Hendler R W, Drachev L A, Bose S, Joshi M K
Laboratory of Cell Biology, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Eur J Biochem. 2000 Oct;267(19):5879-90. doi: 10.1046/j.1432-1327.2000.01620.x.
The kinetics of the bacteriorhodopsin photocycle, measured by voltage changes in a closed membrane system using the direct electrometrical method (DEM) of Drachev, L.A., Jasaitus, A.A., Kaulen, A.D., Kondrashin, A.A., Liberman, E.A., Nemecek, I.B., Ostroumov, S.A., Semenov, Yu, A. & Skulachev, V.P. (1974) Nature 249, 321-324 are sixfold slower than the kinetics obtained in optical studies with suspensions of purple membrane patches. In this study, we have investigated the reasons for this discrepancy. In the presence of the uncouplers carbonyl cyanide m-chlorophenylhydrazone or valinomycin, the rates in the DEM system are similar to the rates in suspensions of purple membrane. Two alternative explanations for the effects of uncouplers were evaluated: (a) the 'back-pressure' of the Deltamicro;H+ slows the kinetic steps leading to its formation, and (b) the apparent difference between the two systems is due to slow major electrogenic events that produce little or no change in optical absorbance. In the latter case, the uncouplers would decrease the RC time constant for membrane capacitance leading to a quicker discharge of voltage and concomitant decrease in photocycle turnover time. The experimental results show that the primary cause for the slower kinetics of voltage changes in the DEM system is thermodynamic back-pressure as described by Westerhoff, H.V. & Dancshazy, Z. (1984) Trends Biochem. Sci. 9, 112-117.
用德拉切夫、亚赛图斯、考伦、孔德拉申、利伯曼、涅梅切克、奥斯特鲁莫夫、谢苗诺夫、尤·A.和斯库拉乔夫（1974年，《自然》249卷，321 - 324页）的直接电测法（DEM），通过封闭膜系统中的电压变化来测量细菌视紫红质光循环的动力学，其速度比用紫色膜片悬浮液进行光学研究得到的动力学速度慢六倍。在本研究中，我们探究了这种差异的原因。在存在解偶联剂羰基氰化物间氯苯腙或缬氨霉素的情况下，DEM系统中的速率与紫色膜悬浮液中的速率相似。对解偶联剂的作用评估了两种替代解释：（a）ΔμH⁺的“背压”减缓了导致其形成的动力学步骤；（b）两个系统之间的明显差异是由于缓慢的主要电致事件，这些事件在光吸收上几乎没有变化或没有变化。在后一种情况下，解偶联剂会降低膜电容的RC时间常数，导致电压更快放电，并伴随着光循环周转时间的减少。实验结果表明，DEM系统中电压变化动力学较慢的主要原因是韦斯特霍夫和丹奇哈齐（1984年，《生物化学趋势》9卷，112 - 117页）所描述的热力学背压。
