Cholesterol modulation of molecular activity of reconstituted shark Na+,K(+)-ATPase.
The cholesterol content of liposome bilayers has been varied between 0-40 mol% to study the effects on reconstituted Na+,K(+)-ATPase. The maximum hydrolytic activity of reconstituted Na+,K(+)-ATPase was increased by cholesterol at concentrations above 10 mol% for both the physiological Na+/K(+)-exchange reactions, as well as for the partial reactions Na+/Na(+)-exchange and uncoupled Na+ efflux. Omission of cholesterol from the liposome bilayer modified the activation by cytoplasmic Na+, indicating effects on both Vmax and on the Na(+)-affinity. Several other kinetic parameters were found to be strongly influenced as well, most notable the steady-state phosphorylation level, and the characteristics of the phosphorylation/dephosphorylation reactions. These results indicate that cholesterol interacts directly with the Na+,K(+)-ATPase as an essential effector perhaps by affecting its conformational mobility or monomer interaction.