31P nuclear magnetic resonance studies of the association of basic proteins with multilayers of diacyl phosphatidylserine

Abstract

Lysozyme, cytochrome c, poly( l-lysine), myelin basic protein and ribonuclease were used to form multilayer dispersions containing about 50% protein (by weight) with bovine brain diacyl phosphatidylserine (PS). 31P nuclear magnetic resonance shift anisotropies, spin-spin ( T 2) and spin-lattice ( T 1) relaxation times for the lipid headgroup phosphorus were measured at 36.44 MHz. At pH 7.5, lysozyme, cytochrome c, poly( l-lysine) and ribonuclease were shown to increase the chemical shift anisotropy of PS by between 12–20%. Myelin basic protein altered the shape of the phosphate resonance, suggesting the presence of two lipid components, one of which had a modified headgroup conformation. The presence of cytochrome c led to the formation of a narrow spike at the isotropic shift position of the spectrum. Of the various proteins or peptides we have studied, only poly( l-lysine) and cytochrome c had any effect on the T 1 of PS (1050 ms). Both caused a 20–30% decrease in T 1 of the lamellar-phase phosphate peak. The narrow peak in the presence of cytochrome c had a very short T 1 of 156 ms. The possibility is considered that the cytochrome Fe 3+ contributes to the phosphate relaxation in this case. The effect of all proteins on the T 2 of the phosphorus resonance was to cause an increase from the value for pure PS (1.6 ms) to between 2 and 5 ms. The results obtained with proteins are compared with the effects of small ions and intrinsic membrane proteins on the order and motion of the headgroups of lipids in bilayers.