Use of energy transfer to assay the asseciation of proteins with lipid membranes

Abstract

A method is described for the rapid and quantitative assay of the association of proteins with lipid membranes. The method utilizes the nonradiative transfer of energy between fluorescent groups in the protein and fluorescent groups attached to the polar heads of the lipids in the membrane. The association constant for the electrostatic association of trypsin with bilayers of the negatively charged lipid, 1,2-dimyristoyl- sn-glycerol-3-phosphoric acid methyl ester (MPA) at a salt concentration of 5 m m and pH 8 was found to be (2.32 ± 0.07) × 10 5 m −1. The stoichiometry of lipid to protein in the association complex is 40–50 lipid molecules per trypsin molecule. Honey-bee venom melittin and the serum apolipoprotein, apo-Lp-Ala, were shown to associate with lecithin vesicles. Acetylcholine esterase isolated from the electric organ of the electric eel according to the method of Leuzinger and Baker [(1967) Proc. Nat. Acad. Sci. USA 57, 446–451] was shown to be incapable of associating with lecithin or MPA bilayers.