tRNA-guanine transglycosylase from E-coli:: a ping-pong kinetic mechanism is consistent with nucleophilic catalysis
tRNA-guanine transglycosylase from E-coli:: a ping-pong kinetic mechanism is consistent with nucleophilic catalysis
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DOI：
10.1016/s0045-2068(03)00069-5
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发表时间：
2003-08-01
影响因子：
5.1
通讯作者：
Garcia, GA
中科院分区：
文献类型：
--
作者：
Goodenough-Lashua, DM;Garcia, GA
tRNA-鸟嘌呤转糖基酶（TGT）是吡咯并嘧啶碱基鸟嘌呤对某些tRNA进行转录后修饰的关键酶。TGT是福氏志贺菌（一种人类病原体）致病所必需的，因此是一种潜在的新型抗菌靶标。先前的工作已经表明，TGT反应通过共价酶-tRNA复合物进行[Biochemistry 40（2001）14123]。为了进一步证实这一机制，进行了TGT反应的动力学机制的测定。计算和图形分析的初始速度数据是最符合乒乓运动机制。7-甲基鸟嘌呤对鸟嘌呤（竞争性）和tRNA（非竞争性）的抑制模式表明，tRNA首先与酶结合。该动力学机制与共价中间体化学机制和我们早期对基于机制的抑制剂[7-氟甲基-7-脱氮鸟嘌呤，Biochemistry 34（1995）15539]的研究一致，其中TGT失活依赖于tRNA的存在。(C)2003 Elsevier Science（美国）。All rights reserved.
tRNA-guanine transglycosylase (TGT) is a key enzyme in the post-transcriptional modification of certain tRNAs with the pyrrolopyrimidine base queuine. TGT is required for pathogenicity in Shigella flexneri, a human pathogen, and therefore is potentially a novel antibacterial target. Previous work has indicated that the TGT reaction proceeds through a covalent enzyme-tRNA complex [Biochemistry 40 (2001) 14123]. To further substantiate this mechanism, the determination of the kinetic mechanism for the TGT reaction was undertaken. Computational and graphical analyses of initial velocity data are most consistent with a ping-pong kinetic mechanism. The modes of inhibition of 7-methylguanine with respect to both guanine (competitive) and tRNA (uncompetitive) indicate that tRNA binds first to the enzyme. This kinetic mechanism is consistent with the covalent intermediate chemical mechanism and with our earlier study of a mechanism-based inhibitor [7-fluoromethyl-7-deazaguanine, Biochemistry 34 (1995) 15539] in which TGT inactivation was dependent upon the presence of tRNA. (C) 2003 Elsevier Science (USA). All rights reserved.