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https://www.readbyqxmd.com/read/28445847/to-elute-or-not-to-elute-in-immunocapture-bottom-up-lc-ms
#1
Maren Christin Stillesby Levernæs, Marianne Nordlund Broughton, Léon Reubsaet, Trine Grønhaug Halvorsen
Immunocapture-based bottom-up LC-MS is a promising technique for the quantification of low abundant proteins. Magnetic immunocapture beads provide efficient enrichment from complex samples through the highly specific interaction between the target protein and its antibody. In this article, we have performed the first thorough comparison between digestion of proteins while bound to antibody coated beads versus after elution from the beads. Two previously validated immunocapture based MS methods for the quantification of pro-gastrin releasing peptide (ProGRP) and human chorionic gonadotropin (hCG) were used as model systems...
April 14, 2017: Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences
https://www.readbyqxmd.com/read/28445766/inhibitory-effect-of-emulsifiers-in-sedds-on-protease-activity-just-an-illusion
#2
Noemi Lupo, Una Fidahic, Gergely Hetényi, Janine Griesser, Andreas Bernkop-Schnürch
AIM: Evaluation of inhibitory effect of emulsifiers on pancreatic trypsin and α-chymotrypsin. METHODS: The inhibitory effect of Cremophor EL, Cremophor RH 40, Brij O10, Tween 20, polyethylene glycol 8000, polyethylene glycol 400, Carbitol, Pemulen TR-1, Pemulen TR-2, Carbopol Ultrez 20 and Carbopol Ultrez 21 on pancreatic trypsin and α-chymotrypsin was tested. BAEE (Nα-Benzoyl-L-arginine ethyl ester), BTEE (N-Benzoyl-L-tyrosine ethyl ester), casein and insulin were used as substrates for trypsin and α-chymotrypsin...
April 23, 2017: International Journal of Pharmaceutics
https://www.readbyqxmd.com/read/28445094/proteinases-during-early-development-of-the-pacific-whiteleg-shrimp-penaeus-vannamei
#3
Patricia Hernandez-Cortes, Crisalejandra Rivera-Pérez, Fernando García-Carreño, Diana Martínez-Alarcón
During shrimp larval development, changes occur in molecular components. Enzyme activity and mRNA expression of proteinases were assayed in Penaeus vannamei during larval development, which consists of 5 nauplius stages, 3 protozoeal stages, 3 mysis stages, and 12 postlarval stages. Trypsin activity reached a maximum at the beginning of postlarval stages 1 and 2, and significantly decreased in subsequent postlarval stages. Chymotrypsin activity increased at the third protozoeal stage, then significantly decreased in subsequent stages...
February 2017: Biological Bulletin
https://www.readbyqxmd.com/read/28442447/detailed-two-dimensional-gel-proteomic-mapping-of-the-feces-of-the-house-dust-mite-dermatophagoides-pteronyssinus-and-comparison-with-d-farinae-reduced-trypsin-protease-content-in-d-pteronyssinus-and-different-isoforms
#4
Tomas Erban, Karel Harant, Jan Hubert
Major domestic mite allergens are present in feces. We present a detailed 2D-E-MS/MS proteomic analysis of the Dermatophagoides pteronyssinus feces. Precise cultivation yielded a pure fecal extract. We detected differences in fecal allergens/digestive enzymes between D. pteronyssinus and D. farinae using 2D-E fingerprinting, including unique information on species-specific protease isoforms. Proteomic analysis was performed by 2D-E coupled with MALDI-TOF/TOF identification. The species-specific differences in the fecal extracts of the mites were attributed to trypsin-like proteases known as group 3 allergens...
April 22, 2017: Journal of Proteomics
https://www.readbyqxmd.com/read/28441731/polyphenolic-compounds-and-digestive-enzymes-in-vitro-non-covalent-interactions
#5
REVIEW
Alejandra I Martinez-Gonzalez, Ángel G Díaz-Sánchez, Laura A de la Rosa, Claudia L Vargas-Requena, Ismael Bustos-Jaimes, And Emilio Alvarez-Parrilla
The digestive enzymes-polyphenolic compounds (PCs) interactions behind the inhibition of these enzymes have not been completely studied. The existing studies have mainly analyzed polyphenolic extracts and reported inhibition percentages of catalytic activities determined by UV-Vis spectroscopy techniques. Recently, pure PCs and new methods such as isothermal titration calorimetry and circular dichroism have been applied to describe these interactions. The present review focuses on PCs structural characteristics behind the inhibition of digestive enzymes, and progress of the used methods...
