Labahn A, Fromme P, Gräber P
Max Volmer Institut für Biophysikalische und Physikalische Chemie, Technische Universität Berlin, FRG.
FEBS Lett. 1990 Oct 1;271(1-2):116-8. doi: 10.1016/0014-5793(90)80385-v.
Uni-site ATP synthesis was measured with thylakoids. The membrane-bound ATP-synthase, CF0F1, was brought into the active, reduced state by illumination in the presence of thioredoxin, dithiothreitol and phosphate. This enzyme contains two tightly bound ATP per CF0F1. ATP was released from the enzyme when ADP was added in substoichiometric amounts during illumination. Experiments with [14C]ADP indicated that after binding the same nucleotide was phosphorylated and released as [14C]ATP, i.e. only one site is involved in ATP-synthesis ('uni-site ATP-synthesis'). The two tightly bound ATP are not involved in the catalytic turnover. The rate constant for ADP binding was (4 +/- 2) x 10(6) M-1s-1. Compared to deenergized conditions the rate constant for ADP binding and that for ATP-release were drastically increased, i.e. membrane energization increased the rate constants for the ATP-synthesis direction.
利用类囊体测量单位点ATP合成。在存在硫氧还蛋白、二硫苏糖醇和磷酸盐的情况下，通过光照使膜结合的ATP合酶CF0F1进入活性还原状态。该酶每个CF0F1含有两个紧密结合的ATP。在光照期间以亚化学计量加入ADP时，ATP从酶中释放出来。用[14C]ADP进行的实验表明，结合后相同的核苷酸被磷酸化并作为[14C]ATP释放，即只有一个位点参与ATP合成（“单位点ATP合成”）。两个紧密结合的ATP不参与催化周转。ADP结合的速率常数为(4±2)×10(6) M-1s-1。与去能条件相比，ADP结合的速率常数和ATP释放的速率常数急剧增加，即膜 energization增加了ATP合成方向的速率常数。
