Integration of F1 and the membrane sector of the proton-ATPase of Escherichia coli. Role of subunit "b" (uncF protein).
Membranes derived from the Escherichia coli strain AN1460 which carries the multicopy plasmid pAN45 (unc+) (Downie, J. A., Langman, L., Cox, G. B., Yanofsky, C., and Gibson, (1980) J. Bacteriol. 143, 8-17) were enriched 5- to 10-fold in proton-ATPase activity. Incubation of F1-depleted AN1460 membranes with trypsin abolished F1-binding ability but did not inhibit proton transport through the membrane sector (F0). Sodium dodecyl sulfate-gel electrophoresis indicated that subunit "b" (uncF protein) of F0 was cleaved by trypsin and prebound F1 protected against the trypsin effect. Subunits "a" (uncB protein) and "c" (uncE protein) were unaffected by the trypsin treatment. A water-soluble fragment (Mr = 14,800) was liberated after trypsin treatment and appeared to arise from subunit b. Studies of enzyme hybridization and of F1 binding to membranes derived from strains containing mutations in uncB, F, and E genes supported the suggestion that subunit b is involved in F1 binding to the F0. Also, extraction of membranes with KSCN increased the relative proportion of subunit b in the membrane and this coincided with a parallel increase in trypsin-sensitive F1-binding ability. It is proposed that subunit b is involved in binding of F1 to the F0; this agrees with the presumed role of the protein as deduced from predictions of its secondary and tertiary structure (Walker, J. E., Saraste, M., and Gay, N. J. (1982) Nature (Lond.) 298, 867-869; Senior, A. E. (1983) Biochim. Biophys. Acta, in press).
D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D009876:Operon; D006180:Proton-Translocating ATPases; D014357:Trypsin / Q000378:metabolism
D001426:Bacterial Proteins; D046911:Macromolecular Substances; D014357:Trypsin; D000251:Adenosine Triphosphatases; D006180:Proton-Translocating ATPases