Nonlinear electron paramagnetic resonance studies of the interaction of cytochrome c oxidase with spin-labeled lipids in gel-phase membranes

The interaction of lipids, spin-labeled at different positions in the sn-2 chain, with cytochrome c oxidase reconstituted in gel-phase membranes of di myristoylphosphatidylglycerol has been studied by electron paramagnetic res onance (EPR) spectroscopy. Nonlinear EPR methods, both saturation transfer EPR and progressive saturation EPR, were used. Interaction with the protein largely removes the flexibility gradient of the lipid chains in gel-phase membranes. The rotational mobility of the chain segments is reduced, relati ve to that for gel-phase lipids, by the intramembranous interaction with cy tochrome c oxidase. This holds for all positions of chain labeling, but the relative effect is greater for chain segments closer to the terminal methy l ends. Modification of the paramagnetic metal-ion centers in the protein b y binding azide has a pronounced effect on the spin-lattice relaxation of t he lipid spin labels. This demonstrates that the centers modified are suffi ciently close to the first-shell lipids to give appreciable dipolar interac tions and that their vertical location in the membrane is closer to the 5-p osition than to the 14-position of the lipid chains.