Mixed membranes of sphingolipids and glycerolipids as studied by spin-label ESR spectroscopy. A search for domain formation

The temperature dependences of the ESR spectra from different positional is omers of sphingomyelin and of phosphatidylcholine spin-labeled in their acy l chain have been compared in mixed membranes composed of sphingolipids and glycerolipids. The purpose of the study was to identify the possible forma tion of sphingolipid-rich in-plane membrane domains. The principal mixtures that were studied contained sphingomyelin and the corresponding glycerolip id phosphatidylcholine, both from egg yolk. Other sphingolipids that were i nvestigated were brain cerebrosides and brain gangliosides, in addition to sphingomyelins from brain and mill;. The outer hyperfine splittings in the ESR spectra of sphingomyelin and of phosphatidylcholine spin-labeled on C-5 of the acyl chain were consistent with mixing of the sphingolipid and glyc erolipid components, in fluid-phase membranes. In the gel phase of egg sphi ngomyelin and its mixtures with phosphatidylcholine, the outer hyperfine sp littings of sphingomyelin spin-labeled at C-14 of the acyl chain of sphingo myelin are smaller than those of the corresponding sn-2 chain spin-labeled phosphatidylcholine. This is in contrast to the situation with sphingomyeli n and phosphatidylcholine spin-labeled at C-5, for which the outer hyperfin e splitting is always greater for the spin-labeled sphingomyelin. The behav ior of the C-14 spin-labels is attributed to a different geometry of the ac yl chain attachments of the sphingolipids and glycerolipids that is consist ent with their respective crystal structures. The two-component ESR spectra of sphingomyelin and phosphatidylcholine spin-labeled at C-14 of the acyl chain directly demonstrate a broad two-phase region with coexisting gel and fluid domains in sphingolipid mixtures with phosphatidylcholine. Domain fo rmation in membranes composed of sphingolipids and glycerolipids alone is r elated primarily to the higher chain-melting transition temperature of the sphingolipid component.