title: Redox-active cysteines of a membrane electron transporter DsbD show dual compartment accessibility.
abstract: The membrane-embedded domain of the unusual electron transporter DsbD (DsbDbeta) uses two redox-active cysteines to catalyze electron transfer between thioredoxin-fold polypeptides on opposite sides of the bacterial cytoplasmic membrane. How the electrons are transferred across the membrane is unknown. Here, we show that DsbDbeta displays an inherent functional and structural symmetry: first, the two cysteines of DsbDbeta can be alkylated from both the cytoplasm and the periplasm. Second, when the two cysteines are disulfide-bonded, cysteine scanning shows that the C-terminal halves of the cysteine-containing transmembrane segments 1 and 4 are exposed to the aqueous environment while the N-terminal halves are not. Third, proline residues located pseudo-symmetrically around the two cysteines are required for redox activity and accessibility of the cysteines. Fourth, mixed disulfide complexes, apparent intermediates in the electron transfer process, are detected between DsbDbeta and thioredoxin molecules on each side of the membrane. We propose a model where the two redox-active cysteines are located at the center of the membrane, accessible on both sides of the membrane to the thioredoxin proteins.
authors.affiliation: Department of Microbiology and Molecular Genetics, Harvard Medical School, Harvard University, Boston, MA 02115, USA.
authors.forename: Seung-Hyun
authors.initials: SH
authors.lastname: Cho
authors.forename: Amir
authors.initials: A
authors.lastname: Porat
authors.forename: Jiqing
authors.initials: J
authors.lastname: Ye
authors.forename: Jon
authors.initials: J
authors.lastname: Beckwith
mesh_terms: D000595:Amino Acid Sequence; D003545:Cysteine / Q000737:chemistry*; D004220:Disulfides / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D017386:Sequence Homology, Amino Acid
chemical_list: D004220:Disulfides; D026901:Membrane Transport Proteins; D003545:Cysteine