Cooperative Electrostatic Interactions Drive Functional Evolution in the Alkaline Phosphatase Superfamily.
Cooperative Electrostatic Interactions Drive Functional Evolution in the Alkaline Phosphatase Superfamily.
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DOI：
10.1021/jacs.5b03945
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发表时间：
2015-07-22
影响因子：
15
通讯作者：
Kamerlin SC
中科院分区：
文献类型：
--
作者：
Barrozo A;Duarte F;Bauer P;Carvalho AT;Kamerlin SC
人们普遍认为，催化混杂，即单一酶催化多个不同化学底物周转的能力，在新酶功能的进化中起着关键作用。在这种背景下，碱性磷酸酶超家族的成员被广泛地作为模型系统来研究，以了解酶的多功能现象。在目前的工作中，我们模拟了这个超家族中两个多重混杂成员的选择性，即来自石竹伯克霍尔德氏菌和豆根瘤菌的磷酸单酯水解酶。我们对野生型酶和几个实验表征的突变体的酶反应进行了广泛的模拟。我们的计算模型与关键的实验观测结果是一致的，例如观察到的野生型酶的活性，对实验pH速率曲线的定性解释，以及几个活性位点突变体之间的活性趋势。在所有情况下，感兴趣的底物都以类似的构象与酶结合，在水溶液中具有与其对应的类似物基本不受干扰的过渡态。对过渡态几何结构和单个残基对计算的激活势垒的贡献的研究表明，这些酶的广泛混杂源于活性部位的协同静电相互作用，使每个酶能够适应不同底物的静电需求。通过比较几种碱性磷酸酶的结构和静电特征，我们认为这一现象是一个普遍的特征，驱动了这个超家族的选择性和混杂，进而可以用于人工酶设计。
It is becoming widely accepted that catalytic promiscuity, i.e., the ability of a single enzyme to catalyze the turnover of multiple, chemically distinct substrates, plays a key role in the evolution of new enzyme functions. In this context, the members of the alkaline phosphatase superfamily have been extensively studied as model systems in order to understand the phenomenon of enzyme multifunctionality. In the present work, we model the selectivity of two multiply promiscuous members of this superfamily, namely the phosphonate monoester hydrolases from Burkholderia caryophylli and Rhizobium leguminosarum. We have performed extensive simulations of the enzymatic reaction of both wild-type enzymes and several experimentally characterized mutants. Our computational models are in agreement with key experimental observables, such as the observed activities of the wild-type enzymes, qualitative interpretations of experimental pH-rate profiles, and activity trends among several active site mutants. In all cases the substrates of interest bind to the enzyme in similar conformations, with largely unperturbed transition states from their corresponding analogues in aqueous solution. Examination of transition-state geometries and the contribution of individual residues to the calculated activation barriers suggest that the broad promiscuity of these enzymes arises from cooperative electrostatic interactions in the active site, allowing each enzyme to adapt to the electrostatic needs of different substrates. By comparing the structural and electrostatic features of several alkaline phosphatases, we suggest that this phenomenon is a generalized feature driving selectivity and promiscuity within this superfamily and can be in turn used for artificial enzyme design.