Recent studies have revealed numerous examples in which oxidation and reduction of cysteines in proteins are integrated into specific cascades of biological regulatory systems. In general, these reactions proceed as thiol-disulfide exchange events. However, it is not exactly understood how a disulfide bond is created de novo. DsbB, an Escherichia coli plasma membrane protein, is one of the enzymes that create a new disulfide bond within itself and in DsbA, the direct catalyst of protein disulfide bond formation in the periplasmic space. DsbB is associated with a cofactor, either ubiquinone or menaquinone, as a source of an oxidizing equivalent. The DsbB-bound quinone undergoes transition to a pink (λmax, ≈500 nm, ubiquinone) or violet (λmax, ≈550 nm, menaquinone)-colored state during the course of the DsbB enzymatic reaction. Here we show that not only the thiolate form of Cys-44 previously suggested but also Arg-48 in the α-helical arrangement is essential for the quinone transition. Quantum chemical simulations indicate that proper positioning of thiolate anion and ubiquinone in conjunction with positively charged guanidinium moiety of arginine allows the formation of a thiolate-ubiquinone charge transfer complex with absorption peaks at ≈500 nm as well as a cysteinylquinone covalent adduct. We propose that the charge transfer state leads to the transition state adduct that accepts a nucleophilic attack from another cysteine to generate a disulfide bond de novo. A similar mechanism is conceivable for a class of eukaryotic dithiol oxidases having a FAD cofactor.
|Original language||English (US)|
|Number of pages||6|
|Journal||Proceedings of the National Academy of Sciences of the United States of America|
|State||Published - Jan 10 2006|
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