The F1-Fo ATP synthase bears 6 nucleotide binding sites, only 3 of which turn over during catalysis. The remaining 3 are occupied by slowly exchanging ATP in vivo, although at least 1 molecule is generally lost on isolation of the enzyme in the absence of nucleotide. It is proposed that the function of the slowly exchanging (NC) nucleotides is to participate in catalysis, the terminal phosphate of the bound ATP acting as an acid catalyst in the cleavage/synthesis of the phosphate anhydride bond in the catalytic sites. Such a role has been demonstrated for the bound pyridoxal phosphate moiety in glycogen phosphorylase. Evidence is presented that (i) the NC nucleotide spans the interface between an α subunit and its partner β, interacting near the catalytic binding site on β; (ii) the phosphate moieties of the catalyzed and NC nucleotide are close in space; and (iii) occupation of the NC nucleotide sites promotes ATP hydrolysis by F1 or its subfragments. All of these findings are required by the proposed mechanism. Relationships between phosphorylase and F1 structures are discussed.
