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Protein denaturation

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https://www.readbyqxmd.com/read/28231319/development-of-a-keratinase-activity-assay-using-recombinant-chicken-feather-keratin-substrates
#1
Hyeon-Su Jin, Seon Yeong Park, Kyungmin Kim, Yong-Jik Lee, Gae-Won Nam, Nam Joo Kang, Dong-Woo Lee
Poultry feathers consist mainly of the protein keratin, which is rich in β-pleated sheets and consequently resistant to proteolysis. Although many keratinases have been identified, the reasons for their substrate specificity towards β-keratin remain unclear due to difficulties in preparing a soluble feather keratin substrate for use in activity assays. In the present study, we overexpressed Gallus gallus chromosomes 2 and 27 β-keratin-encoding genes in Escherichia coli, purified denatured recombinant proteins by Ni2+ affinity chromatography, and refolded by stepwise dialysis to yield soluble keratins...
2017: PloS One
https://www.readbyqxmd.com/read/28224610/structural-changes-in-cartilage-and-collagen-studied-by-high-temperature-raman-spectroscopy
#2
Mark Fields, Nicholas Spencer, Jayesh Dudhia, Paul F McMillan
Understanding the high temperature behavior of collagen and collagenous tissue is important for surgical procedures and biomaterials processing for the food, pharmaceutical and cosmetics industries. One primary event for proteins is thermal denaturation that involves unfolding the polypeptide chains while maintaining the primary structure intact. Collagen in the extracellular matrix of cartilage and other connective tissue is a hierarchical material containing bundles of triple-helical fibers associated with water and proteoglycan components...
February 22, 2017: Biopolymers
https://www.readbyqxmd.com/read/28224394/native-like-and-denatured-cytochrome-c-ions-yield-cation-to-anion-proton-transfer-reaction-products-with-similar-collision-cross-sections
#3
Kenneth J Laszlo, John H Buckner, Eleanor B Munger, Matthew F Bush
The relationship between structures of protein ions, their charge states, and their original structures prior to ionization remains challenging to decouple. Here, we use cation-to-anion proton transfer reactions (CAPTR) to reduce the charge states of cytochrome c ions in the gas phase, and ion mobility to probe their structures. Ions were formed using a new temperature-controlled nanoelectrospray ionization source at 25 °C. Characterization of this source demonstrates that the temperature of the liquid sample is decoupled from that of the atmospheric pressure interface, which is heated during CAPTR experiments...
February 21, 2017: Journal of the American Society for Mass Spectrometry
https://www.readbyqxmd.com/read/28223518/integrated-view-of-internal-friction-in-unfolded-proteins-from-single-molecule-fret-contact-quenching-theory-and-simulations
#4
Andrea Soranno, Andrea Holla, Fabian Dingfelder, Daniel Nettels, Dmitrii E Makarov, Benjamin Schuler
Internal friction is an important contribution to protein dynamics at all stages along the folding reaction. Even in unfolded and intrinsically disordered proteins, internal friction has a large influence, as demonstrated with several experimental techniques and in simulations. However, these methods probe different facets of internal friction and have been applied to disparate molecular systems, raising questions regarding the compatibility of the results. To obtain an integrated view, we apply here the combination of two complementary experimental techniques, simulations, and theory to the same system: unfolded protein L...
February 21, 2017: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/28223313/active-multi-enzyme-assemblies-for-long-chain-olefinic-hydrocarbon-biosynthesis
#5
James K Christenson, Matthew R Jensen, Brandon R Goblirsch, Fatuma Mohamed, Wei Zhang, Carrie M Wilmot, Lawrence P Wackett
Bacteria from different Phyla produce long-chain olefinic hydrocarbons derived from an OleA-catalyzed Claisen condensation of two fatty acyl-CoA substrates, followed by reduction and oxygen elimination reactions catalyzed by the proteins OleB, OleC, and OleD. In this report, OleA, OleB, OleC, and OleD were individually purified as soluble proteins, and all were found to be essential for reconstituting hydrocarbon biosynthesis. Recombinant co-expression of tagged OleABCD proteins from Xanthomonas campestris in Escherichia coli and purification over His6x- and FLAG-columns resulted in OleA separating whilst OleBCD purified together, irrespective of which of the four Ole proteins were tagged...
