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1.
Lieza M. Danan Zhihao Yu Peter J. Ludden Weitao Jia Kevin L. Moore Julie A. Leary 《Journal of the American Society for Mass Spectrometry》2010,21(9):1633-1642
Human tyrosylprotein sulfotransferases catalyze the transfer of a sulfuryl moiety from the universal sulfate donor PAPS to
the hydroxyl substituent of tyrosine residues in proteins and peptides to yield tyrosine sulfated products and PAP. Tyrosine
sulfation occurs in the trans-Golgi network, affecting an estimated 1% of the tyrosine residues in all secreted and membrane-bound
proteins in higher order eukaryotes. In this study, an effective LC-MS-based TPST kinetics assay was developed and utilized
to measure the kinetic properties of human TPST-2 and investigate its catalytic mechanism when G protein-coupled CC-chemokine
receptor 8 (CCR8) peptides were used as acceptor substrates. Through initial rate kinetics, product inhibition studies, and
radioactive-labeling experiments, our data strongly suggest a two-site ping-pong model for TPST-2 action. In this mechanistic
model, the enzyme allows independent binding of substrates to two distinct sites, and involves the formation of a sulfated
enzyme covalent intermediate. Some insights on the important amino acid residues at the catalytic site of TPST-2 and its covalent
intermediate are also presented. To our knowledge, this is the first detailed study of the reaction kinetics and mechanism
reported for human TPST-2 or any other Golgi-resident sulfotransferase. 相似文献
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3.
Danan LM Yu Z Hoffhines AJ Moore KL Leary JA 《Journal of the American Society for Mass Spectrometry》2008,19(10):1459-1466
Protein tyrosine O-sulfation, a widespread post-translational modification, is mediated by two Golgi enzymes, tyrosylprotein
sulfotransferase-1 and-2. These enzymes catalyze the transfer of sulfate from the universal sulfate donor 3′-phosphoadenosine-5′-phosphosulfate
(PAPS) to the hydroxyl group of tyrosine residues to form tyrosine O-sulfate ester and PAP. More than 60 proteins have been identified to be tyrosine sulfated including several G protein-coupled
receptors, such as CC-chemokine receptor 8 (CCR8) that is implicated in allergic inflammation, asthma, and atherogenesis.
However, the kinetic properties of purified tyrosylprotein sulfotransferase (TPST)-1 and −2 have not been previously reported.
Moreover, currently there is no available quantitative TPST assay that can directly monitor individual sulfation of a series
of tyrosine residues, which is present in most known substrates. We chose an MS-approach to address this limitation. In this
study, a liquid chromatography electrospray ionisation mass spectrometry (LC/ESI-MS)-based TPST assay was developed to determine
the kinetic parameters of individual TPSTs and a mixture of both isozymes using CCR8 peptides as substrates that have three
tyrosine residues in series. Our method can differentiate between mono-and disulfated products, and our results show that
the Km,app for the monosulfated substrate was 5-fold less than the nonsulfated substrate. The development of this method is the initial
step in the investigation of kinetic parameters of the sequential tyrosine sulfation of chemokine receptors by TPSTs and in
determining its catalytic mechanism. 相似文献
4.
第十七届全国中学生物理竞赛预赛试题获得中学物理教师的普遍好评.题目灵活,难度适中,既能与中学物理教学相衔接,又能考出考生的能力与水平,而且结合了一些近代物理知识,起到了很好的导向作用. 相似文献
5.
2,3-Diazido-2,3-dideoxy-beta-D-mannopyranoside derivatives were synthesized in order to prepare beta-glycosides of 2,3-diacetamido-2,3-dideoxy-D-mannuronic acid, a rare moiety of bacterial oligosaccharides. A direct glycosyl donor, 4,6-di-O-acetyl-2,3-diazido-2,3-dideoxy-alpha-D-mannopyranosyl bromide, was prepared, and its synthetic capacity was tested in glycosylation reactions. An indirect route was also elaborated: 3-azido-3-deoxy-beta-D-glucopyranosides were converted into beta-D-mannopyranosides. The cis vicinal diazido function successfully tolerated the conditions of mild acidic hydrolysis, tritylation, Jones oxidation, TEMPO oxidation, acetolysis, and bromination with TiBr(4). 相似文献
6.
Synthesis of a trimeric gp120 epitope mimic conjugated to a T-helper peptide to improve antigenicity
Schellinger JG Danan-Leon LM Hoch JA Kassa A Srivastava I Davis D Gervay-Hague J 《Journal of the American Chemical Society》2011,133(10):3230-3233
A fully synthetic trivalent mimotope of gp120 conjugated to pan allelic HLA DR binding epitope was prepared using solid-phase peptide synthesis and optimized copper-catalyzed azide-alkyne cycloaddition. The methodology efficiently provides chemically uniform heteromultimeric peptide constructs with enhanced binding, avidity, and specificity toward an established HIV-neutralizing human antibody, MAb b12. The versatile synthetic strategy serves as a powerful platform for the development of synthetic peptides as potential HIV-1 vaccine candidates. 相似文献
7.
如图所示,光滑绝缘水平面上的M、N两点各放完全相同的金属球A和B,A和B的带电量分别为-2Q和-Q. 相似文献
8.
Mediterranean Journal of Mathematics - This paper represents a continuation of Barboteu et al. (Analysis of a contact problem with unilateral constraint and slip-dependent friction, Math... 相似文献
9.
层状盐岩力学特性研究进展 总被引:31,自引:0,他引:31
盐岩具有孔隙度低,渗透率小,损伤自恢复和塑性变形能力大等特性.本文着眼于深部盐岩能源地下储备,在简要介绍国内外盐岩力学特性研究现状基础上,着重综述了近期关于层状沉积型盐岩工程力学特性研究的进展.层状盐岩压缩试验表明泥质硬石膏夹层对盐岩体的变形和破坏特性有明显的影响,揭示了复合岩体的变形和破坏机制;层状盐岩流变试验表明层状盐岩的蠕变主要由盐岩层控制,硬夹层的存在对溶腔围岩的长期蠕变起到抑制作用;为了深入了解界面的力学特性,开展了层状盐岩直接剪切试验、间接拉伸试验和界面微观分析试验,试验表明层状盐岩体中盐岩夹层界面具有较强的黏结力,这十分有利于盐岩溶腔的密闭性和稳定性;针对特殊的层状岩体提出了复合岩体Cosserat介质扩展本构模型,并应用于层状盐岩构造中能源储库稳定性分析中.最后指出了层状盐岩工程力学特性研究中值得关注的几个重点研究方向. 相似文献
10.
高效液相色谱法同时测定化妆品中七种性激素 总被引:17,自引:0,他引:17
建立了化妆品中7种性激素(雌二醇、雌三醇、雌酮、睾酮、甲基睾酮、孕酮、己烯雌酚)同时测定的高效液相色谱法。先在试样中加入20 g/L氢氧化钠溶液与油脂进行皂化反应,然后用二氯甲烷-乙酸乙酯(体积比为40∶1)混合液在酸性条件下(pH 3)萃取,选择XTerraTMRP18色谱柱,以水-甲醇-乙腈(体积比为50∶32∶18)混合液为流动相,在波长230 nm处检测。7种性激素分离良好并排除了样品中杂质峰的干扰,低、高浓度平均回收率范围为75.6%~97.8%;相对标准偏差为1.9%~7.2%;检出限为3.7 相似文献