Temperature-sensitive contacts in disordered loops tune enzyme I activity.
Temperature-sensitive contacts in disordered loops tune enzyme I activity.
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DOI：
10.1073/pnas.2210537119
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发表时间：
2022-11-22
影响因子：
11.1
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中科院分区：
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温度影响所有酶的催化速率。然而，温度对酶催化活性的影响是蛋白质序列、结构和动力学的复杂函数。因此，人们对在不同温度下调节催化速率的酶的分子特征仍然知之甚少。在此，我们通过模拟和诱变实验揭示了细菌酶l C结构域的嗜温和嗜热同源物的温度调节机制。我们发现酶可以通过位于无序环中的温度敏感残基接触网络来调节其生理温度。此外，我们发现一些温度敏感触点之间表现出线性温度依赖性，另一些则表现出非线性温度依赖性。这些线索为调节酶活性提供了一种有前景的基于物理的方法。具有相同折叠的同源酶通常表现出不同的热和动力学行为。了解酶序列如何在功能最佳温度下编码催化活性是生物物理学的一个基本问题。最近的研究表明，调节细菌酶 I (EIC) C 端结构域的嗜热/嗜温变体的催化活性的残基主要位于无序环内，为研究这种现象提供了一个模型系统。在这项工作中，我们利用分子动力学模拟和诱变实验揭示了 EIC 同系物的序列依赖性活性调节机制。我们发现催化回路中的接触网络对温度变化特别敏感，一些接触表现出明显的线性或非线性温度依赖性趋势。此外，这些趋势定义了结构上聚集的动力学模式，并且可以区分在较高温度下趋于有序或无序的区域。分析了几种嗜热EIC突变体，我们发现对温度最敏感的位置的互补嗜温突变表现出最强的活性增强，而对温度相对不敏感的位置的突变则表现出最弱的增强活性。这些结果提供了序列依赖性温度调节的机制解释，并为合理的酶修饰提供了计算方法。
Temperature affects the catalytic rates of all enzymes. However, the impact of temperature on an enzyme's catalytic activity is a complex function of protein sequence, structure, and dynamics. Therefore, the molecular features of enzymes that tune catalytic rates at different temperatures remain poorly understood. Herein we use simulations and mutagenesis experiments to reveal the temperature-tuning mechanism of mesophilic and thermophilic homologs of the C domain of bacterial enzyme l. We find that enzymes can be tuned to their physiological temperatures through a network of temperature-sensitive residue contacts localized in the disordered loops. Furthermore, we find that some exhibit linear and others nonlinear temperature dependence among temperature-sensitive contacts. These clues offer a promising physics-based approach for tuning enzyme activity. Homologous enzymes with identical folds often exhibit different thermal and kinetic behaviors. Understanding how an enzyme sequence encodes catalytic activity at functionally optimal temperatures is a fundamental problem in biophysics. Recently it was shown that the residues that tune catalytic activities of thermophilic/mesophilic variants of the C-terminal domain of bacterial enzyme I (EIC) are largely localized within disordered loops, offering a model system with which to investigate this phenomenon. In this work, we use molecular dynamics simulations and mutagenesis experiments to reveal a mechanism of sequence-dependent activity tuning of EIC homologs. We find that a network of contacts in the catalytic loops is particularly sensitive to changes in temperature, with some contacts exhibiting distinct linear or nonlinear temperature-dependent trends. Moreover, these trends define structurally clustered dynamical modes and can distinguish regions that tend toward order or disorder at higher temperatures. Assaying several thermophilic EIC mutants, we show that complementary mesophilic mutations to the most temperature-sensitive positions exhibit the most enhanced activity, while mutations to relatively temperature insensitive positions exhibit the least enhanced activities. These results provide a mechanistic explanation of sequence-dependent temperature tuning and offer a computational method for rational enzyme modification.
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影响因子：
16.8
作者：
Bhabha, Gira;Ekiert, Damian C.;Jennewein, Madeleine;Zmasek, Christian M.;Tuttle, Lisa M.;Kroon, Gerard;Dyson, H. Jane;Godzik, Adam;Wilson, Ian A.;Wright, Peter E.
通讯作者：
Wright, Peter E.
影响因子：
5.5
作者：
Stirnemann, Guillaume;Sterpone, Fabio
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Sterpone, Fabio
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5.8
作者：
Pronk, Sander;Pall, Szilard;Lindahl, Erik
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Lindahl, Erik
影响因子：
6.3
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Tribello, Gareth A.;Bonomi, Massimiliano;Bussi, Giovanni
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Bussi, Giovanni
影响因子：
4.4
作者：
Fukunishi, H;Watanabe, O;Takada, S
通讯作者：
Takada, S