Turnover-dependent covalent inactivation of Staphylococcus aureus coenzyme A-disulfide reductase by coenzyme A-mimetics: mechanistic and structural insights.
Turnover-dependent covalent inactivation of Staphylococcus aureus coenzyme A-disulfide reductase by coenzyme A-mimetics: mechanistic and structural insights.
复制标题
辅酶 A-模拟物对金黄色葡萄球菌辅酶 A-二硫键还原酶的周转依赖性共价失活：机制和结构见解。
DOI：
10.1021/bi301026c
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发表时间：
2012
期刊：
影响因子：
2.9
通讯作者：
Claiborne,Al
中科院分区：
文献类型：
--
作者：
Wallace,BretD;Edwards,JonathanS;Wallen,JamieR;Moolman,WesselJA;vanderWesthuyzen,Renier;Strauss,Erick;Redinbo,MatthewR;Claiborne,Al
金黄色葡萄球菌中不寻常的基于硫醇的氧化还原动态平衡机制的破坏代表了一个独特的机会来确定新的代谢过程和新的干预目标。针对Coash生物合成和氧化还原功能的不常见方面。在金黄色葡萄球菌中，抗生素CJ-15,801最近被证明是这种生物中Coash生物合成途径的抗代谢物；含有α，β-不饱和砜和羧基的COAS模拟物也被开发为不可逆的OFS抑制剂。金黄色辅酶A-二硫键还原酶(SaCoADR)。在这项工作中，我们确定了三个这样的共价SaCoADR-抑制剂络合物的晶体结构，这些络合物是通过在周转过程中灭活野生型酶而制备的。其结构揭示了活性中心Cys43-Sγ与乙烯基砜或羧基的Cβ之间的共价键。每个酶二聚体占据两个抑制分子，结合对野生型/C43S杂二聚体的动力学分析，表明在正常的催化周转过程中，半位反应活性不是一个因素。此外，我们提供了SaCoADR活性位点突变体的结构；特别是Tyr419‘-OH在指导Cys43-SSCoA氧化还原中心的分子内还原、在NADPH滴定中观察到的两个FAD/二聚体的氧化还原不对称以及在催化中发挥着重要作用。在C43S突变结构中观察到的Ser43侧链的两种构象支持Cys43-Sγ在其催化的Cys43-SSCoA/Cys43-SH氧化还原循环中的构象开关。最后，这三种抑制剂复合体的结构为设计对几种主要细菌病原体具有治疗潜力的更有效的抑制剂提供了一个框架。
Disruption of the unusual thiol-based redox homeostasis mechanisms inStaphylococcus aureusrepresents a unique opportunity to identify new metabolic processes and new targets for intervention. Targeting uncommon aspects of CoASH biosynthetic and redox functions inS. aureus, the antibiotic CJ-15,801 has recently been demonstrated to be an antimetabolite of the CoASH biosynthetic pathway in this organism; CoAS-mimetics containing α,β-unsaturated sulfone and carboxyl moieties have also been exploited as irreversible inhibitors ofS. aureuscoenzyme A-disulfide reductase (SaCoADR). In this work we have determined the crystal structures of three of these covalentSaCoADR-inhibitor complexes, prepared by inactivation of wild-type enzyme during turnover. The structures reveal the covalent linkage between the active-site Cys43-Sγand Cβof the vinyl sulfone or carboxyl moiety. The full occupancy of two inhibitor molecules per enzyme dimer, together with kinetic analyses of the wild-type/C43S heterodimer, indicates that half-sites-reactivity is not a factor during normal catalytic turnover. Further, we provide the structures ofSaCoADR active-site mutants; in particular, Tyr419′-OH plays dramatic roles in directing intramolecular reduction of the Cys43-SSCoA redox center, in the redox asymmetry observed for the two FAD per dimer in NADPH titrations, and in catalysis. The two conformations observed for the Ser43 side chain in the C43S mutant structure lend support to a conformational switch for Cys43-Sγduring its catalytic Cys43-SSCoA/Cys43-SH redox cycle. Finally, the structures of the three inhibitor complexes provide a framework for design of more effective inhibitors with therapeutic potential against several major bacterial pathogens.