Characterization of Streptococcus pneumoniae enoyl-(acyl-carrier protein) reductase (FabK)
Characterization of Streptococcus pneumoniae enoyl-(acyl-carrier protein) reductase (FabK)
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DOI：
10.1042/bj20021699
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发表时间：
2003-03-15
影响因子：
4.1
通讯作者：
Wallis, NG
中科院分区：
文献类型：
--
作者：
Marrakchi, H;DeWolf, WE;Wallis, NG
烯酰基-（酰基-载体蛋白）（ACIP）还原酶催化11型脂肪酸合酶系统中脂肪酸延伸的每个循环中的最后一步。广泛表征的NADH依赖性还原酶Fab 1广泛分布于细菌和植物中，而烯酰-ACP还原酶FabK是在肺炎链球菌中发现的该酶组的明显不同的成员。我们无法从链球菌中删除fabK基因。pneumoniae，表明这是该生物体中唯一的烯酰-ACP还原酶。FabK酶的纯化和还原酶的生化性质进行了检查。纯化的蛋白质的可见吸收光谱表明存在黄素辅因子，其通过MS鉴定为FMN，并且与蛋白质以1：1的摩尔比存在。FabK特异性地需要NADH，并且蛋白质活性被铵离子刺激。FabK在无底物存在下也表现出NADH氧化酶活性。链球菌肺炎杆菌属于芽孢杆菌/乳杆菌/链球菌组，包括金黄色葡萄球菌和枯草芽孢杆菌。这两种生物体也含有FabK相关基因，表明它们也可能表达FabK样烯酰-ACP还原酶。然而，这些基因没有互补fabI（Ts）突变体，纯化的黄素蛋白不能在体外还原烯酰-ACP，也没有表现出NAD（P）H氧化酶活性，表明它们不是烯酰-ACP还原酶。FabK烯酰-ACP还原酶的限制性发生可能与底物非依赖性NADH氧化在氧依赖性厌氧能量代谢中的作用有关。
The enoyl-(acyl-carrier protein) (ACIP) reductase catalyses the last step in each cycle of fatty acid elongation in the type 11 fatty acid synthase systems. An extensively characterized NADH-dependent reductase, Fab1, is widely distributed in bacteria and plants, whereas the enoyl-ACP reductase, FabK, is a distinctly different member of this enzyme group discovered in Streptococcus pneumoniae. We were unable to delete the fabK gene from Strep. pneumoniae, suggesting that this is the only enoyl-ACP reductase in this organism. The FabK enzyme was purified and the biochemical properties of the reductase were examined. The visible absorption spectrum of the purified protein indicated the presence of a flavin cofactor that was identified as FMN by MS, and was present in a 1: 1 molar ratio with protein. FabK specifically required NADH and the protein activity was stimulated by ammonium ions. FabK also exhibited NADH oxidase activity in the absence of substrate. Strep. pneumoniae belongs to the Bacillus/Lactobacillus/Streptococcus group that includes Staphylococcus aureus and Bacillus subtilis. These two organisms also contain FabK-related genes, suggesting that they may also express a FabK-like enoyl-ACP reductase. However, the genes did not complement a fabI(Ts) mutant and the purified flavoproteins were unable to reduce enoyl-ACP in vitro and did not exhibit NAD(P)H oxidase activity, indicating they were not enoyl-ACP reductases. The restricted occurrence of the FabK enoyl-ACP reductase may be related to the role of substrate-independent NADH oxidation in oxygen-dependent anaerobic energy metabolism.