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Four cysteines of the membrane protein DsbB act in concert to oxidize its substrate DsbA
Author(s) -
Kadokura Hiroshi,
Beckwith Jon
Publication year - 2002
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1093/emboj/21.10.2354
Subject(s) - dsba , periplasmic space , protein disulfide isomerase , cysteine , oxidoreductase , redox , biology , escherichia coli , biochemistry , biophysics , chemistry , disulfide bond , enzyme , organic chemistry , gene
Protein disulfide bond formation in Escherichia coli is catalyzed by the periplasmic protein DsbA. A cytoplasmic membrane protein DsbB maintains DsbA in the oxidized state by transferring electrons from DsbA to quinones in the respiratory chain. Here we show that DsbB activity can be reconstituted by co‐expression of N‐ and C‐terminal fragments of the protein, each containing one of its redox‐active disulfide bonds. This system has allowed us (i) to demonstrate that the two DsbB redox centers interact directly through a disulfide bond formed between the two DsbB domains and (ii) to identify the specific cysteine residues involved in this covalent interaction. Moreover, we are able to capture an intermediate in the process of electron transfer from one redox center to the other. These results lead us to propose a model that describes how the cysteines cooperate in the early stages of oxidation of DsbA. DsbB appears to adopt a novel mechanism to oxidize DsbA, using its two pairs of cysteines in a coordinated reaction to accept electrons from the active cysteines in DsbA.