis played an important role in protein (MGMT)-DNA cross-linking repair. • DVC1 acts as a proteolytic enzyme in cross-linking repair in a p

Protective effect of C-peptide on experimentally induced diabetic nephropathy and the possible link between C-peptide and nitric oxide.

Science.gov (United States)

Elbassuoni, Eman A; Aziz, Neven M; El-Tahawy, Nashwa F

2018-06-01

Diabetic nephropathy one of the major microvascular diabetic complications. Besides hyperglycemia, other factors contribute to the development of diabetic complications as the proinsulin connecting peptide, C-peptide. We described the role of C-peptide replacement therapy on experimentally induced diabetic nephropathy, and its potential mechanisms of action by studying the role of nitric oxide (NO) as a mediator of C-peptide effects by in vivo modulating its production by N G -nitro-l-arginine methyl ester (L-NAME). Renal injury markers measured were serum urea, creatinine, tumor necrosis factor alpha, and angiotensin II, and malondialdehyde, total antioxidant, Bcl-2, and NO in renal tissue. In conclusion, diabetic induction resulted in islet degenerations and decreased insulin secretion with its metabolic consequences and subsequent renal complications. C-Peptide deficiencies in diabetes might have contributed to the metabolic and renal error, since C-peptide treatment to the diabetic rats completely corrected these errors. The beneficial effects of C-peptide are partially antagonized by L-NAME coadministration, indicating that NO partially mediates C-peptide effects.

Laser-induced corneal cross-linking upon photorefractive ablation with riboflavin

Directory of Open Access Journals (Sweden)

Kornilovskiy IM

2016-04-01

Full Text Available Igor M Kornilovskiy,1 Elmar M Kasimov,2 Ayten I Sultanova,2 Alexander A Burtsev1 1Department of Eye Diseases, Federal State Budgetary Institution “National Pirogov Medical Surgical Centre”, Ministry of Health, Moscow, Russia; 2Department of Eye Diseases, Zarifa Aliyeva National Ophthalmology Center, Ministry of Health, Baku, Azerbaijan Aim: To estimate the biomechanical effect of the laser-induced cross-linking resulting from photorefractive ablation of the cornea with riboflavin.Methods: Excimer laser ablation studies were performed ex vivo (32 eyes of 16 rabbits by phototherapeutic keratectomy (PTK and in vivo (24 eyes of 12 rabbits by transepithelial photorefractive keratectomy (TransPRK, with and without riboflavin saturation of the stroma. Then, we performed corneal optical coherence tomography on 36 eyes of 18 patients with varying degrees of myopia at different times after the TransPRK was performed with riboflavin saturation of the stroma.Results: Biomechanical testing of corneal samples saturated with riboflavin revealed cross-linking effect accompanied by the increase in tensile strength and maximum strength. PTK showed increase in tensile strength from 5.1±1.4 to 7.2±1.6 MPa (P=0.001, while TransPRK showed increase in tensile strength from 8.8±0.9 to 12.8±1.3 MPa (P=0.0004. Maximum strength increased from 8.7±2.5 to 12.0±2.8 N (P=0.005 in PTK and from 12.8±1.6 to 18.3±1.2 N (P=0.0004 in TransPRK. Clinical optical coherence tomography studies of the biomicroscopic transparent cornea at different times after TransPRK showed increased density in the surface layers of the stroma and membrane-like structure beneath the epithelium.Conclusion: Photorefractive ablation of the preliminary corneal stroma saturation with riboflavin causes the effect of laser-induced cross-linking, which is attended with an increase in corneal tensile strength, maximum strength, increased density in the surface layers of the stroma, and formation of

In vitro cross-linking of bovine lens proteins photosensitized by promazines

International Nuclear Information System (INIS)

Merville, M.P.; Decuyper, J.; Piette, J.; Calberg-Bacq, C.M.; Van de Vorst, A.

1984-01-01

Promazine derivatives induce cross-linking of bovine lens crystallins in vitro by irradiation with near-ultraviolet (UV) light in the presence of O 2 , as revealed by electrophoresis after denaturation. With the five derivatives tested (promazine [PZ], chlorpromazine [CPZ], triflupromazine [TFPZ], methoxypromazine [MTPZ], and acepromazine [ACPZ]), single-hit kinetics are observed. Evidence implicating the cation radicals of the PZ derivatives as the causative agent of this in vitro effect is presented. Hydroxyl radicals do not appear to be involved in the photo-cross-linking reaction. Sodium ascorbate protects against damage induced either by PZ derivatives plus light or by PZ cation radicals in the dark. These findings are discussed with respect to development of cataracts induced by these drugs in vivo

Effect of dual modification with hydroxypropylation and cross-linking on physicochemical properties of taro starch.

