ATP HYDROLYSIS-LINKED STRUCTURAL-CHANGES IN THE N-TERMINAL PART OF THE GAMMA-SUBUNIT OF ESCHERICHIA-COLI F1-ATPASE EXAMINED BY CROSS-LINKING STUDIES

A mutant of Escherichia coli F1-ATPase (ECF1) in which the serine resi due in position 8 of the gamma subunit has been replaced by a cysteine residue (gammaS8C) has been used to study nucleotide-dependent cross- linking of the gamma subunit to a beta subunit. When examined in the p resence of ADP + Mg2+, either supplied directly or as produced during catalytic turnover of ATP + Mg2+, the main cross-linked product genera ted using the heterobifunctional, photoactivatable, cross-linker tetra fluorophenylazide maleimide-6 had a M(r(app)) of 108,000. When ATP hyd rolysis was inhibited, either by cold or by reaction with sodium azide , or when ATP hydrolysis was prevented by the use of adenyl-5'-yl beta ,gamma-imidodiphosphate, the main cross-linked products were species w ith M(r(app)) of 102,000 and 84,000. The nucleotide-dependent switchin g from one cross-linking pattern to another could only be observed whe n the epsilon subunit was bound to ECF1; it was not seen in ECF1, an enzyme preparation missing delta and epsilon subunits, but was observe d in preparations selectively depleted of the delta subunit. We conclu de that the changes detected in these cross-linking experiments are oc curring during the hydrolysis of ATP when the beta-gamma phosphate bon d is cleaved and that they are related to the coupling of ATP hydrolys is to proton translocation.