Further study on the role of Mg2+ in lipid-protein interaction in reconstituted porcine heart mitochondrial H+-ATPase.
Porcine heart mitochondrial H+-ATPase was reconstituted by cholate dialysis method in liposomes containing neutral (PC, PE), acidic (PG, PI, PA, PS, DPG) or neutral and acidic phospholipids. The Mg2+ effect on the ATPase activity and its sensitivity to oligomycin, ATP-induced delta psi and delta pH formation was observed for the proteoliposomes containing acidic but not neutral phospholipids. Maleimide spin labels with varying arm lengths or bromoacetamide spin probe were used to monitor the conformational difference of H+-ATPase in the Mg2+-containing and Mg2+-'free' samples. A difference in W/S ratio (weakly immobilized/strongly immobilized component in the ESR spectra) could be detected for the F0.F1-containing and F1-depleted, (F0)-containing proteoliposomes, suggesting conformational difference in the F0-F1 complex and F0 portion induced by the Mg2+ effect. A conformational change of the beta-subunits in the F1 portion was also deduced from the ATP-induced fluorescence quenching of aurovertin-complex for Mg2+-containing samples. The results obtained are in favor of our previous assumption that Mg2+ may play its role by altering the physical state of the lipid bilayer, which would induce a conformational change in F0 (buried in the lipid core), which in turn is transmitted to the catalytic F1, resulting in a higher enzyme activity.
Zhang
X F
XF
Institute of Biophysics, Academia Sinica, Beijing, People's Republic of China.
Yang
F Y
FY
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D003497:Cyclic N-Oxides; D004578:Electron Spin Resonance Spectroscopy; D006863:Hydrogen-Ion Concentration; D008051:Lipid Bilayers / Q000378:metabolism*; D008081:Liposomes / Q000378:metabolism; D008274:Magnesium / Q000494:pharmacology*; D008301:Maleimides; D008563:Membrane Lipids / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D008929:Mitochondria, Heart / Q000201:enzymology*; D009840:Oligomycins / Q000494:pharmacology; D011487:Protein Conformation / Q000187:drug effects; D006180:Proton-Translocating ATPases / Q000378:metabolism*; D013113:Spin Labels; D013552:Swine
D003497:Cyclic N-Oxides; D008051:Lipid Bilayers; D008081:Liposomes; D008301:Maleimides; D008563:Membrane Lipids; D008565:Membrane Proteins; D009840:Oligomycins; D013113:Spin Labels; C028369:tempo 4-bromoacetamide; C043592:maleimide; D000255:Adenosine Triphosphate; D006180:Proton-Translocating ATPases; D008274:Magnesium