Binding of alkylurea inhibitors to epoxide hydrolase implicates active site tyrosines in substrate activation
Binding of alkylurea inhibitors to epoxide hydrolase implicates active site tyrosines in substrate activation
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DOI：
10.1074/jbc.m000278200
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发表时间：
2000-05-19
影响因子：
4.8
通讯作者：
Christianson, DW
中科院分区：
文献类型：
--
作者：
Argiriadi, MA;Morisseau, C;Christianson, DW
用X-射线晶体学方法测定了两种与鼠可溶性环氧化物水解酶复合的烷基脲抑制剂的结构。每种抑制剂的烷基取代基在可溶性环氧化物水解酶活性部位形成广泛的疏水接触，每个脲羰基氧接受Tyr（381）和Tyr（465）的酚羟基的氢键。这些氢键相互作用表明Tyr（381）和/或Tyr（465）是促进水解反应第一步中的环氧化物开环的通用酸催化剂; Tyr（485）在所有环氧化物水解酶中高度保守，并且Tyr（381）在可溶性环氧化物水解酶中保守。在一种酶抑制剂复合物中，尿素羰基氧还与Gln相互作用（382）。如果在催化中发生类似的相互作用，则Gln（382）可以提供环氧化物氧上部分负电荷的静电稳定。Asp（333）的羧酸侧链接受来自每个酶抑制剂复合物中的一个脲NH基团的氢键。由于Asp（333）是催化亲核试剂，其与脲NH基团上的部分正电荷的相互作用模拟其接近环氧化物底物的亲电碳上的部分正电荷。因此，烷基脲抑制剂模仿功能遇到的反应坐标环氧化物开环，并提出了基于结构的机制白细胞毒素环氧化物水解。
The structures of two alkylurea inhibitors complexed with murine soluble epoxide hydrolase have been determined by x-ray crystallographic methods, The alkyl substituents of each inhibitor make extensive hydrophobic contacts in the soluble epoxide hydrolase active site, and each urea carbonyl oxygen accepts hydrogen bonds from the phenolic hydroxyl groups of Tyr(381) and Tyr(465). These hydrogen bond interactions suggest that Tyr(381) and/or Tyr(465) are general acid catalysts that facilitate epoxide ring opening in the first step of the hydrolysis reaction; Tyr(485) is highly conserved among all epoxide hydrolases, and Tyr(381) is conserved among the soluble epoxide hydrolases. In one enzyme-inhibitor complex, the urea carbonyl oxygen additionally interacts with Gln(382). If a comparable interaction occurs in catalysis, then Gln(382) may provide electrostatic stabilization of partial negative charge on the epoxide oxygen. The carboxylate side chain of Asp(333) accepts a hydrogen bond from one of the urea NH groups in each enzyme-inhibitor complex. Because Asp(333) is the catalytic nucleophile, its interaction with the partial positive charge on the urea NH group mimics its approach toward the partial positive charge on the electrophilic carbon of an epoxide substrate. Accordingly, alkylurea inhibitors mimic features encountered in the reaction coordinate of epoxide ring opening, and a structure-based mechanism is proposed for leukotoxin epoxide hydrolysis.