CONFORMATIONAL-CHANGES IN THE ESCHERICHIA-COLI ATP SYNTHASE (ECF(1)F(0)) MONITORED BY NUCLEOTIDE-DEPENDENT DIFFERENCES IN THE REACTIVITY OFCYS-87 OF THE GAMMA-SUBUNIT IN THE MUTANT BETA-GLU-381-]ALA

Cys-87, one of two intrinsic cysteines of the gamma subunit of the Esc herichia coli ATP synthase (ECF(1)F(0)), is in a short segment of this subunit that binds to the bottom domain of a beta subunit close to a glutamate (Glu-381), Cys-87 was unreactive to maleimides under all con ditions in wild-type ECF(1) and ECF(1)F(0) but became reactive when Gl u-381 of beta was replaced by a cysteine or alanine, The reactivity of Cys-87 with maleimides was nucleotide-dependent, occurring with ATP o r ADP + EDTA in catalytic sites, in the presence of AMP PNP + Mg2+ but not with ADP + Mg2+ bound, whether P-i was present or not, and not wh en nucleotide binding sites were empty, Binding of N-ethylmaleimide ha d no effect, whereas 7 diethyl-amino 3-(4'-maleimidylphenyl)-4-methylc oumarin increased the ATPase activity of ECF, more than 2-fold by reac tion with Cys-87, In ECF(1)F(0), these reagents inhibited activity, Th e nucleotide dependence of the reaction of Cys-87 of the gamma subunit depended on the presence of the epsilon subunit, In epsilon subunit-f ree ECF(1), maleimides reacted with Cys-87 under all nucleotide condit ions, including when catalytic sites were empty, These results are dis cussed in terms of nucleotide-dependent movements of the gamma subunit during functioning of the F1F0-type ATPase.