February 21, 2017: Journal of Bacteriology
https://www.readbyqxmd.com/read/28223212/n-butilydenephthalide-exhibits-protection-against-neurotoxicity-through-regulation-of-tryptophan-2-3-dioxygenase-in-spinocerebellar-ataxia-type-3
#6
Karthyayani Rajamani, Jen-Wei Liu, Cheng-Han Wu, I-Tsang Chiang, Deng-Huwei You, Si-Yin Lin, Dean-Kuo Hsieh, Shinn-Zong Lin, Horng-Jyh Harn, Tzyy-Wen Chiou
Spinocerebellar ataxia type 3 or Machado-Joseph disease (SCA3/MJD) is characterized by the repetition of a CAG codon in the ataxin-3 gene (ATXN3), which leads to the formation of an elongated mutant ATXN3 protein that can neither be denatured nor undergo proteolysis in the normal manner. This abnormal proteolysis leads to the accumulation of cleaved fragments, which have been identified as toxic and further they act as a seed for more aggregate formation, thereby increasing toxicity in neuronal cells. To date, there have been few studies or treatment strategies that have focused on controlling toxic fragment formation...
February 18, 2017: Neuropharmacology
https://www.readbyqxmd.com/read/28219688/mesoporous-phenylalanine-ammonia-lyase-microspheres-with-improved-stability-through-calcium-carbonate-templating
#7
Jiandong Cui, Yamin Zhao, Zhilei Tan, Cheng Zhong, Peipei Han, Shiru Jia
Cross-linked enzyme aggregates (CLEAs) have recently emerged as a promising method for enzyme immobilization due to its simplicity and low cost. However, a lack of good size and morphological control over the as-prepared CLEAs has limited their practical applications in some cases. Here, monodisperse spherical CLEAs of phenylalanine ammonia lyase (PAL microspheres) were prepared based on CaCO3 microtemplates. The preparation procedure involves filling porous CaCO3 microtemplates with the protein by salt precipitation, glutaraldehyde crosslinking, and dissolution of the microtemplates...
February 17, 2017: International Journal of Biological Macromolecules
https://www.readbyqxmd.com/read/28214695/the-effects-of-atmospheric-pressure-cold-plasma-treatment-on-microbiological-physical-chemical-and-sensory-characteristics-of-vacuum-packaged-beef-loin
#8
A Bauer, Y Ni, S Bauer, P Paulsen, M Modic, J L Walsh, F J M Smulders
Effects on vacuum packaged and non-packaged beef longissimus samples exposed to atmospheric cold plasma (ACP) generated at different powers were studied over a 10day period of vacuum-, and a subsequent 3day period of aerobic storage. Exposure of non-covered beef samples under high power ACP conditions resulted in increased a*, b*, Chroma and Hue values, but ACP treatment of packaged loins did not impact colour (L*, a*, b*, Chroma, Hue), lipid peroxidation, sarcoplasmic protein denaturation, nitrate/nitrite uptake, or myoglobin isoform distribution...
February 8, 2017: Meat Science
https://www.readbyqxmd.com/read/28214340/molecular-tiling-on-the-surface-of-a-bacterial-spore-the-exosporium-of-the-bacillus-anthracis-cereus-thuringiensis-group
#9
Cassandra Terry, Shuo Jiang, David S Radford, Wan Qiang, Svetomir Tzokov, Anne Moir, Per A Bullough
Bacteria of the genera Bacillus and Clostridium form highly resistant spores, which in the case of some pathogens act as the infectious agents. An exosporium forms the outermost layer of some spores; it plays roles in protection, adhesion, dissemination, host targeting in pathogens, and germination control. The exosporium of the Bacillus cereus group, including the anthrax pathogen, contains a 2D-crystalline basal layer, overlaid by a hairy nap. BclA and related proteins form the hairy nap, and require ExsFA (BxpB) for their localisation on the basal layer...
February 18, 2017: Molecular Microbiology
https://www.readbyqxmd.com/read/28213141/thermostability-of-photosystem-i-trimers-and-monomers-from-the-cyanobacterium-thermosynechococcus-elongatus
#10
Vladimir V Shubin, Irina V Terekhova, Yulia V Bolychevtseva, Eithar El-Mohsnawy, Matthias Rögner, Werner Mäntele, Marta J Kopczak, Enela Džafić
The performance of solar energy conversion into alternative energy sources in artificial systems highly depends on the thermostability of photosystem I (PSI) complexes Terasaki et al. (2007), Iwuchukwu et al. (2010), Kothe et al. (2013) . To assess the thermostability of PSI complexes from the thermophilic cyanobacterium Thermosynechococcus elongatus heating induced perturbations on the level of secondary structure of the proteins were studied. Changes were monitored by Fourier transform infrared (FT-IR) spectra in the mid-IR region upon slow heating (1°C per minute) of samples in D2O phosphate buffer (pD 7...
