Translocation of pro-OmpA across inner membrane vesicles of Escherichia coli occurs in two consecutive energetically distinct steps.
The rate of energy-dependent transfer of pro-OmpA across Escherichia coli inner membrane vesicles in vitro was found to be a function of the ATP concentration. At concentrations above 0.1 mM ATP, the addition of a transmembrane electrochemical potential (proton motive force or pmf) increased the rate of pro-OmpA translocation. Additional experiments demonstrated that the overall reaction proceeded by at least two distinct energy-requiring steps. The first step required only ATP, was nearly unaffected by the pmf, and resulted in the insertion of the amino-terminal domain of pro-OmpA across the membrane. The insertion exposed the signal sequence cleavage site to the periplasmic side of the membrane, as measured by the appearance of a mature length translocation intermediate. However, this intermediate was partially exposed to the cytoplasmic side of the membrane. In a second energy-dependent step, either ATP or the pmf was sufficient to complete the translocation of mature length OmpA across the membrane.
Geller
B L
BL
Department of Microbiology, Center for Gene Research and Biotechnology, Oregon State University, Corvallis 97331-3804.
Green
H M
HM
D000255:Adenosine Triphosphate / Q000378:metabolism*; D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism*; D007700:Kinetics; D008962:Models, Theoretical; D011498:Protein Precursors / Q000378:metabolism*; D013462:Sulfur Radioisotopes
D001425:Bacterial Outer Membrane Proteins; D011498:Protein Precursors; D013462:Sulfur Radioisotopes; D000255:Adenosine Triphosphate