pH-Induced conformational change of ATPase inhibitor from yeast mitochondria. A proton magnetic resonance study.
The ATPase inhibitor from yeast, Saccharomyces cerevisiae, was found to strongly inhibit the ATPase activity when it was preincubated in an acidic pH region. This paper describes the conformational changes of the inhibitor in the acidic pH region, as studied by 1H NMR spectroscopy. In the pH range from 7.43 to 4.48, two conformations were detected. With lowering of the pH in this pH region, one conformation increased at the expense of the other. In the two conformations, His 39 had different pKa values which were determined at 23 degrees C to be 6.08 +/- 0.04 and 5.91 +/- 0.03 from the pH-titration curves of His 39. The exchange between the two conformations was reversible with pH and the rate of the conformational change was estimated to be slower than 22 s-1 at 23 degrees C as estimated by the line-widths (7.0 Hz) of the peaks from the C2-proton of His 39 in the two conformations. The equilibrium constant between the two conformations was dependent on the temperature. From the equilibrium constants in the temperature range from 23 to 45 degrees C, the apparent delta H'a of the conformational change was calculated to be 14 kcal/mol at pH 6.95. It was confirmed by 1H NMR spectra that at pH 6.95 the structure of the inhibitor was reversible with temperature at least below 80 degrees C. The reversibility is consistent with the high thermal stability of the protein.
D006863:Hydrogen-Ion Concentration; D009682:Magnetic Resonance Spectroscopy; D008928:Mitochondria / Q000032:analysis; D011487:Protein Conformation; D011506:Proteins / Q000302:isolation & purification; D011522:Protons; D012441:Saccharomyces cerevisiae / Q000032:analysis*; D013696:Temperature; D000097795:ATPase Inhibitory Protein
D011506:Proteins; D011522:Protons; D000097795:ATPase Inhibitory Protein