Catalytic Promiscuity of Ancestral Esterases and Hydroxynitrile Lyases.
Catalytic Promiscuity of Ancestral Esterases and Hydroxynitrile Lyases.
复制标题
祖先酯酶和羟基腈裂解酶的催化混杂性。
DOI：
10.1021/jacs.5b12209
复制
发表时间：
2016-01-27
影响因子：
15
通讯作者：
Kazlauskas RJ
中科院分区：
文献类型：
--
作者：
Devamani T;Rauwerdink AM;Lunzer M;Jones BJ;Mooney JL;Tan MA;Zhang ZJ;Xu JH;Dean AM;Kazlauskas RJ
催化混杂性是一种有用但偶然的酶性质，因此在自然界中寻找催化混杂性酶是低效的。一些祖先酶是新酶进化的分支点，假设是混杂的。为了验证祖先酶比其现代后代更混杂的假设，我们重建了大约1亿年前酯酶与羟基腈裂解酶（HNL’s）分化的四个分支点上的祖先酶。两种酶类型均为α/β-水解酶折叠酶，具有相同的催化三联体，但反应类型和机理不同。酯酶通过酰基酶中间体催化水解，而裂解酶催化消除而不需要中间体。用6种酯酶底物和6种裂解酶底物筛选祖先酶及其现代后代发现，祖先酶的催化混交性较高（P <0.01）。祖先酯酶比现代酯酶更容易催化裂解酶反应，祖先HNL比现代HNL更容易催化酯水解。一个祖先酶（HNL1）沿着从酯酶到羟基腈裂解酶的路径是特别混杂的，催化水解和裂解酶与许多底物的反应。更广泛的筛选测试了没有被进化选择的机械相关反应：脱羧、Michael加成、γ-内酰胺水解和1,5-二酮水解。祖先的酶比现代后代更混杂（P = 0.04）。因此，这些重建的祖先酶在催化上是混杂的，但HNL1尤其如此。
Catalytic promiscuity is a useful, but accidental, enzyme property, so finding catalytically promiscuous enzymes in nature is inefficient. Some ancestral enzymes were branch points in the evolution of new enzymes and are hypothesized to have been promiscuous. To test the hypothesis that ancestral enzymes were more promiscuous than their modern descendants, we reconstructed ancestral enzymes at four branch points in the divergence hydroxynitrile lyases (HNL’s) from esterases ~100 million years ago. Both enzyme types are α/β-hydrolase-fold enzymes and have the same catalytic triad, but differ in reaction type and mechanism. Esterases catalyze hydrolysis via an acyl enzyme intermediate, while lyases catalyze an elimination without an intermediate. Screening ancestral enzymes and their modern descendants with six esterase substrates and six lyase substrates found higher catalytic promiscuity among the ancestral enzymes (P <0.01). Ancestral esterases were more likely to catalyze a lyase reaction than modern esterases and the ancestral HNL was more likely to catalyze ester hydrolysis than modern HNL’s. One ancestral enzyme (HNL1) along the path from esterase to hydroxynitrile lyases was especially promiscuous and catalyzed both hydrolysis and lyase reactions with many substrates. A broader screen tested mechanistically related reactions that were not selected for by evolution: decarboxylation, Michael addition, γ-lactam hydrolysis and 1,5-diketone hydrolysis. The ancestral enzymes were more promiscuous than their modern descendants (P = 0.04). Thus, these reconstructed ancestral enzyme are catalytically promiscuous, but HNL1 is especially so.