A kinetic model for the Ca2+ + Mg2+-activated ATPase of sarcoplasmic reticulum

Abstract

The Ca2+ + Mg2+-activated ATPase of sarcoplasmic reticulum exhibits complex kinetics of activation with respect to ATP. ATPase activity is pH-dependent, with similar pH-activity profiles at high and low concentrations of ATP. Low concentrations of Ca2+ in the micromolar range activate the ATPase, whereas activity is inhibited by Ca2+ at millimolar concentrations. The pH-dependence of this Ca2+ inhibition and the effect of the detergent C12E8 (dodecyl octaethylene glycol monoether) on Ca2+ inhibition are similar to those observed on activation by low concentrations of Ca2+. On the basis of these and other studies we present a kinetic model for the ATPase. The ATPase is postulated to exist in one of two conformations: a conformation (E1) of high affinity for Ca2+ and MgATP and a conformation (E2) of low affinity for Ca2+ and MgATP. Ca2+ binding to E2 and to the phosphorylated form E2P are equal. Proton binding at the Ca2+-binding sites in the E1 and E2 conformations explains the pH-dependence of Ca2+ effects. Binding of MgATP to the phosphorylated intermediate E1'PCa2 and to E2 modulate the rates of the transport step E1'PCa-E2'PCa2 and the return of the empty Ca2+ sites to the outside surface of the sarcoplasmic reticulum, as well as the rate of dephosphorylation of E2P. Only a single binding site for MgATP is postulated.

An investigation of functional similarities between the sarcoplasmic reticulum and platelet calcium-dependent adenosinetriphosphatases with the inhibitors quercetin and calmidazolium

“…In Figure 1, substrate ligation occurs in a random order, with the preferred pathway (or combination of pathways) being de-termined by the concentration of ATP and calcium. This type of ligation scheme has been proposed (Inesi et al, 1980;Tanford et al, 1985;Reynolds et al, 1985;Gould et al, 1986) to account for transient and steady-state kinetic measurements (Neet & Green, 1977;Inesi et al, 1980;Moller et al, 1980;Gould et al, 1986) involving substrate activation of the sarcoplasmic reticulum Ca 2+ -ATPase. The scheme in Figure 1 includes only steps that can be probed by varying calcium and ATP concentration on a steady-state time scale.…”

Modelling coral polyp calcification in relation to ocean acidification

“…Kinetic activity measurements reveal saturating activity of the Ca-ATPase with increasing calcium concentrations, typical for ion transporters (Ip et al, 1991). Furthermore, the removal of protons from the calcifying fluid is likely connected to the activity of the calcium pump as the Ca 2+ -mediated-ATPase in eukaryotic cell membranes acts as a proton-calcium antiporter (Gould et al, 1986;MacLennan et al, 1997;Allemand et al, 2004). Therefore, the assumption of either a weak or a strong proton pump, or the maintenance of a constant proton gradient in corals (Ries, 2011) has implications for the calcium transport.…”

Probing the nucleotide‐binding site of sarcoplasmic reticulum (Ca2+‐Mg2+)‐ATPase with anti‐fluorescein antibodies

“…All members of this class are believed to pump cations by a similar mechanism involving two conformational extremes of the enzyme, designated E1 and E2. In the case of (Ca2+-Mg2+)-ATPase it is the phosphorylation of the E1 form of the ATPase (with its 2 high affinity outward facing calciumbinding sites) by ATP to give Ca2E1-P and the conformational change to give Ca2E2-P (with its 2 low affinity outward facing calcium-binding sites) which constitutes the transport event [2][3][4].…”