FATTY-ACID PH TITRATION AND THE SELECTIVITY OF INTERACTION WITH EXTRINSIC PROTEINS IN DIMYRISTOYLPHOSPHATIDYLGLYCEROL DISPERSIONS - SPIN LABEL ESR STUDIES

The effects of pH titration on the ESR spectra of 5-(4,4-dimethyloxazolidine-N-oxyl) stearic acid in bilayers of dimyristoyl phosphatidylglycerol complexed with either myelin basic protein, apocytochrome c, cytochrome c or lysozyme have been studied. Binding of the protein increases the outer hyperfine splitting of the stearic acid spin label to an extent which depends on the protonation state of the fatty acid carboxyl group for all proteins studied. The hyperfine splittings have been analysed to determine the relative selectivities of the protonated and unprotonated forms of the fatty acid by assuming that the spectra correspond to fast exchange on the ESR timescale. For myelin basic protein and apocytochrome c, the relative association constants are approximately 80% and 30% greater, respectively for the charged state than for the protonated state of the fatty acid. The pKa of the stearic acid spin label has been determined from the pH titration behaviour of the outer hyperfine splitting in the ESR spectra. The pKa at the lipid/water interface in the absence of protein is 8.0, and at the lipid/peripheral protein interface is 9.6, 9.2, 8.5 and 8.5 for the case of bilayers complexed with the myelin basic protein, apocytochrome c, cytochrome c and lysozyme, respectively. The large upward shifts in pKa are interpreted in terms of a strongly reduced polarity at the lipid/protein interface, suggesting that the dimyristoyl phosphatidylglycerol headgroups become largely dehydrated on binding of either myelin basic protein or apocytochrome c. The extent of this dehydraton is considerably less for cytochrome c and lysozyme. © 1990.