title: The membrane ATPase of alkalophilic Bacillus firmus RAB is an F1-type ATPase.
abstract: At the optimal pH for growth (pH 10.5), alkalophilic Bacillus firmus RAB, an obligate aerobe, exhibits normal rates of oxidative phosphorylation despite the low transmembrane proton electrochemical gradient, about -60 mV (delta psi = -180 mV and delta pH = +120 mV). This bioenergetic problem might be resolved by use of an Na+ coupled ATP synthase; otherwise an F1F0-ATPase must be able to utilize low driving forces in this organism. The ATPase activity was extracted from everted membrane vesicles by low ionic strength treatment and purified to homogeneity by hydrophobic interaction chromatography and sucrose density gradient centrifugation. The ATPase preparation had the characteristic F1-ATPase subunit structure, with Mr values of 51,500 (alpha), 48,900 (beta), 34,400 (gamma), 23,300 (delta), and 14,500 (epsilon); the identity of the alpha and beta subunits was confirmed by immunoblotting with anti-beta of Escherichia coli and anti-B. firmus RAB F1. Methanol and octyl glucoside, agents that stimulated the low basal membrane ATPase activity 10- to 12-fold, dramatically elevated the MgATPase activity of the purified F1, more than 150-fold, to 50 mumol min-1 mg protein-1. Anti-F1 inhibited membrane ATPase activity greater than or equal to 80%. The membranes exhibited no Na+-stimulated or vanadate-sensitive ATPase activity when prepared in the absence or presence of Na+ or ATP. These findings, which are consistent with previous studies, establish that in alkalophilic bacteria, ATP hydrolysis, and presumably ATP synthesis is catalyzed by an F1F0-ATPase rather than a Na+ ATPase.
mesh_terms: D001407:Bacillus / Q000201:enzymology*; D017301:Ca(2+) Mg(2+)-ATPase / Q000378:metabolism; D004563:Electrochemistry; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D007163:Immunosorbent Techniques; D008670:Metals / Q000378:metabolism; D010085:Oxidative Phosphorylation; D011188:Potassium / Q000378:metabolism; D006180:Proton-Translocating ATPases / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism
chemical_list: D008670:Metals; D012964:Sodium; D017301:Ca(2+) Mg(2+)-ATPase; D006180:Proton-Translocating ATPases; D011188:Potassium