A single amino acid substitution in SecY stabilizes the interaction with SecA.
The SecYEG complex constitutes a protein conducting channel across the bacterial cytoplasmic membrane. It binds the peripheral ATPase SecA to form the translocase. When isoleucine 278 in transmembrane segment 7 of the SecY subunit was replaced by a unique cysteine, SecYEG supported an increased preprotein translocation and SecA translocation ATPase activity, and allowed translocation of a preprotein with a defective signal sequence. SecY(I278C)EG binds SecA with a higher affinity than normal SecYEG, in particular in the presence of ATP. The increased translocation activity of SecY(I278C)EG was confirmed in a purified system consisting of SecYEG proteoliposomes, while immunoprecipitation in detergent solution reveal that translocase-preprotein complexes are more stable with SecY(I278C) than with normal SecY. These data imply an important role for SecY transmembrane segment 7 in SecA binding. As improved SecA binding to SecY was also observed with the prlA4 suppressor mutation, it may be a general mechanism underlying signal sequence suppression.
Manting
E H
EH
Department of Microbiology and Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Kerklaan 30, 9751 NN Haren, The Netherlands.
Kaufmann
A
A
van der Does
C
C
Driessen
A J
AJ
D000251:Adenosine Triphosphatases / Q000737:chemistry* / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D001692:Biological Transport; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D008969:Molecular Sequence Data; D000069816:SEC Translocation Channels; D000081416:SecA Proteins; D013329:Structure-Activity Relationship
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D000069816:SEC Translocation Channels; C057841:SecY protein, E coli; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D000081416:SecA Proteins