Crystal structure of a bacterial cocaine esterase
Crystal structure of a bacterial cocaine esterase
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DOI：
10.1038/nsb742
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发表时间：
2002-01-01
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影响因子：
--
通讯作者：
Wilson, IA
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文献类型：
--
作者：
Larsen, NA;Turner, JM;Wilson, IA
在这里，我们报告了可卡因降解酶的第一个结构。细菌酯酶 cocE 将具有药理活性的 (-)-可卡因水解为非精神活性代谢物，其速度比任何其他报道的可卡因酯酶都要快（k(cat) = 7.8 s(-1) 和 K-M = 640 nM）。由于 cocE 的高催化能力，它是用于治疗可卡因过量的新型蛋白质疗法的有吸引力的候选者。通过多重异常分散 (MAD) 方法解析的 cocE 晶体结构表明，cocE 是一种丝氨酸酯酶，由三个结构域组成：(i) 典型的 α/β 水解酶折叠 (ii) 覆盖活性位点的 α 螺旋结构域，以及 (iii) 与其他两个结构域广泛相互作用的果冻卷状 β 结构域。通过分析过渡态类似加合物和产物复合物的晶体结构，分别在 1.58 埃和 1.63 埃分辨率下精炼，确定了所有三个域界面内的活性位点。这些结构研究表明，底物识别部分源于可卡因的苯甲酰基部分和高度进化的特异性口袋之间的相互作用。
Here we report the first structure of a cocaine-degrading enzyme. The bacterial esterase, cocE, hydrolyzes pharmacologically active (-)-cocaine to a nonpsychoactive metabolite with a rate faster than any other reported cocaine esterase (k(cat) = 7.8 s(-1) and K-M = 640 nM). Because of the high catalytic proficiency of cocE, it is an attractive candidate for novel protein- based therapies for cocaine overdose. The crystal structure of cocE, solved by multiple anomalous dispersion (MAD) methods, reveals that cocE is a serine esterase composed of three domains: (i) a canonical alpha/beta hydrolase fold (ii) an alpha-helical domain that caps the active site and (iii) a jelly-roll-like beta-domain that interacts extensively with the other two domains. The active site was identified within the interface of all three domains by analysis of the crystal structures of transition state analog adduct and product complexes, which were refined at 1.58 Angstrom and 1.63 Angstrom resolution, respectively. These structural studies suggest that substrate recognition arises partly from interactions between the benzoyl moiety of cocaine and a highly evolved specificity pocket.