Reversible, nonionic, and pH-dependent association of cytochrome c with cardiolipin-phosphatidylcholine liposomes.
Membrane association of cytochrome c (cyt c) was monitored by the efficiency of resonance energy transfer from a pyrene-fatty acid containing phospholipid derivative (1-palmitoyl-2[6-(pyren-1-yl)]hexanoyl-sn-glycero-3-phosphocholine (PPHPC)) to the heme of cyt c. Liposomes consisted of 85 mol% egg phosphatidylcholine (egg PC), 10 mol% cardiolipin, and 5 mol% PPHPC. Cardiolipin was necessary for the membrane binding of cyt c over the pH range studied, from 4 to 7. In accordance with the electrostatic nature of the membrane association of cyt c at neutral pH both 2 mM MgCl2 and 80 mM NaCl dissociated cyt c from the vesicles completely. At neutral pH also adenine nucleotides in millimolar concentrations were able to displace cyt c from liposomes, their efficiency decreasing in the sequence ATP > ADP > AMP. In addition, both CTP and GTP were equally effective as ATP. The detachment of cyt c from liposomes by nucleotides is likely to result from a competition between cardiolipin and the nucleotides for a common binding site in cyt c. When pH was decreased to 4 there was a small yet significant increase in the apparent affinity of cyt c to cardiolipin containing liposomes. Notably, at pH 4 the above nucleotides as well as NaCl and MgCl2 were no longer able to dissociate cyt c and, on the contrary, they slightly enhanced the quenching of pyrene fluorescence by cyt c. The above results do suggest that the membrane association of cyt c at acidic pH was non-ionic and presumably due to hydrogen bonding. The pH-dependent binding of cyt c to membranes was fully reversible. Accordingly, in the presence of sufficient concentrations of either nucleotides or salts rapid detachment and membrane association of cyt c could be induced by varying pH between neutral and acidic values, respectively.
Rytömaa
M
M
Department of Medical Chemistry, University of Helsinki, Finland.
Mustonen
P
P
Kinnunen
P K
PK
D000227:Adenine Nucleotides / Q000494:pharmacology; D002308:Cardiolipins / Q000737:chemistry*; D003570:Cytidine Triphosphate / Q000494:pharmacology; D003574:Cytochrome c Group / Q000737:chemistry*; D006160:Guanosine Triphosphate / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D008051:Lipid Bilayers / Q000737:chemistry; D008081:Liposomes; D010713:Phosphatidylcholines / Q000737:chemistry*; D012965:Sodium Chloride / Q000494:pharmacology; D013050:Spectrometry, Fluorescence
D000227:Adenine Nucleotides; D002308:Cardiolipins; D003574:Cytochrome c Group; D008051:Lipid Bilayers; D008081:Liposomes; D010713:Phosphatidylcholines; D012965:Sodium Chloride; D003570:Cytidine Triphosphate; D006160:Guanosine Triphosphate