A NON EQUILIBRIUM THERMODYNAMICS MODEL OF RECONSTITUTED CA2-ATPASE()

A non-equilibrium thermodynamics (NET) model describing the action of completely coupled or 'slipping' reconstituted Ca2+-ATPase is presente d. Variation of the coupling stoichiometries with the magnitude of the electrochemical gradients, as the ATPase hydrolyzes ATP, is an indica tion of molecular slip. However, the Ca2+ and H+ membrane-leak conduct ances may also be a function of their respective gradients. Such non-o hmic leak typically yields 'flow-force' relationships that are similar to those that are obtained when the pump slips; hence, caution needs to be exercised when interpreting data of Ca2+-ATPase-mediated fluxes that display a non-linear dependence on the electrochemical proton (De lta<(mu)over tilde>(H)) and/or calcium gradients (Delta<(mu)over tilde >(Ca)). To address this issue, three experimentally verifiable relatio nships differentiating between membrane leak and enzymic slip were der ived. First, by measuring Delta<(mu)over tilde>(H) as a function Of th e rate of ATP hydrolysis by the enzyme. Second, by measuring the overa ll 'efficiency' of the pump as a function of Delta<(mu)over tilde>(H). Third, by measuring the proton ejection rate by the pump as a functio n of its ATP hydrolysis rate.