Crystal structures of Bacillus caldovelox arginase in complex with substrate and inhibitors reveal new insights into activation, inhibition and catalysis in the arginase superfamily
Crystal structures of Bacillus caldovelox arginase in complex with substrate and inhibitors reveal new insights into activation, inhibition and catalysis in the arginase superfamily
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DOI：
10.1016/s0969-2126(99)80056-2
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发表时间：
1999-04-15
期刊：
影响因子：
--
通讯作者：
Baker, EN
中科院分区：
文献类型：
--
作者：
Bewley, MC;Jeffrey, PD;Baker, EN
背景：精氨酸酶是一种锰依赖性酶，催化L-精氨酸水解为L-鸟氨酸和尿素。在输尿管动物中，精氨酸酶是尿素循环的最终酶，但在许多物种中，它具有更广泛的作用，控制精氨酸用于其他代谢目的的使用，包括肌酸、多胺、脯氨酸和一氧化氮的产生。精氨酸酶活性受多种小分子调节，包括产物 L-鸟氨酸。这些结构研究的目的是测试催化机制的各个方面并研究精氨酸酶抑制的结构基础。结果：我们在此报告了来自 Bacillus caldovelox 的精氨酸酶在 pH 5.6 和 pH 8.5 下的晶体结构，以及该酶与 L-精氨酸、L-鸟氨酸和 L-赖氨酸在 pH 8.5 下的二元复合物的晶体结构。精氨酸酶单体包含单个紧凑的α/β结构域，其进一步缔合成六聚四级结构。二元复合物揭示了配体结合的常见模式，该模式将底物置于邻近二锰中心的位置。我们还观察到影响活性位点的构象变化，并与胍或精氨酸占据外部位点相结合。结论：此处报告的结构阐明了活性位点的各个方面，并表明了催化机制的关键特征，包括与其中一个锰离子的底物配位以及相邻组氨酸残基的定向作用。发现 L-氨基酸的立体特异性取决于它们在活性位点边缘的精确识别。远离活性位点的第二个精氨酸结合位点的鉴定以及相关的构象变化使我们提出该位点在底物水解中的调节作用。
Background: Arginase is a manganese-dependent enzyme that catalyzes the hydrolysis of L-arginine to L-ornithine and urea. In ureotelic animals arginase is the final enzyme of the urea cycle, but in many species it has a wider role controlling the use of arginine for other metabolic purposes, including the production of creatine, polyamines, proline and nitric oxide. Arginase activity is regulated by various small molecules, including the product L-ornithine. The aim of these structural studies was to test aspects of the catalytic mechanism and to investigate the structural basis of arginase inhibition.Results: We report here the crystal structures of arginase from Bacillus caldovelox at pH 5.6 and pH 8.5, and of binary complexes of the enzyme with L-arginine, L-ornithine and L-lysine at pH 8.5. The arginase monomer comprises a single compact alpha/beta domain that further associates into a hexameric quaternary structure. The binary complexes reveal a common mode of ligand binding, which places the substrate adjacent to the dimanganese centre. We also observe a conformational change that impacts on the active site and is coupled with the occupancy of an external site by guanidine or arginine.Conclusions: The structures reported here clarify aspects of the active site and indicate key features of the catalytic mechanism, including substrate coordination to one of the manganese ions and an orientational role for a neighboring histidine residue. Stereospecificity for L-amino acids is found to depend on their precise recognition at the active-site rim. Identification of a second arginine-binding site, remote from the active site, and associated conformational changes lead us to propose a regulatory role for this site in substrate hydrolysis.