Regulation mechanism of the DsbA/DsbB system that catalyzes disulfine bond formation

在大肠杆菌蛋白二硫键形成途径中，膜蛋白DsbB对还原的质周DsbA（二硫键引入酶）进行再氧化。我们的研究结果表明，二硫键形成的氧化当量是由氧通过有氧生长的大肠杆菌细胞呼吸电子传递系统提供的。DsbB中的Cys-41-Val-Leu-Cys-44基序被呼吸醌分子强烈氧化。我们的插入诱变结果表明，仅c端毗邻CXXC基序的片段对于DsbB的呼吸耦合氧化至关重要(4)Ala取代一个或所有的I le45-tyr46-Glu47-Arg48残基，这些残基由CXXC基序和假定的膜泛区标记，并没有消除呼吸耦合。相反，从该片段中删除一个或多个残基以及在其中插入一个或多个Ala会严重损害氧化。因此，后一种突变蛋白积累为还原形式或最后的特征二硫苏糖醇抗性。我们认为Ile45-tyr46-Glu47-Arg48片段的重要性可能在于它的物理长度，而不是每个氨基酸的化学性质。可以想象，CXXC基序相对于膜表面的适当定位对于其与膜锚定醌分子的有效相互作用至关重要。然后，泛醌泛醇与（CXXC）ox/(CXXC)red的氧化还原电位之间的巨大差异可以驱动该反应。

In the Escherichia coli protein disulphide bond formation pathway, a membrane protein DsbB reoxidizes reducd periplasmic DsbA, the disulphide boud-introducing enzyme. Our results indicate that the oxidizing equivalent for disulphide bond formation is provided by oxygen through the respiratory electron transfer system in aerobically growin E. coli cells. The Cys-41-Val-Leu-Cys-44 motif in DsbB is strongly oxidized by respiratory quinone molecules. The results of our insertion mutagenesis indicated that a segment just C-terminally adjacent to CXXC motif is crucial for the respiration-coupled oxidation of DsbB(4)Ala substitution for one or all of the I le45-tyr46-Glu47-Arg48 residues, which are franked by the CXXC motif and the presumed membrane-panning region, did not abolish the respiratory coupling. In contrast,deletion of one or more residues from this segment as well as insertions of one or more Ala into it severely impaired the oxidation.Thus,the latter mutant proteins accumulated as reduced forms or last the characteristic dithiothreitol resistance. We prose that the importance of the Ile45-tyr46-Glu47-Arg48 segment may lie in its physical length rather than the chemical nature of each amino acid. It may be conceivable that a proper positioning of the CXXC motif relative to the membrane surface is crucial for its efficient interaction with membrane-anchored quinonc molecules.The reaction Could then be driven by the large difference between the redox potentials of ubiquinone ubiquinol vs(CXXC)ox/(CXXC)red.