April 22, 2017: Molecules: a Journal of Synthetic Chemistry and Natural Product Chemistry
https://www.readbyqxmd.com/read/28439086/quality-and-freshness-of-human-bone-marrow-derived-mesenchymal-stem-cells-decrease-over-time-after-trypsinization-and-storage-in-phosphate-buffered-saline
#6
Tae Hwan Shin, Seungah Lee, Ki Ryung Choi, Da Yeon Lee, Yongman Kim, Man Jeong Paik, Chan Seo, Seok Kang, Moon Suk Jin, Tae Hyeon Yoo, Seong Ho Kang, Gwang Lee
Human bone marrow-derived mesenchymal stem cells (hBM-MSCs) have been studied for their therapeutic potential. However, evaluating the quality of hBM-MSCs before transplantation remains a challenge. We addressed this issue in the present study by investigating deformation, the expression of genes related to reactive oxygen species (ROS) generation, changes in amino acid profiles, and membrane fluidity in hBM-MSCs. Deformability and cell size were decreased after storage for 6 and 12 h, respectively, in phosphate-buffered saline...
April 24, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28437072/the-single-disulfide-directed-%C3%AE-hairpin-fold-dynamics-stability-and-engineering
#7
Balasubramanyam Chittoor, Krishnarjuna Bankala, Rodrigo A W Morales, Christopher Andrew MacRaild, Maiada Sadek, Eleanor Ww Leung, Samuel Robinson, Michael Pennington, Raymond S Norton
Grafting bioactive peptide sequences onto small cysteine-rich scaffolds is a promising strategy to enhance their stability and value in novel peptide-based therapeutics. However, correctly folded disulfide-rich peptides can be challenging to produce by either recombinant or synthetic means. The single disulfide-directed β-hairpin (SDH) fold, first observed in contryphan-Vc1, provides a potential alternative to complex disulfide-rich scaffolds. We have undertaken recombinant production of full-length contryphan-Vc1 (rCon-Vc1[Z1Q]) and a truncated analog (rCon-Vc11-22[Z1Q]), analyzed the backbone dynamics of rCon-Vc1[Z1Q], and probed the conformational and proteolytic stability of these peptides to evaluate the potential of contryphan-Vc1 as a molecular scaffold...
April 24, 2017: Biochemistry
https://www.readbyqxmd.com/read/28436273/proteome-analysis-of-sheep-b-lymphocytes-in-the-course-of-bovine-leukemia-virus-induced-leukemia
#8
Michal Reichert
Presented are the results of a study of the expression pattern of different proteins in the course of bovine leukemia virus-induced leukemia in experimental sheep and I discuss how the obtained data may be useful in gaining a better understanding of the pathogenesis of the disease, diagnosis, and for the selection of possible therapeutic targets. In cattle, the disease is characterized by life-long persistent lymphocytosis leading to leukemia/lymphoma in about 5% of infected animals. In sheep, as opposed to cattle, the course of the disease is always fatal and clinical symptoms usually occur within a three-year period after infection...
January 1, 2017: Experimental Biology and Medicine
https://www.readbyqxmd.com/read/28434814/novel-protein-biomarkers-associated-with-coronary-artery-disease-in-statin-treated-patients-with-familial-hypercholesterolemia
#9
Sven Bos, Michael Phillips, Gerald F Watts, Adrie J M Verhoeven, Eric J G Sijbrands, Natalie C Ward
BACKGROUND: Familial hypercholesterolemia (FH) is the most common and serious monogenic disorder of lipid metabolism. The incidence of coronary artery disease (CAD) varies among both treated and untreated FH patients. OBJECTIVE: The aim of the study was to utilize proteomics to identify novel protein biomarkers that differentiate genetically confirmed heterozygous patients with FH at high CAD risk from those at low CAD risk. METHODS: Sixty genetically confirmed FH patients were recruited and stratified into (1) asymptomatic FH with low atherosclerotic burden (FH, n = 20); (2) asymptomatic FH with high atherosclerotic burden (FH + Ca, n = 20); and (3) FH with previously confirmed symptomatic CAD (FH + CAD, n = 20)...
April 4, 2017: Journal of Clinical Lipidology
https://www.readbyqxmd.com/read/28434171/peroxidatic-cysteine-residue-of-peroxiredoxin-2-separated-from-human-red-blood-cells-treated-by-tert-butyl-hydroperoxide-is-hyperoxidized-into-sulfinic-and-sulfonic-acids
#10
Yo-Ichi Ishida, Mariko Aki, Sohta Fujiwara, Masami Nagahama, Yuki Ogasawara
Peroxiredoxin 2 (Prx2) is a redox enzyme that is abundantly expressed in red blood cells (RBCs) and has been the focus of clinical attention for monitoring the oxidative status. We previously developed a method to quantify the reduced and hyperoxidized forms of Prx2 in human RBCs using reverse-phase high-performance liquid chromatography (HPLC). In the present study, we investigated the hyperoxidative status of Prx2 at the molecular level in a post-translational modification analysis using a liquid chromatography-tandem mass spectrometry (LC-MS/MS) system...
