Effect of heptane treatment on the response of sarcoplasmic reticulum preparations to phosphate

The calcium-stimulated (extra) ATPase and calcium uptake activities of sarcoplasmic reticulum (SR) preparations treated with aqueous heptane mixtures were compared with those of untreated SR, and with those of SR treated with aqueous ether. Both treatments altered the kinetic behaviour of the extra ATPase, the Lineweaver-Burk plot being changed from its normal non-linear shape to a straight line. Kinetic constants, Vmax, Km for ATP and Ki for phosphate, were measured. The extra ATPase activity of heptane-treated SR was inhibited by phosphate as was that of ether-treated SR, to a lesser extent. The magnitude of this inhibition by phosphate was found to be considerably less than the degree of stimulation of the extra ATPase activity of untreated SR caused by phosphate through its calcium-precipitating action. The steady-state concentrations of the phosphoryl-enzyme intermediates were measured and together with the Km and Ki values they indicate that the bidning of ATP to heptane-treated SR is weaker than it is to untreated SR, and that phosphate is an efficient competitor for the binding sites.