THE CONTROL OF PROTEIN SURFACE CONCENTRATION ON PROTEOLIPOSOMES

Proteoliposomes having surface-bound succinylated concanavalin A (sConA) have been prepared by sonication of small unilamellar vesicles (SUV) and reverse-phase evaporation (REV) from lipid mixtures of dipalmitoylphosphatidylcholine, phosphatidylinositol and the reactive lipid phosphatidylethanolamine derivatised with maleimidobenzoyl-N-hydroxy-succinimide. The liposomes were conjugated with protein by reaction with the N-succinimidyl-S-thioacetate (SATA) derivative of sConA and had weight-average diameters (d(w)BAR) in the range 45-260 nm, as determined by photon correlation spectroscopy, and weight-average numbers of protein molecules per proteoliposome (P(w)BAR) up to approximately 2000. The factors controlling the extent of conjugation, including activation of the SATA derivative of sConA, the molar ratio of SATA to sConA and the reactive lipid content of the liposomes, have been studied. The surface concentration of sConA is directly controllable by manipulation of the reactive lipid content of the proteoliposomes. The results are compared with previous studies on wheatgerm agglutinin-conjugated proteoliposomes, and it is shown that for both types of proteoliposome the area per protein in the liposomal surfaces reaches limiting values of approximately 100 nm2 for REV and 40 nm2 for SUV. This observation is discussed in terms of the projected excluded area of protein on the liposomal surface.