Domain formation in sphingomyelin/cholesterol mixed membranes studied by spin-label electron spin resonance spectroscopy.
Interactions of palmitoylsphingomyelin with cholesterol in multilamellar vesicles have been studied over a wide range of compositions and temperatures in excess water by using electron spin resonance (ESR) spectroscopy. Spin labels bearing the nitroxide free radical group on the 5 or 14 C-atom in either the sn-2 stearoyl chain of phosphatidylcholine (predominantly 1-palmitoyl) or the N-stearoyl chain of sphingomyelin were used to determine the mobility and ordering of the lipids in the different phases. Two-component ESR spectra of the 14-position spin labels demonstrate the coexistence first of gel (L(beta)) and liquid-ordered (L(o)) phases and then of liquid-ordered and liquid-disordered (L(alpha)) phases, with progressively increasing temperature. These phase coexistences are detected over a limited range of cholesterol contents. ESR spectra of the 5-position spin labels register an abrupt increase in ordering at the L(alpha)-L(o) transition and a biphasic response at the L(beta)-L(o) transition. Differences in outer splitting between the C14-labeled sphingomyelin and phosphatidylcholine probes are attributed to partial interdigitation of the sphingomyelin N-acyl chains across the bilayer plane in the L(o) state. In the region where the two fluid phases, L(alpha) and L(o), coexist, the rate at which lipids exchange between phases (<<7 x 10(7) s(-)(1)) is much slower than translational rates in the L(alpha) phase, which facilitates resolution of two-component spectra.
Collado
M Isabel
MI
Departamento de Bioquímica, Universidad del País Vasco, Apartado 644, E-48080 Bilbao, Spain.
Goñi
Félix M
FM
Alonso
Alicia
A
Marsh
Derek
D
D002784:Cholesterol / Q000737:chemistry* / Q000378:metabolism; D003497:Cyclic N-Oxides / Q000378:metabolism; D005782:Gels; D008051:Lipid Bilayers / Q000737:chemistry* / Q000378:metabolism; D044367:Phase Transition; D010713:Phosphatidylcholines / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D013109:Sphingomyelins / Q000737:chemistry* / Q000378:metabolism; D013113:Spin Labels; D013696:Temperature
C025274:2-(16'-(N-oxyl-4',4'-dimethyloxazolidine)stearoyl)phosphatidylcholine; D003497:Cyclic N-Oxides; D005782:Gels; D008051:Lipid Bilayers; D010713:Phosphatidylcholines; D013109:Sphingomyelins; D013113:Spin Labels; C033171:palmitoylsphingomyelin; D002784:Cholesterol