Lysine 155 in beta-subunit is a catalytic residue of Escherichia coli F1 ATPase.

Abstract

The "homology A" ("glycine-rich" or "P-loop") consensus sequence occurs in the catalytic sites of F1F0 ATP synthase enzymes. The conserved lysine of this motif is beta-subunit Lys-155 in Escherichia coli F1. The role of this lysine in binding and catalysis at the high affinity ATP binding site was studied with the mutants beta K155Q and beta K155E by measuring the rates of ATP binding/release, ATP hydrolysis/synthesis, and Pi release as a function of pH varied from 5.5 to 9.5. In wild type, protonated beta Lys-155 appears to contribute significantly to high affinity binding of ATP, probably through hydrogen bonding to the gamma-phosphate. ATP hydrolysis and synthesis were impaired strongly in the mutants, and the reaction equilibrium constant, which was pH-independent in wild type, was highly pH-dependent in beta K155Q and beta K155E. Studies of steady-state ATPase turnover showed that positive catalytic cooperativity was virtually absent and the pH-dependent component of positive catalytic cooperativity was abolished or reversed in the mutants. The data demonstrate that residue beta K155 is a critical catalytic residue in F1 ATPase.