April 22, 2017: Human Cell
https://www.readbyqxmd.com/read/28433552/roscovitine-attenuates-intimal-hyperplasia-via-inhibiting-nf-%C3%AE%C2%BAb-and-stat3-activation-induced-by-tnf-%C3%AE-in-vascular-smooth-muscle-cells
#11
Ming He, Chao Wang, Jia-Huan Sun, Yu Liu, Hong Wang, Jing-Shan Zhao, Yun-Feng Li, Hong Chang, Jian-Ming Hou, Jun-Na Song, Ai-Ying Li, En-Sheng Ji
Roscovitine is a selective CDK inhibitor originally designed as anti-cancer agent, which has also been shown to inhibit proliferation in vascular smooth muscle cells (VSMCs). However, its effect on vascular remodeling and its mechanism of action remain unknown. In our study, we created a new intimal hyperplasia model in male Sprague-Dawley rats by trypsin digestion method, which cause to vascular injury as well as the model of rat carotid balloon angioplasty. Roscovitine administration led to a significant reduction in neointimal formation and VSMCs proliferation after injury in rats...
April 19, 2017: Biochemical Pharmacology
https://www.readbyqxmd.com/read/28432272/establishment-of-an-in-vitro-culture-model-of-theca-cells-from-hierarchical-follicles-in-ducks
#12
Xiang Gan, da Chen, Yan Deng, Junsong Yuan, Bo Kang, Jiamin Qiu, Wenqiang Sun, Chunchun Han, Jiwei Hu, Liang Li, Jiwen Wang
Theca cells, including theca interna cells and theca externa cells, are vital components of ovarian follicles. The aim of this study is to identify a reliable method for the in vitro culture of theca cells from duck ovarian hierarchical (F4-F2) follicles. We improved the method for cell separation by using trypsin to further remove granular cells, and we increased the concentration of fetal bovine serum used in in vitro culture to improve cytoactivity. Cell antibody immunofluorescence showed that all inoculated cells could be stained by the CYP17A1/19A1 antibody but not by the FSHR antibody, which could stain granulosa cells...
April 21, 2017: Bioscience Reports
https://www.readbyqxmd.com/read/28426228/ultrafast-dynamics-of-a-triazene-excited-state-pathways-and-the-impact-of-binding-to-the-minor-groove-of-dna-and-further-biomolecular-systems
#13
Lena Grimmelsmann, Alireza Marefat Khah, Christian Spies, Christof Hättig, Patrick Nuernberger
Many synthetic DNA minor groove binders exhibit a strong increase in fluorescence when bound to DNA. The pharmaceutical-relevant berenil (diminazene aceturate) is an exception with an extremely low fluorescence quantum yield (on the order of 10(-4)). We investigate the ultrafast excited-state dynamics of this triazene by femtosecond time-resolved fluorescence experiments in water, ethylene glycol, and buffer and bound to the enzyme β-trypsin, the minor groove of AT-rich DNA, and G-quadruplex DNA. Ab initio calculations provide additional mechanistic insight...
April 20, 2017: Journal of Physical Chemistry Letters
https://www.readbyqxmd.com/read/28426042/antibodies-to-h2a-and-h2b-histones-from-the-sera-of-hiv-infected-patients-catalyze-site-specific-degradation-of-these-histones
#14
Svetlana V Baranova, Pavel S Dmitrienok, Nikita V Ivanisenko, Valentina N Buneva, Georgy A Nevinsky
Histones and their post-translational modifications have key roles in chromatin remodeling and gene transcription. Besides intranuclear functions, histones act as damage-associated molecules when they are released into the extracellular space. Administration of histones to animals leads to systemic inflammatory and toxic responses. Autoantibodies with enzymatic activities (abzymes) are distinctive features of some autoimmune and viral diseases. Electrophoretically homogeneous IgGs containing no canonical enzymes were isolated from the sera of HIV-infected patients by chromatography on several affinity sorbents including anti-histone Sepharose...
April 20, 2017: Molecular BioSystems
https://www.readbyqxmd.com/read/28425100/mass-spectrometry-analysis-and-in-silico-prediction-of-allergenicity-of-peptides-in-tryptic-hydrolysates-of-the-proteins-from-ruditapes-philippinarum
#15
Yue Yu, Hongwei Liu, Maolin Tu, Meiling Qiao, Zhenyu Wang, Ming Du
BACKGROUND: The Ruditapes philippinarum is nutrient-rich and widely-distributed, but little attentions had been paid on the identification and characterization of bioactive peptides in the bivalve. In this study, we evaluated the peptides of the Ruditapes philippinarum enzymolysised by trypsin using a combination of UPLC separation and a ESI-Q-TOF-MS/MS spectrometer, followed by data processing, sequence-similarity database searching. The potential allergenicity of the peptides was assessed in silico...
