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Abstract

Despite the thermodynamic problem imposed on alkaliphilic bacteria of synthesizing adenosine triphosphate (ATP) against a large inverted pH gradient and consequently a low electrochemical proton potential, these bacteria still utilize a proton-coupled F₁F_o-ATP synthase to synthesize ATP. One potential solution to this apparent thermodynamic problem would be the operation of a larger oligomeric c ring, which would raise the ion to ATP ratio, thus facilitating the conversion of a low electrochemical potential into a significant phosphorylation potential. To address this hypothesis, we have purified the oligomeric c ring from the thermoalkaliphilic bacterium Bacillus sp. strain TA2.A1 and determined the number of c-subunits using a novel mass spectrometry method, termed ‘laser-induced liquid bead ion desorption’ (LILBID). This technique allows the mass determination of non-covalently assembled, detergent-solubilized membrane protein complexes, and hence enables an accurate determination of c ring stoichiometries. We show that the Bacillus sp. strain TA2.A1 ATP synthase harbours a tridecameric c ring. The operation of a c ring with 13 subunits renders the thermodynamic problem of ATP synthesis at alkaline pH less severe and may represent a strategy for ATP synthesis at low electrochemical potential.