title: Identification of two interaction sites in SecY that are important for the functional interaction with SecA.
abstract: The motor protein SecA drives the translocation of (pre-)proteins across the SecYEG channel in the bacterial cytoplasmic membrane by nucleotide-dependent cycles of conformational changes often referred to as membrane insertion/de-insertion. Despite structural data on SecA and an archaeal homolog of SecYEG, the identity of the sites of interaction between SecA and SecYEG are unknown. Here, we show that SecA can be cross-linked to several residues in cytoplasmic loop 5 (C5) of SecY, and that SecA directly interacts with a part of transmembrane segment 4 (TMS4) of SecY that is buried in the membrane region of SecYEG. Mutagenesis of either the conserved Arg357 in C5 or Glu176 in TMS4 interferes with the catalytic activity of SecA but not with binding of SecA to SecYEG. Our data explain how conformational changes in SecA could be directly coupled to the previously proposed opening mechanism of the SecYEG channel.
authors.affiliation: Department of Molecular Microbiology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, 9751 NN Haren, The Netherlands.
authors.forename: Eli O
authors.initials: EO
authors.lastname: van der Sluis
authors.forename: Nico
authors.initials: N
authors.lastname: Nouwen
authors.forename: Joachim
authors.initials: J
authors.lastname: Koch
authors.forename: Jeanine
authors.initials: J
authors.lastname: de Keyzer
authors.forename: Chris
authors.initials: C
authors.lastname: van der Does
authors.forename: Robert
authors.initials: R
authors.lastname: Tampé
authors.forename: Arnold J M
authors.initials: AJ
authors.lastname: Driessen
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000378:metabolism*; D000595:Amino Acid Sequence; D001120:Arginine / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D003545:Cysteine / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism*; D005973:Glutamine / Q000378:metabolism; D007700:Kinetics; D026901:Membrane Transport Proteins / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010455:Peptides / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary; D000069816:SEC Translocation Channels; D000081416:SecA Proteins; D013329:Structure-Activity Relationship
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D010455:Peptides; D000069816:SEC Translocation Channels; C065493:SecE protein, E coli; C057841:SecY protein, E coli; D005973:Glutamine; D001120:Arginine; D000251:Adenosine Triphosphatases; D000081416:SecA Proteins; D003545:Cysteine