Department of Chemistry and Biochemistry, Butler University, Indianapolis, Indiana, United States of America.
PLoS One. 2020 Mar 17;15(3):e0230166. doi: 10.1371/journal.pone.0230166. eCollection 2020.
Over 100 metabolic serine hydrolases are present in humans with confirmed functions in metabolism, immune response, and neurotransmission. Among potentially clinically-relevant but uncharacterized human serine hydrolases is OVCA2, a serine hydrolase that has been linked with a variety of cancer-related processes. Herein, we developed a heterologous expression system for OVCA2 and determined the comprehensive substrate specificity of OVCA2 against two ester substrate libraries. Based on this analysis, OVCA2 was confirmed as a serine hydrolase with a strong preference for long-chain alkyl ester substrates (>10-carbons) and high selectivity against a variety of short, branched, and substituted esters. Substitutional analysis was used to identify the catalytic residues of OVCA2 with a Ser117-His206-Asp179 classic catalytic triad. Comparison of the substrate specificity of OVCA2 to the model homologue FSH1 from Saccharomyces cerevisiae illustrated the tighter substrate selectivity of OVCA2, but their overlapping substrate preference for extended straight-chain alkyl esters. Conformation of the overlapping biochemical properties of OVCA2 and FSH1 was used to model structural information about OVCA2. Together our analysis provides detailed substrate specificity information about a previously, uncharacterized human serine hydrolase and begins to define the biological properties of OVCA2.
人类中存在超过 100 种代谢丝氨酸水解酶，这些酶在代谢、免疫反应和神经递质传递中具有明确的功能。在具有潜在临床相关性但尚未确定的人类丝氨酸水解酶中，有 OVCA2，这是一种与多种癌症相关过程有关的丝氨酸水解酶。在此，我们开发了 OVCA2 的异源表达系统，并针对两种酯底物文库确定了 OVCA2 的全面底物特异性。基于此分析，OVCA2 被确认为一种丝氨酸水解酶，对长链烷基酯底物（>10 个碳原子）有强烈的偏好，并且对各种短链、支链和取代酯具有很高的选择性。取代分析用于确定 OVCA2 的催化残基，其具有 Ser117-His206-Asp179 经典催化三联体。将 OVCA2 的底物特异性与酿酒酵母的模型同源物 FSH1 进行比较，表明 OVCA2 的底物选择性更严格，但它们对扩展直链烷基酯的底物偏好重叠。OVCA2 和 FSH1 的重叠生化特性的构象用于对 OVCA2 的结构信息进行建模。总的来说，我们的分析提供了关于以前未被表征的人类丝氨酸水解酶的详细底物特异性信息，并开始定义 OVCA2 的生物学特性。
