Conformational changes in the Escherichia coli ATP synthase (ECF(1)F(0)) monitored by nucleotide-dependent differences in the reactivity of Cys-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 Escherichia 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 conditions in wild-type ECF(1) and ECF(1)F(0) but became reactive when Glu-381 of beta was replaced by a cysteine or alanine, The reactivity of Cys-87 with maleimides was nucleotide-dependent, occurring with ATP or 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 when nucleotide binding sites were empty, Binding of N-ethylmaleimide had no effect, whereas 7 diethyl-amino 3-(4'-maleimidylphenyl)-4-methylcoumarin increased the ATPase activity of ECF, more than 2-fold by reaction with Cys-87, In ECF(1)F(0), these reagents inhibited activity, The nucleotide dependence of the reaction of Cys-87 of the gamma subunit depended on the presence of the epsilon subunit, In epsilon subunit-free ECF(1), maleimides reacted with Cys-87 under all nucleotide conditions, including when catalytic sites were empty, These results are discussed in terms of nucleotide-dependent movements of the gamma subunit during functioning of the F1F0-type ATPase.