Free energy study of the catalytic mechanism of Trypanosoma cruzi trans-sialidase. From the Michaelis complex to the covalent intermediate.
Free energy study of the catalytic mechanism of Trypanosoma cruzi trans-sialidase. From the Michaelis complex to the covalent intermediate.
复制标题
DOI：
10.1021/bi2009618
复制
发表时间：
2011-11-22
期刊：
影响因子：
2.9
通讯作者：
Roitberg, Adrian E.
中科院分区：
文献类型：
--
作者：
Pierdominici-Sottile, Gustavo;Horenstein, Nicole A.;Roitberg, Adrian E.
克氏锥虫转唾液酸酶（TcTS）是克氏锥虫感染的关键酶，克氏锥虫是导致人类恰加斯病的原生动物。它催化唾液酸从宿主的糖缀合物转移到寄生虫的糖缀合物。基于动力学同位素效应（KIE）研究，可以确定在过渡态有强亲核参与；最近，精心设计的实验使用2 - 脱氧 - 2,3 - 二氟唾液酸作为底物，并能够在催化机制过程中捕获一种长寿命的共价中间体（CI）。在本文中，我们计算了KIE，并使用计算工具研究了TcTS中CI形成步骤的整个机制途径。特别是，自由能结果表明，在过渡态时Tyr342有强亲核参与，在此之后，体系坍缩形成稳定的CI。我们发现该反应没有碳正离子中间体。通过能量分解方法，我们确定了对催化影响最大的残基。这项研究有助于理解TcTS的催化机制，并可作为未来抑制剂设计研究的指南。
Trypanosoma cruzi trans -sialidase (TcTS) is a crucial enzyme for the infection of Trypanosoma cruzi, the protozoa responsible for Chagas’ disease in humans. It catalyzes the transfer of sialic acids from the host’s glycoconjugates to the parasite’s glycoconjugates. Based on kinetic isotope effect (KIE) studies, a strong nucleophilic participation at the transition state could be determined and; recently, elaborate experiments used 2-deoxy-2,3-difluorosialic acid as substrate and were able to trap a long-lived covalent intermediate (CI) during the catalytic mechanism. In this paper, we compute the KIE and address the entire mechanistic pathway of the CI formation step in TcTS using computational tools. Particularly, the free energy results indicate that in the transition state there is a strong nucleophilic participation of Tyr342 and after this, the system collapsed into a stable CI. We find that there is no carbocation intermediate for this reaction. By means of the energy decomposition method, we identify the residues that have the biggest influence on catalysis. This study facilitates the understanding of the catalytic mechanism of TcTS and can serve as a guide for future inhibitor design studies.
登录
查看更多内容
影响因子：
64.8
作者：
Dinner, AR;Blackburn, GM;Karplus, M
通讯作者：
Karplus, M
影响因子：
15
作者：
Horenstein, BA;Bruner, M
通讯作者：
Bruner, M
影响因子：
18.3
作者：
GUTHRIE, RD;JENCKS, WP
通讯作者：
JENCKS, WP
影响因子：
2.9
作者：
Damager, Iben;Buchini, Sabrina;Withers, Stephen G.
通讯作者：
Withers, Stephen G.
影响因子：
15
作者：
CREMER, D;POPLE, JA
通讯作者：
POPLE, JA