February 6, 2017: Spectrochimica Acta. Part A, Molecular and Biomolecular Spectroscopy
https://www.readbyqxmd.com/read/28203642/epitopes-within-recombinant-%C3%AE-actinin-protein-is-serodiagnostic-target-for-trichomonas-vaginalis-sexually-transmitted-infections
#11
J F Alderete
To date there is no commercially-available serodiagnostic for women and men infected with Trichomonas vaginalis. Thirteen epitopes of the immunogenic T. vaginalis α-actinin (106.2-kDa) are detected by sera of women patients, and 5 epitopes, a subset of the 13, are detected by sera of men. A truncated recombinant protein called ACT-P2 (63.5-kDa) encoding the 5 epitopes is used for screening by ELISA for antibody in sera of women and men. Because ACT-P2 is poorly expressed in E. coli, we wanted alternative recombinant α-actinin proteins as serodiagnostic targets...
January 2017: Heliyon
https://www.readbyqxmd.com/read/28199087/expression-purification-and-properties-of-a-human-arachidonoyl-specific-isoform-of-diacylglycerol-kinase
#12
William Jennings, Sejal Doshi, Prasanta Kumar Hota, Aaron Prodeus, Stephanie Black, Richard M Epand
Diacylglycerol kinase ε (DGKε) catalyzes the phosphorylation of diacylglycerol, producing phosphatidic acid. DGKε demonstrates exquisite specificity for the acyl chains of diacylglycerol. This contributes to the enrichment of particular acyl chains within the lipids of the phosphatidylinositol cycle. Phosphatidylinositol is highly enriched with 1-stearoyl-2-arachidonoyl, which is important for maintaining cellular health. Dysregulation of DGKε perturbs lipid signaling and biosynthesis, which has been linked to epilepsy, Huntington's disease, and heart disease...
February 24, 2017: Biochemistry
https://www.readbyqxmd.com/read/28194738/collidoscope-an-improved-tool-for-computing-collisional-cross-sections-with-the-trajectory-method
#13
Simon A Ewing, Micah T Donor, Jesse W Wilson, James S Prell
Ion mobility-mass spectrometry (IM-MS) can be a powerful tool for determining structural information about ions in the gas phase, from small covalent analytes to large, native-like or denatured proteins and complexes. For large biomolecular ions, which may have a wide variety of possible gas-phase conformations and multiple charge sites, quantitative, physically explicit modeling of collisional cross sections (CCSs) for comparison to IMS data can be challenging and time-consuming. We present a "trajectory method" (TM) based CCS calculator, named "Collidoscope," which utilizes parallel processing and optimized trajectory sampling, and implements both He and N2 as collision gas options...
February 13, 2017: Journal of the American Society for Mass Spectrometry
https://www.readbyqxmd.com/read/28193489/what-can-we-get-from-varying-scan-rate-in-protein-differential-scanning-calorimetry
#14
Mojtaba Amani, Ali A Moosavi-Movahedi, Boris I Kurganov
Differential scanning calorimetry has many advantages over other techniques to study the thermal stability of proteins due to its direct measurement of thermodynamic parameters. Most proteins undergo irreversible thermal denaturation causing their thermogram to be scan rate dependent. We modeled reversible and irreversible protein thermograms at varying scan rates. The complete Lumry-Eyring model was used to model the irreversible thermograms at various values of T1/2 (temperature at which equilibrium constant equals unity) and T* (temperature at which rate constant equals 1min(-1))...
February 10, 2017: International Journal of Biological Macromolecules
https://www.readbyqxmd.com/read/28193434/the-effect-of-non-standard-heat-treatment-of-sheep-s-milk-on-physico-chemical-properties-sensory-characteristics-and-the-bacterial-viability-of-classical-and-probiotic-yogurt
#15
Šimun Zamberlin, Dubravka Samaržija
Classical and probiotic set yogurt were made using non-standard heat treatment of sheep's milk at 60°C/5min. Physico-chemical properties, sensory characteristics, and the viability of bacteria that originated from cultures in classical and probiotic yogurt were analysed during 21days of storage at 4°C. For the production of yogurt, a standard yogurt culture and a probiotic strain Lactobacillus rhamnosus GG were used. At the end of storage time of the classical and probiotic yogurt the totals of non-denatured whey proteins were 92...
