Unisite hydrolysis of [gamma 32 P]ATP by soluble mitochondrial F1-ATPase and its release by excess ADP and ATP. Effect of trifluoperazine.
Some of the characteristics of unisite hydrolysis of [gamma 32P]ATP as well as the changes that occur on the transition to multisite catalysis were further studied. It was found that a fraction of [gamma 32P]ATP bound at the catalytic sites of F1 under unisite conditions undergoes both hydrolysis and release induced by medium nucleotides upon addition of millimolar concentrations of ADP or ATP. The fraction of [gamma 32P]ATP that undergoes release is similar to the fraction that undergoes hydrolytic cleavage, indicating that the rates of the release and hydrolytic reactions of bound [gamma 32P]ATP are in the same range. As part of studies on the mechanisms through which trifluoperazine inhibits ATP hydrolysis, its effect on unisite hydrolysis of [gamma 32P]ATP was also studied. Trifluoperazine diminishes the rate of unisite hydrolysis by 30-40%. The inhibition is accompanied by a nearly tenfold increase in the ratio of [gamma 32P]ATP/32Pi bound at the catalytic site and a 50% diminution in the rate of 32Pi release from the enzyme into the media. Trifluoperazine also induces heterogeneity of the three catalytic sites of F1 in the sense that in a fraction of F1 molecules, the high-affinity catalytic site has a turnover rate lower than the other two. Trifluoperazine does not modify the release of previously bound [gamma 32P]ATP induced by medium nucleotides. The latter indicates that hindrances in the release of Pi do not necesarily accompany alterations in the release of ATP even though both species lie in the same site.
García
J J
JJ
Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, México, D. F. México.
Gómez-Puyou
A
A
de Gómez-Puyou
M T
MT
D000244:Adenosine Diphosphate / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D002384:Catalysis; D006868:Hydrolysis; D008928:Mitochondria / Q000201:enzymology*; D010710:Phosphates; D010761:Phosphorus Radioisotopes; D006180:Proton-Translocating ATPases / Q000378:metabolism*; D012995:Solubility; D014268:Trifluoperazine / Q000494:pharmacology*
D010710:Phosphates; D010761:Phosphorus Radioisotopes; D014268:Trifluoperazine; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D006180:Proton-Translocating ATPases