The role of the mature domain of proOmpA in the translocation ATPase reaction.
The export of proOmpA, the precursor of outer membrane protein A from Escherichia coli, requires preprotein translocase, which is comprised of SecA, SecY/E, and acidic phospholipids. Previous studies of proOmpA translocation intermediates (Schiebel, E., Driessen, A. J. M., Hartl, F.-U., and Wickner, W. (1991) Cell 64, 927-939) suggested that the "slippage" of the translocating polypeptide chain and the high level of ATP hydrolysis, characteristic of the "translocation ATPase," were part of a futile cycle. To examine the role of the mature domain of proOmpA in its translocation-dependent ATP hydrolysis, we used chemical cleavage to generate NH2-terminal fragments of this preprotein. Each fragment contained the 21-residue leader region and either 53 or 228 residues of the mature domain (preproteins P74 and P249, respectively). As observed with full-length proOmpA, the translocation of each fragment requires ATP and both the SecA and SecY/E domains of translocase and is stimulated by the transmembrane proton electrochemical gradient. The apparent maximal velocities of P74 and proOmpA translocation are similar. While the translocation of P74 and of proOmpA show the same apparent Km for ATP, far less ATP is hydrolyzed during the translocation of P74. Thus, the mature carboxyl-terminal domain of proOmpA has a major role in supporting the translocation ATPase.
Bassilana
M
M
Molecular Biology Institute, University of California, Los Angeles 90024-1570.
Arkowitz
R A
RA
Wickner
W
W
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D001425:Bacterial Outer Membrane Proteins / Q000302:isolation & purification / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins; D007700:Kinetics; D008565:Membrane Proteins; D008970:Molecular Weight; D010446:Peptide Fragments / Q000302:isolation & purification; D011498:Protein Precursors / Q000302:isolation & purification / Q000378:metabolism*; D012697:Serine Endopeptidases
D001425:Bacterial Outer Membrane Proteins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D010446:Peptide Fragments; D011498:Protein Precursors; C057184:outer membrane protein A precursor (E coli); D000255:Adenosine Triphosphate; D010450:Endopeptidases; D012697:Serine Endopeptidases; C026515:type I signal peptidase; D000251:Adenosine Triphosphatases