Science.gov (United States)

Hazarika, Bidyut Jyoti; Sit, Nandan

2016-04-20

Dual modification of taro starch by hydroxypropylation and cross-linking was carried out and the properties of the modified starches were investigated. Two different levels of hydroxypropylation (5 and 10%) and cross-linking (0.05 and 0.10%) were used in different sequences. The amylose contents of the starch decreased due to single and dual modification. For the dual-modified starches, the swelling, solubility and clarity was found to increase with level of hydroxypropylation and decrease with level of cross-linking. The freeze-thaw stability of the dual-modified starches was also affected by the sequence of modification. The viscosities of the cross-linked and dual-modified starches were more than native and hydroxypropylated starches. The firmness of the dual-modified starches was also higher than native and single modified starches. The dual-modified starches have benefits of both type of modifications and could be used for specific purposes e.g. food products requiring high viscosity as well as freeze-thaw stability. Copyright © 2015 Elsevier Ltd. All rights reserved.

Specific cross-linking of capsid proteins to virus RNA by ultraviolet irradiation of polio virus

Energy Technology Data Exchange (ETDEWEB)

Wetz, K.; Habermehl, K.O. (Freie Univ. Berlin (Germany, F.R.))

1982-04-01

Poliovirus was irradiated with u.v. light under conditions causing approx. 5% cross-linking of capsid protein to virus RNA. Cross-linked RNA-protein complexes, freed from unbound protein, were treated with nuclease, and then analysed on SDS-polyacrylamide gels. The smallest capsid polypeptide VP4 was found to be associated with the RNA to the greatest degree, followed by VP2 and VP1, while VP3 was attached only in trace amounts. Low radiation doses, which produced cross-linking of RNA to protein, did not cause breakdown of the virus particles or conformational changes of the capsid as examined physically and serologically. However, higher doses caused structural alterations of the virus capsid.

Specific cross-linking of capsid proteins to virus RNA by ultraviolet irradiation of polio virus

International Nuclear Information System (INIS)

Wetz, K.; Habermehl, K.-O.

1982-01-01

Poliovirus was irradiated with u.v. light under conditions causing approx. 5% cross-linking of capsid protein to virus RNA. Cross-linked RNA-protein complexes, freed from unbound protein, were treated with nuclease, and then analysed on SDS-polyacrylamide gels. The smallest capsid polypeptide VP4 was found to be associated with the RNA to the greatest degree, followed by VP2 and VP1, while VP3 was attached only in trace amounts. Low radiation doses, which produced cross-linking of RNA to protein, did not cause breakdown of the virus particles or conformational changes of the capsid as examined physically and serologically. However, higher doses caused structural alterations of the virus capsid. (author)

Pel is a cationic exopolysaccharide that cross-links extracellular DNA in the Pseudomonas aeruginosa biofilm matrix.

Science.gov (United States)

Jennings, Laura K; Storek, Kelly M; Ledvina, Hannah E; Coulon, Charlène; Marmont, Lindsey S; Sadovskaya, Irina; Secor, Patrick R; Tseng, Boo Shan; Scian, Michele; Filloux, Alain; Wozniak, Daniel J; Howell, P Lynne; Parsek, Matthew R

2015-09-08

Biofilm formation is a complex, ordered process. In the opportunistic pathogen Pseudomonas aeruginosa, Psl and Pel exopolysaccharides and extracellular DNA (eDNA) serve as structural components of the biofilm matrix. Despite intensive study, Pel's chemical structure and spatial localization within mature biofilms remain unknown. Using specialized carbohydrate chemical analyses, we unexpectedly found that Pel is a positively charged exopolysaccharide composed of partially acetylated 1→4 glycosidic linkages of N-acetylgalactosamine and N-acetylglucosamine. Guided by the knowledge of Pel's sugar composition, we developed a tool for the direct visualization of Pel in biofilms by combining Pel-specific Wisteria floribunda lectin staining with confocal microscopy. The results indicate that Pel cross-links eDNA in the biofilm stalk via ionic interactions. Our data demonstrate that the cationic charge of Pel is distinct from that of other known P. aeruginosa exopolysaccharides and is instrumental in its ability to interact with other key biofilm matrix components.

Chemical Synthesis of Circular Proteins*

Science.gov (United States)

Tam, James P.; Wong, Clarence T. T.

