title: Four cysteines of the membrane protein DsbB act in concert to oxidize its substrate DsbA.
abstract: 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.
authors: Department of Microbiology and Molecular Genetics, Harvard Medical School, 200 Longwood Avenue, Boston, MA 02115, USA.
authors: Hiroshi
authors: H
authors: Kadokura
authors: Jon
authors: J
authors: Beckwith
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D001483:Base Sequence; D002462:Cell Membrane / Q000201:enzymology; D003545:Cysteine; D017931:DNA Primers; D004579:Electron Transport; D004926:Escherichia coli / Q000201:enzymology*; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D011485:Protein Binding; D011487:Protein Conformation; D019704:Protein Disulfide-Isomerases / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D017931:DNA Primers; C079503:DsbB protein, Bacteria; D008565:Membrane Proteins; D019704:Protein Disulfide-Isomerases; D003545:Cysteine