June 15, 2017: Food Chemistry
https://www.readbyqxmd.com/read/28192906/coupling-high-throughput-microfluidics-and-small-angle-x-ray-scattering-to-study-protein-crystallization-from-solution
#16
Nhat Pham, Dimitri Radajewski, Adam Round, Martha Brennich, Petra Pernot, Béatrice Biscans, Françoise Bonneté, Sébastien Teychené
In this work, we propose the combination of small-angle X-ray scattering (SAXS) and high throughput, droplet based microfluidics as a powerful tool to investigate macromolecular interactions, directly related to protein solubility. For this purpose, a robust and low cost microfluidic platform was fabricated for achieving the mixing of proteins, crystallization reagents, and buffer in nanoliter volumes and the subsequent generation of nanodroplets by means of a two phase flow. The protein samples are compartmentalized inside droplets, each one acting as an isolated microreactor...
February 8, 2017: Analytical Chemistry
https://www.readbyqxmd.com/read/28192141/glossoscolex-paulistus-hemoglobin-with-fluorescein-isothiocyanate-steady-state-and-time-resolved-fluorescence
#17
Ana E B Barros, Marina B Barioni, Francisco A O Carvalho, Amando Siuiti Ito, Marcel Tabak
Glossoscolex paulistus extracellular hemoglobin (HbGp) stability has been followed, in the presence of urea, using fluorescein isothiocyanate (FITC). Binding of FITC to HbGp results in a significant quenching of probe fluorescence. Tryptophan emission decays present four characteristic lifetimes: two in the sub-nanosecond/picosecond, and two in the nanosecond time ranges. Tryptophan decays for pure HbGp and HbGp-FITC systems are similar. In the absence of denaturant, and up to 2.5mol/L of urea, the shorter lifetimes predominate...
February 10, 2017: International Journal of Biological Macromolecules
https://www.readbyqxmd.com/read/28187517/selective-cysteine-modification-of-metal-free-human-metallothionein-1a-and-its-isolated-domain-fragments-solution-structural-properties-revealed-via-esi-ms
#18
Gordon W Irvine, Melissa Santolini, Martin J Stillman
Human metallothionein 1a, a protein with two cysteine-rich metal-binding domains (α with 11 Cys and β with 9), was analyzed in its metal-free form by selective, covalent Cys modification coupled with ESI-MS techniques. The modification profiles of the isolated β- and α-fragments reacted with p-benzoquinone (Bq), N-ethylmalemide (NEM) and iodoacetamide (IAM) were compared with the full length protein using ESI-mass spectral data to follow the reaction pathway. Under denaturing conditions at low pH, the reaction profile with each modifier followed pathways that resulted in stochastic, Normal distributions of species whose maxima was equal to the mol...
February 10, 2017: Protein Science: a Publication of the Protein Society
https://www.readbyqxmd.com/read/28185459/soy-soluble-polysaccharide-as-a-nanocarrier-for-curcumin
#19
Fei-Ping Chen, Shiyi Ou, Zhong Chen, Chuan-He Tang
The complexation between soy soluble polysaccharide (SSPS) and curcumin at pH 7.0 and 4.0, as well as some physicochemical characteristics of the resultant complexes were investigated. The encapsulation efficiency and loading amount of curcumin in the complexes at pH 4.0 reached 67.3% and 4.49 g/mg SSPS, respectively. Ethanol-induced denaturation and structural unfolding of the protein fraction in SSPS was essential for the complex formation. The complexation with curcumin resulted in aggregation of SSPS, and subsequently formation of compacted nanoparticles with curcumin as the core...
February 10, 2017: Journal of Agricultural and Food Chemistry
https://www.readbyqxmd.com/read/28185187/preservation-of-specific-protein-signaling-states-using-heat-based-stabilizor-system
#20
Mats Borén
The ability to adequately measure the phosphorylation state of a protein has major biological as well as clinical relevance. Due to its variable nature, reversible protein phosphorylations are sensitive to changes in the tissue environment. Stabilizor(TM) T1 is a system for rapid inactivation of enzymatic activity in biological samples. Enzyme inactivation is accomplished using thermal denaturation in a rapid, homogeneous, and reproducible fashion without the need of added chemical inhibitors. Using pCREB(Ser133) as a model system, the applicability of the Stabilizor system to preserve a rapidly lost phosphorylation is shown...
2017: Methods in Molecular Biology
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