The binding of a second divalent metal ion is necessary for the activation of ATP hydrolysis and its inhibition by tightly bound ADP in the ATPase from Halobacterium saccharovorum.

B Schobert

doi:10.1016/s0021-9258(19)50011-4

Abstract

A binding site for divalent metal ions on the ATPase from Halobacterium saccharovorum is proposed. This site is different from the catalytic site which binds ATP and a complexed divalent metal ion. Occupation of the second site greatly stimulates the rate of ATP hydrolysis and the affinity of the catalytic site for the metal ion-ATP complex. The time-dependent inhibition of the ATPase, which occurs during catalysis and which is known to be caused by the retention of ADP, is also dependent on the occupation of this metal ion binding site. The binding of the metal ion apparently induces extremely tight binding of ADP after the departure of Pi. Mg2+, Mn2+, Zn2+, Co2+, and Ca2+ were tested and showed both the activating and the inhibitory effects, although their binding constants for ATP and the second metal ion binding site were quite different. The characteristic shapes of the nonlinear ATP hydrolysis curves obtained with different metal ions, and different ratios of metal ion and ATP, could be explained with the established dissociation constants. On this basis, a model for the ATPase was developed with two catalytic cycles: one in which the second metal ion binding site is occupied, and another in which it is empty. These pathways are connected by metal ion-dependent equilibria.

Highlights

A binding site for divalent metal ions on the ATPasdeetect the short lag-phase, and, in an earlier investigation of from Halobacterium saccharovorumis proposed
The results demonstrate the functions of a divalent metal ion, associated with a site on the H. saccharouorum ATPase which is distinct from the Me
The effect of the bound metal ion wasboth to activate andto inhibit, depending on the stage in the catalytic cycle. Occupation of this metal ion binding site greatly stimulated the initial rate of ATP hydrolysis (Fig. 7, A and B )

Summary

Brigitte Schobert

The prerequisites for this tight binding of ADP On this basis, a model for the ATPase was developed are not clear; puzzling are the variations in the with two catalytic cycles: one in which the secondshampestaolf the nonlinear hydrolysis curves during different ion bindingsite is occupied, and another in whiciht is assay conditions (Schobert, 1991).it is noctlear empty. A model for the ATPase was developed are not clear; puzzling are the variations in the with two catalytic cycles: one in which the secondshampestaolf the nonlinear hydrolysis curves during different ion bindingsite is occupied, and another in whiciht is assay conditions (Schobert, 1991).it is noctlear empty These pathways are connected by metal ion- if the tight binding of the inhibitory ADP differs from the dependent equilibria.

RESULTS

Metal ion Medium

Metal ion at catalytic site

DISCUSSION