Flavin analogue studies of pig kidney general acyl-CoA dehydrogenase.
Flavin analogue studies of pig kidney general acyl-CoA dehydrogenase.
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DOI：
10.1021/bi00281a029
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发表时间：
1983-06
期刊：
影响因子：
2.9
通讯作者：
C. Thorpe;V. Massey
中科院分区：
文献类型：
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作者：
C. Thorpe;V. Massey
Colin Thorpe和Vincent Massey摘要：猪肾通用酰辅酶A脱氢酶的载脂蛋白由8个环状修饰的黄素腺嘌呤二核苷酸(FAD)类似物重组，以探索黄素结合的环境以及酶与乙酰辅酶A衍生物的相互作用。8-氯-，7-溴-和2-硫代-FAD的加成可产生可被辛酰辅酶A还原的全酶，并且在以吩嗪甲硫酸盐为介体的测定体系中表现出显著的酶活性。相反，与具有比FAD更负的标准氧化还原电位的类似物，1-去氮杂-，5-去氮杂-，6-羟基-，8-羟基-和8-巯基-FAD重组酶，产生的衍生物是不活跃的，并且不会被硫酯底物显著还原。相反，5-脱氮-FADH2脱氢酶被巴豆酰-辅酶A迅速重新氧化，这表明高度不受欢迎的5-去氮杂黄半喹酮不是该反应的必备中间体。8-氯-和8-巯基-FAD的实验-最近，人们对酰基-辅酶A脱氢酶，特别是参与哺乳动物脂肪酸氧化的一般酰基-辅酶A脱氢酶重新产生了兴趣。这种线粒体黄素蛋白首先是由Beinert等人研究的(Crane等人，1956；Beinert，1963)，并显示出广泛的底物专一性，对中链长度的酰基CoA硫酯具有最佳活性(Crane等人，1956；Hall&Kamin，1975；Thorpe等人，1979)。这些底物导致黄素假体快速漂白
Colin Thorpe* and Vincent Massey abstract: The apoprotein of pig kidney general acyl-CoA dehydrogenase has been reconstituted with eight ring-modified flavin adenine dinucleotide (FAD) analogues to probe the environment of the bound flavin and theinteraction of the enzyme with acyl-CoAderivatives. Addition of 8-C1-, 7-Br-, and 2-thio-FAD yields holoenzymes that can be reduced by octanoyl-CoA and that exhibit appreciable enzymatic activity in an assay system utilizing phenazine methosulfate as a mediator. In contrast, reconstitution of the enzyme with analogues that have more negative standard redox potentials than FAD, 1-deaza-, 5-deaza-, 6-OH-, 8-OH-, and 8-mercapto-FAD, generates derivatives that are inactive and not significantly reduced by thioester substrates. In the reverse direction, the 5-deaza-FADH2 dehydrogenase is rapidly re-oxidized by crotonyl-CoA, suggesting that the highly unfavored 5-deazaflavosemiquinone is not an obligatory intermediate in this reaction. Experiments with 8-C1-and 8-mercapto-FAD-Recently, there has been a resurgence of interest in the acyl-CoA dehydrogenases and in particular in the general acyl-CoA dehydrogenase that participates in mammalian fatty acid oxidation. This mitochondrial flavoprotein was first studied by Beinert and co-workers (Crane et al., 1956; Beinert, 1963) and exhibits a broad substrate specificity with optimal activity toward medium chain length acyl-CoA thioesters (Crane et al., 1956; Hall & Kamin, 1975; Thorpe et al., 1979). These substrates induce rapid bleaching of the flavin prosthetic