Substrate induced population shifts and stochastic gating in the PBCV-1 mRNA capping enzyme.
Substrate induced population shifts and stochastic gating in the PBCV-1 mRNA capping enzyme.
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DOI：
10.1021/ja808064g
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发表时间：
2009-04-15
影响因子：
15
通讯作者：
McCammon JA
中科院分区：
文献类型：
--
作者：
Swift RV;McCammon JA
PBCV-1 mRNA加帽酶催化第二个酶促反应，在新生mRNA的5′端形成N-7-甲基-GMP帽。它由两个球形结构域组成，由一个短的柔性肽接头结合，已显示其经历打开和关闭事件。小的尺寸和实验证明的结构域移动性使PBCV-1加帽酶成为理想的合适模型系统，以探索底物结合背景下的结构域移动性。在这里，我们具体解决以下四个问题：（1）底物结合如何影响相对结构域的流动性：系统更好地描述了诱导适合或人口转移机制？(2)能够结合底物的构象的总体特征是什么？(3)活性位点的“域门控”是否影响底物结合的速率？(4)受体构象波动的大小是否通过空间上封闭特定大小或几何形状的分子而赋予底物特异性？我们用理论、布朗动力学和分子动力学的结合来回答这些问题。我们的研究结果表明，结合效率是构象的函数，但有效和无效的结合构象之间的异构化不影响底物缔合速率。此外，我们表明，构象的灵活性本身是不足以解释单链mRNA的特异性。虽然我们的结果是特定的PBCV-1 mRNA加帽酶，它们提供了一个有用的背景下，类似结构的酶或蛋白质的底物结合行为可以考虑。
The 317 residue PBCV-1 mRNA capping enzyme catalyzes the second enzymatic reaction in the formation of an N-7-methyl-GMP cap on the 5′-end of the nascent mRNA. It is composed of two globular domains bound by a short flexible peptide linker, which have been shown to undergo opening and closing events. The small size and experimentally demonstrated domain mobility make the PBCV-1 capping enzyme an ideally suited model system to explore domain mobility in context of substrate binding. Here, we specifically address the following four questions: (1) How does substrate binding affect relative domain mobility: is the system better described by an induced fit or population shift mechanism? (2) What are the gross characteristics of a conformation capable of binding substrate? (3) Does “domain gating” of the active site affect the rate of substrate binding? (4) Does the magnitude of receptor conformational fluctuations confer substrate specificity by sterically occluding molecules of a particular size or geometry? We answer these questions using a combination of theory, Brownian dynamics, and molecular dynamics. Our results show that binding efficiency is a function of conformation but that isomerization between efficient and inefficient binding conformations does not impact the substrate association rate. Additionally, we show that conformational flexibility alone is insufficient to explain single stranded mRNA specificity. While our results are specific to the PBCV-1 mRNA capping enzyme, they provide a useful context within which the substrate binding behavior of similarly structured enzymes or proteins may be considered.
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影响因子：
56.9
作者：
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通讯作者：
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影响因子：
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作者：
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通讯作者：
McCammon, JA