Chernyak B V, Dukhovich V F
A. N. Belozersky Laboratory of Molecular Biology and Bioorganic Chemistry, Moscow State University, USSR.
Arch Biochem Biophys. 1991 May 1;286(2):604-9. doi: 10.1016/0003-9861(91)90087-y.
A decrease in the rate of ATP hydrolysis was observed after preincubation of intact mitochondria from hepatoma 22a with an uncoupler. This effect is due both to a decrease in the rate of ATP transport and to an inactivation of the F0F1-ATPase. The former effect is shown to result from an uncoupler-induced ADP efflux. In de-energized mitochondria from hepatoma (but not from mice liver), the concentration of adenine nucleotides in the matrix equilibrates with the medium concentration via a carboxyatractyloside (CATR)-insensitive transport system. CATR-insensitive accumulation of medium ADP and stoichiometric exchange of added ATP are observed in energized hepatoma mitochondria. The dependence of the uncoupler-induced inactivation of ATPase activity on delta mu H+, pH, and ATP is consistent with the effect being caused by the natural protein inhibitor (IF1) of F0F1. ATP- and pH-dependent inactivation of the enzyme is also observed after disruption of mitochondria with the detergent Lubrol-WX. Almost all F0F1 in hepatoma mitochondria have IF1 bound in a noninhibitory manner. In the presence of uncoupler, this complex converts, via a reversible pH-dependent and an irreversible ATP-dependent process, to an inhibitory complex. The pH-dependent step can be blocked by Zn2+ and Cd2+ ions which probably bind to negatively charged residues on IF1, thereby preventing their protonation and conversion of the protein to an inhibitory conformation.
用解偶联剂预孵育肝癌22a的完整线粒体后，观察到ATP水解速率降低。这种效应既归因于ATP转运速率的降低，也归因于F0F1 - ATP酶的失活。前一种效应被证明是由解偶联剂诱导的ADP外流所致。在肝癌（而非小鼠肝脏）的去能线粒体中，基质中的腺嘌呤核苷酸浓度通过一种对羧基苍术苷（CATR）不敏感的转运系统与培养基浓度达到平衡。在有能量的肝癌线粒体中观察到培养基ADP的CATR不敏感积累和添加ATP的化学计量交换。解偶联剂诱导的ATP酶活性失活对δμH⁺、pH和ATP的依赖性与该效应是由F0F1的天然蛋白质抑制剂（IF1）引起的一致。在用去污剂Lubrol - WX破坏线粒体后，也观察到酶的ATP和pH依赖性失活。肝癌线粒体中几乎所有的F0F1都以非抑制方式结合有IF1。在存在解偶联剂的情况下，这种复合物通过一个可逆的pH依赖性和一个不可逆的ATP依赖性过程转化为抑制性复合物。pH依赖性步骤可被Zn²⁺和Cd²⁺离子阻断，这些离子可能与IF1上带负电荷的残基结合，从而阻止它们的质子化以及蛋白质转化为抑制性构象。
