Apocytochrome c binding to negatively charged lipid dispersions studied by spin-label electron spin resonance.
The interaction of apocytochrome c with aqueous dispersions of phosphatidylserine from bovine spinal cord and with other negatively charged phospholipids has been studied as a function of pH and salt concentration by using spin-label electron spin resonance (ESR) spectroscopy and chemical binding assays. The ESR spectra of phospholipids spin-labeled at different positions on the sn-2 chain indicate a generalized decrease in mobility of the lipids, while the characteristic flexibility gradient toward the terminal methyl end of the chain is maintained, on binding of apocytochrome c to phosphatidylserine dispersions. This perturbation of the bulk lipid mobility or ordering is considerably greater than that observed on binding of cytochrome c. In addition, a second, more motionally restricted, lipid component is observed with lipids labeled close to the terminal methyl ends of the chains. This second component is not observed on binding of cytochrome c and can be taken as direct evidence for penetration of apocytochrome c into the lipid bilayer. It is less strongly motionally restricted than similar spectral components observed with integral membrane proteins and displays a steep flexibility gradient. The proportion of this second component increases with increasing protein-to-lipid ratio, but the stoichiometry per protein bound decreases from 4.5 lipids per 12 000-dalton protein at low protein contents to 2 lipids per protein at saturating amounts of protein. Apocytochrome c binding to phosphatidylserine dispersions decreases with increasing salt concentration from a saturation value corresponding to approximately 5 lipids per protein in the absence of salt to practically zero at 0.4 M NaCl.(ABSTRACT TRUNCATED AT 250 WORDS)
D000818:Animals; D001059:Apoproteins / Q000378:metabolism*; D003574:Cytochrome c Group / Q000378:metabolism*; D045304:Cytochromes c; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods; D006736:Horses; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008433:Mathematics; D009206:Myocardium / Q000378:metabolism; D010718:Phosphatidylserines / Q000378:metabolism*; D011485:Protein Binding
D001059:Apoproteins; D003574:Cytochrome c Group; D010718:Phosphatidylserines; D045304:Cytochromes c