April 20, 2017: Journal of the Science of Food and Agriculture
https://www.readbyqxmd.com/read/28419926/stability-and-activity-of-immobilized-trypsin-on-carboxymethyl-chitosan-functionalized-magnetic-nanoparticles-cross-linked-with-carbodiimide-and-glutaraldehyde
#16
Jun Sun, Lin Yang, Mengmeng Jiang, Yu Shi, Bin Xu, Hai-le Ma
Enzyme cross-linkers, such as 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and glutaraldehyde (GA), have been used to improve the stability of immobilized enzymes. We have developed a relatively stable and high-activity immobilized trypsin through EDC and GA cross-linking. Carboxymethyl chitosan (CM-CTS)-functionalized magnetic nanoparticles (Fe3O4@CM-CTS) were prepared, and characterized by Fourier transform infrared spectroscopy (FTIR), thermogravimetric analysis, X-ray diffraction, and transmission electron microscopy...
April 10, 2017: Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences
https://www.readbyqxmd.com/read/28418608/methods-for-the-isolation-and-3d-culture-of-dermal-papilla-cells-from-human-hair-follicles
#17
H Topouzi, N J Logan, G Williams, C A Higgins
The dermal papilla is a cluster of mesenchymal cells located at the base of the hair follicle which have a number of important roles in the regulation of hair growth. As a consequence, in vitro models of these cells are widely used to study the molecular mechanisms which underlie hair follicle induction, growth and maintenance. While dermal papilla from rodent hair follicles can be digested prior to cell isolation, the unique extracellular matrix composition found in human dermal papilla renders enzymes such as trypsin and collagenase insufficient for digestion of the dermal papilla into a single cell suspension...
April 18, 2017: Experimental Dermatology
https://www.readbyqxmd.com/read/28416397/monitoring-thermal-and-chemical-unfolding-of-brugia-malayi-calreticulin-using-fluorescence-and-circular-dichroism-spectroscopy
#18
Sunita Yadav, Smita Gupta, Jitendra Kumar Saxena
Calreticulin of Brugia malayi (BmCRT) play very important role in host-parasite interaction. In previous study it was found that BmCRT is responsible for prevention of host classical complement pathway activation via its interaction with first component C1q of the human host. Therefore, BmCRT is an essential protein for parasite survival and an important drug target to fend filariasis. In the present study, we have carried out a systamatic biophysical characterization of BmCRT protein. Unfolding of BmCRT was found to be non-cooperative two-state process in the presence of both denaturant GdmCl and urea...
April 14, 2017: International Journal of Biological Macromolecules
https://www.readbyqxmd.com/read/28416038/the-effects-of-fermentation-and-enzymatic-treatment-of-pea-on-nutrient-digestibility-and-growth-performance-of-broilers
#19
F Goodarzi Boroojeni, M Senz, K Kozłowski, D Boros, M Wisniewska, D Rose, K Männer, J Zentek
The present study examined the impacts of native, fermented or enzymatically treated peas (Pisum sativum L.) inclusion in broiler diets, on growth performance and nutrient digestibility. For the fermentation process, Madonna pea was mixed with water (1/1) containing 2.57×108 Bacillus subtilis (GalliPro®) spores/kg pea and then, incubated for 48 h at 30 °C. For the enzymatic treatment process, the used water for dough production contained three enzymes, AlphaGalTM (α-galactosidase), RONOZYME® ProAct and VP (protease and pectinases respectively - DSM, Switzerland) and the pea dough incubated for 24 h at 30°C...
April 18, 2017: Animal: An International Journal of Animal Bioscience
https://www.readbyqxmd.com/read/28414463/do-fragments-and-crystallization-additives-bind-similarly-to-drug-like-ligands
#20
Malgorzata N Drwal, Célien Jacquemard, Carlos Perez, Jérémy Desaphy, Esther Kellenberger
The success of fragment-based drug design (FBDD) hinges upon the optimization of low molecular weight compounds (MW<300) with weak binding affinities to lead compounds with high affinity and selectivity. Usually, structural information from fragment-protein complexes is used to develop ideas about the binding mode of similar, but drug-like molecules. In this regard, crystallization additives such as cryoprotectants or buffer components, highly abundant in crystal structures, are frequently ignored. Thus, the aim of the study was the investigation of the information present in both protein complexes with fragments as well as additives and how they relate to the binding modes of their drug-like counterparts...
April 17, 2017: Journal of Chemical Information and Modeling
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