Crystal structure of aminopeptidase N (proteobacteria alanyl aminopeptidase) from Escherichia coli and conformational change of methionine 260 involved in substrate recognition
Crystal structure of aminopeptidase N (proteobacteria alanyl aminopeptidase) from Escherichia coli and conformational change of methionine 260 involved in substrate recognition
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DOI：
10.1074/jbc.m605203200
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发表时间：
2006-11-03
影响因子：
4.8
通讯作者：
Yoshimoto, Tadashi
中科院分区：
文献类型：
--
作者：
Ito, Kiyoshi;Nakajima, Yoshitaka;Yoshimoto, Tadashi
来自大肠杆菌的氨肽酶 N 是一种广泛特异性的锌外肽酶，属于氨肽酶 MA 族、M1 家族。无配体形式和酶贝他汀复合物的结构分别以 1.5-A 和 1.6-A 分辨率测定。该酶由四个结构域组成：N 端 β 结构域 (Met(1)-Asp(193))、催化结构域 (Phe(194)-Gly(444))、中间 β 结构域 (Thr(445)-Trp(546)) 和 C 端 α 结构域 (Ser(547)-Ala(870))。该催化结构域的结构与嗜热菌蛋白酶相似，并且在该结构域中发现了金属结合基序（HEXXHX18E）。锌离子由 His(297)、His(301)、Glu(320) 和水分子配位。含有活性位点的催化结构域上的凹槽被C端α结构域覆盖，并且在蛋白质内部形成一个大空腔。然而，C 端 α 结构域的中心存在一个小孔。 bestatin 的 N 末端被 Glu(121) 和 Glu(264) 识别，它们分别位于 N 末端和催化结构域。位于锌离子附近的 Glu(298) 和 Tyr(381) 被认为参与肽裂解。无配体形式和酶-bestatin 复合物之间的差异表明，Met(260) 作为接受具有不同 N 末端残基大小的底物的缓冲垫，导致该酶具有广泛的底物特异性。
Aminopeptidase N from Escherichia coli is a broad specificity zinc exopeptidase belonging to aminopeptidase clan MA, family M1. The structures of the ligand-free form and the enzymebestatin complex were determined at 1.5- and 1.6-A resolution, respectively. The enzyme is composed of four domains: an N-terminal beta-domain (Met(1)-Asp(193)), a catalytic domain (Phe(194)-Gly(444)), a middle beta-domain (Thr(445)-Trp(546)), and a C-terminal alpha-domain (Ser(547)-Ala(870)). The structure of the catalytic domain exhibits similarity to thermolysin, and a metal-binding motif ( HEXXHX18E) is found in the domain. The zinc ion is coordinated by His(297), His(301), Glu(320), and a water molecule. The groove on the catalytic domain that contains the active site is covered by the C-terminal alpha-domain, and a large cavity is formed inside the protein. However, there exists a small hole at the center of the C-terminal alpha-domain. The N terminus of bestatin is recognized by Glu(121) and Glu(264), which are located in the N-terminal and catalytic domains, respectively. Glu(298) and Tyr(381), located near the zinc ion, are considered to be involved in peptide cleavage. A difference revealed between the ligand-free form and the enzyme-bestatin complex indicated that Met(260) functions as a cushion to accept substrates with different N-terminal residue sizes, resulting in the broad substrate specificity of this enzyme.