Stoichiometry of lipid–protein interaction assessed by hydrophobic photolabeling

Here we undertook a comparative study of the composition of the lipid annulus of three ATPases pertaining to the P-type family: plasma membrane calcium pump (PMCA), sarcoplasmic reticulum calcium pump (SERCA) and Na,K-ATPase. The photoactivatable phosphatidylcholine analogue [ 125I]TID-PC/16 was incorporated into mixtures of dimyristoyl phosphatidylcholine (DMPC) and each enzyme with the aid of the nonionic detergent C 12E 10. After photolysis, the extent of the labeling reaction was assessed to determine the lipid:protein stoichiometry: 17 for PMCA, 18 for SERCA, 24 for the Na,K-ATPase (α-subunit) and 5.6 mol PC/mol protein for the Na,K-ATPase (β-subunit).