title: Covalently dimerized SecA is functional in protein translocation.
abstract: The ATPase SecA provides the driving force for the transport of secretory proteins across the cytoplasmic membrane of Escherichia coli. SecA exists as a dimer in solution, but the exact oligomeric state of SecA during membrane binding and preprotein translocation is a topic of debate. To study the requirements of oligomeric changes in SecA during protein translocation, a non-dissociable SecA dimer was formed by oxidation of the carboxyl-terminal cysteines. The cross-linked SecA dimer interacts with the SecYEG complex with a similar stoichiometry as non-cross-linked SecA. Cross-linking reversibly disrupts the SecB binding site on SecA. However, in the absence of SecB, the activity of the disulfide-bonded SecA dimer is indistinguishable from wild-type SecA. Moreover, SecYEG binding stabilizes a cold sodium dodecylsulfate-resistant dimeric state of SecA. The results demonstrate that dissociation of the SecA dimer is not an essential feature of the protein translocation reaction.
authors.affiliation: Department of Molecular Microbiology, Groningen Biomolecular Sciences and Biotechnology Institute and Materials and Science Centre Plus, University of Groningen, Kerklaan 30, 9751 NN Haren, The Netherlands.
authors.forename: Jeanine
authors.initials: J
authors.lastname: de Keyzer
authors.forename: Eli O
authors.initials: EO
authors.lastname: van der Sluis
authors.forename: Robin E J
authors.initials: RE
authors.lastname: Spelbrink
authors.forename: Niels
authors.initials: N
authors.lastname: Nijstad
authors.forename: Ben
authors.initials: B
authors.lastname: de Kruijff
authors.forename: Nico
authors.initials: N
authors.lastname: Nouwen
authors.forename: Chris
authors.initials: C
authors.lastname: van der Does
authors.forename: Arnold J M
authors.initials: AJ
authors.lastname: Driessen
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry*; D001426:Bacterial Proteins / Q000737:chemistry*; D002462:Cell Membrane / Q000378:metabolism; D003432:Cross-Linking Reagents / Q000494:pharmacology; D003545:Cysteine / Q000737:chemistry; D003593:Cytoplasm / Q000378:metabolism; D019281:Dimerization; D004220:Disulfides / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D026901:Membrane Transport Proteins / Q000737:chemistry*; D009154:Mutation; D010100:Oxygen / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D000069816:SEC Translocation Channels; D000081416:SecA Proteins; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry; D020349:Surface Plasmon Resonance; D013997:Time Factors; D014508:Urea / Q000494:pharmacology
chemical_list: D001426:Bacterial Proteins; D003432:Cross-Linking Reagents; D004220:Disulfides; D026901:Membrane Transport Proteins; D000069816:SEC Translocation Channels; D012967:Sodium Dodecyl Sulfate; D014508:Urea; D000251:Adenosine Triphosphatases; D000081416:SecA Proteins; D003545:Cysteine; D010100:Oxygen