Identification of two interaction sites in SecY that are important for the functional interaction with SecA.
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.
van der Sluis
Eli O
EO
Department of Molecular Microbiology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, 9751 NN Haren, The Netherlands.
Nouwen
Nico
N
Koch
Joachim
J
de Keyzer
Jeanine
J
van der Does
Chris
C
Tampé
Robert
R
Driessen
Arnold J M
AJ
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
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