The Kinetic Mechanism of the Human Bifunctional Enzyme ATIC (5-Amino-4-imidazolecarboxamide Ribonucleotide Transformylase/Inosine 5′-Monophosphate Cyclohydrolase)
The Kinetic Mechanism of the Human Bifunctional Enzyme ATIC (5-Amino-4-imidazolecarboxamide Ribonucleotide Transformylase/Inosine 5′-Monophosphate Cyclohydrolase)
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人类双功能酶 ATIC（5-氨基-4-咪唑甲酰胺核糖核苷酸转化酶/肌苷 5-单磷酸环水解酶）的动力学机制
DOI：
--
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发表时间：
2002
影响因子：
4.8
通讯作者：
K. Anderson
中科院分区：
文献类型：
--
作者：
Karen G. Bulock;G. Beardsley;K. Anderson
5-氨基-4-咪唑甲酰胺核糖核苷酸转化酶/IMP环化水解酶（ATIC）是一种双功能蛋白，具有两种酶活性，依次催化肌苷5′-磷酸福尔德从头合成途径的最后两步。这种双功能酶是特别感兴趣的，因为它作为化疗靶点的潜力。此外，这两种催化活性在整个自然界中存在于同一种蛋白质上，这就提出了双功能性是否具有动力学优势的问题。快速化学淬灭，停流吸收，稳态动力学技术被用来阐明完整的动力学机制的人ATIC。动力学模拟程序KINSIM被用来模拟在这项研究中获得的动力学数据。详细的动力学分析，结合动力学模拟，提供了酶反应途径的以下关键特征。1)整个反应中的限速步骤（2.9 ± 0.4 s-1）可能是从甲酰转移酶活性位点释放四氢叶酸或与四氢叶酸释放相关的构象变化。2)反向转化酶反应的速率（6.7 s-1）比正向速率（2.9 s-1）快2.2 -3倍，而环化水解酶反应在正向意义上基本上是单向的。因此，环化水解酶反应将整个双功能反应引向肌苷一磷酸的产生。3)没有动力学证据的中间体，甲酰氨基咪唑甲酰胺核糖核苷酸，甲酰基转移酶和环化水解酶活性位点之间的底物通道。
5-Amino-4-imidazolecarboxamide ribonucleotide transformylase/IMP cyclohydrolase (ATIC) is a bifunctional protein possessing two enzymatic activities that sequentially catalyze the last two steps in the pathway forde novo synthesis of inosine 5′-monophosphate. This bifunctional enzyme is of particular interest because of its potential as a chemotherapeutic target. Furthermore, these two catalytic activities reside on the same protein throughout all of nature, raising the question of whether there is some kinetic advantage to the bifunctionality. Rapid chemical quench, stopped-flow absorbance, and steady-state kinetic techniques were used to elucidate the complete kinetic mechanism of human ATIC. The kinetic simulation program KINSIM was used to model the kinetic data obtained in this study. The detailed kinetic analysis, in combination with kinetic simulations, provided the following key features of the enzyme reaction pathway. 1) The rate-limiting step in the overall reaction (2.9 ± 0.4 s−1) is likely the release of tetrahydrofolate from the formyltransferase active site or a conformational change associated with tetrahydrofolate release. 2) The rate of the reverse transformylase reaction (6.7 s−1) is ∼2–3-fold faster than the forward rate (2.9 s−1), whereas the cyclohydrolase reaction is essentially unidirectional in the forward sense. The cyclohydrolase reaction thus draws the overall bifunctional reaction toward the production of inosine monophosphate. 3) There was no kinetic evidence of substrate channeling of the intermediate, the formylaminoimidazole carboxamide ribonucleotide, between the formyltransferase and the cyclohydrolase active sites.
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鸡肝中5-氨基咪唑-4-甲酰胺-核糖核苷酸转化酶的纯化及作用机制研究
影响因子：
2.9
作者：
Mueller,WT;Benkovic,SJ
通讯作者：
Benkovic,SJ
DOI：
--
发表时间：
1989-01
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
G. Beardsley;B. Moroson;E. C. Taylor;R. Moran
通讯作者：
G. Beardsley;B. Moroson;E. C. Taylor;R. Moran
影响因子：
2.9
作者：
BALDWIN, SW;TSE, A;MORAN, RG
通讯作者：
MORAN, RG
影响因子：
--
作者：
Anderson,KS
通讯作者：
Anderson,KS
影响因子：
15.9
作者：
CRONSTEIN, BN;NAIME, D;OSTAD, E
通讯作者：
OSTAD, E