2012-01-01

Circular proteins, once thought to be rare, are now commonly found in plants. Their chemical synthesis, once thought to be difficult, is now readily achievable. The enabling methodology is largely due to the advances in entropic chemical ligation to overcome the entropy barrier in coupling the N- and C-terminal ends of large peptide segments for either intermolecular ligation or intramolecular ligation in end-to-end cyclization. Key elements of an entropic chemical ligation consist of a chemoselective capture step merging the N and C termini as a covalently linked O/S-ester intermediate to permit the subsequent step of an intramolecular O/S-N acyl shift to form an amide. Many ligation methods exploit the supernucleophilicity of a thiol side chain at the N terminus for the capture reaction, which makes cysteine-rich peptides ideal candidates for the entropy-driven macrocyclization. Advances in desulfurization and modification of the thiol-containing amino acids at the ligation sites to other amino acids add extra dimensions to the entropy-driven ligation methods. This minireview describes recent advances of entropy-driven ligation to prepare circular proteins with or without a cysteinyl side chain. PMID:22700959

Alzheimer’s disease against peptides products of enzymatic cleavage APP protein: Biological, pathobiological and physico-chemical properties of fibrillating peptides

Directory of Open Access Journals (Sweden)

Małgorzata Marszałek

2017-05-01

Full Text Available Various peptides products of enzymatic cleavage of key for Alzheimer’s disease Amyloid Precursor Protein (APP are well known, but still are matter of scientific debate. The Aβ type products are especially challenging for experimental and medical research. This paper outlines several, still poorly known, biological and medical processes such as peptides biology, i.e., formation, biodistribution, translocation, transport and finally removal from brain compartments and body fluids like Intracellular Fluid (ICF, Cerebrospinal Fluid (CSF, Interstitial Fluid (ISF, blood serum or urine. In addition, the following studies concerning AD patients might prove challenging and simultaneously promising: peptides translocation through Blood-Brain – Barrier (BBB and Blood–Cerebrospinal Fluid Barrier (BCSFB and their removal from the brain according to a new concept of glymphatic system; – diagnostic difficulties that stem from physico-chemical properties and the nature of proteins or fibrillating peptides itself like low concentration, short half-live and from experimental-technical problems as well like high adsorption or low solubility of Aβ, tau or amylin. The study of diagnostic parameters is very important, as it may better reflect early changes before the disease develops; one such parameter is the Aβ42/Aβ40 ratio, or the ratio with the total tau concentration combination and other new biomarkers like Aβ1-38; other factors include oxidative stress and inflammation process proteins, complement factor H, alpha-2-macroglobulin, or clusterin. The study of various forms of pathological amyloid deposits that emerge in different but specific brain regions AD patients seems to be crucial as well. The composition of the first initial pathological, pre-fibrillating monomers of fibrillating peptides and their role in AD development and disease progression have been described as well. They are even more challenging for science and simultaneously might be

Improved PET Imaging of uPAR expression using new Cu-64-labeled cross-bridged peptide ligands

DEFF Research Database (Denmark)

Persson, Morten; Hosseini, Masood; Madsen, Jacob

2013-01-01

The correlation between uPAR expression, cancer cell invasion and metastases is now well-established and has prompted the development of a number of uPAR PET imaging agents, which could potentially identify cancer patients with invasive and metastatic lesions. In the present study, we synthesized......, the more stable of the new uPAR PET tracers, (64)Cu-CB-TE2A-PA-AE105, exhibits a significantly reduced liver uptake compared to (64)Cu-DOTA-AE105 as well as (64)Cu-CB-TE2A-AE105, (p...... and characterized two new cross-bridged (64)Cu-labeled peptide conjugates for PET imaging of uPAR and performed a head-to-head comparison with the corresponding and more conventionally used DOTA conjugate. Based on in-source laser-induced reduction of chelated Cu(II) to Cu(I), we now demonstrate the following...... ranking with respect to the chemical inertness of their complexed Cu ions: DOTA-AE105 95%) were achieved in all cases by incubation at 95ºC. In vivo, they display identical tumor uptake after 1h, but differ significantly after 22 hrs, where the DOTA-AE105 uptake remains surprisingly high. Importantly...

Characterization of radiation-cross-linked, high-density polyethylene for thermal energy storage

International Nuclear Information System (INIS)

Whitaker, R.B.; Craven, S.M.; Etter, D.E.; Jendrek, E.F.; Nease, A.B.

1983-01-01