Bakker E P, Randall L L
EMBO J. 1984 Apr;3(4):895-900. doi: 10.1002/j.1460-2075.1984.tb01902.x.
The energy requirement for the maturation and export of the plasmid-encoded TEM beta-lactamase in Escherichia coli K12 was shown to be fulfilled by the total protonmotive force. This was demonstrated by assessing the inhibition of proteolytic processing of the precursor form of beta-lactamase caused by perturbation of the energized state of the membrane in cells treated with valinomycin. The magnitude of the membrane potential was manipulated by varying the concentration of KCl in the medium and the pH gradient was manipulated by varying the external pH. Both components were simultaneously affected by addition of the protonophore carbonylcyanide-p- trifluoromethoxy phenylhydrazone (FCCP). Inhibition of processing was demonstrated in a mutant strain having a defective ATP synthase where protonmotive force could be dissipated without altering the intracellular level of ATP, indicating that the observed inhibition was not the result of decreased ATP concentration. Half-maximal accumulation of precursor of beta-lactamase was observed in all cases when the level of protonmotive force was decreased to approximately 150 mV. Under those conditions the membrane potential varied from 65 to 140 mV (internally negative) and the pH gradient from 95 to 25 mV (internally alkaline). Thus, the energy requirement is satisfied by the total protonmotive force, with no specificity for either the membrane potential or the pH gradient.
已证明，大肠杆菌K12中质粒编码的TEMβ-内酰胺酶成熟和输出所需的能量由总质子动力提供。在用缬氨霉素处理的细胞中，通过评估膜的通电状态受到干扰时对β-内酰胺酶前体形式的蛋白水解加工的抑制作用，证明了这一点。通过改变培养基中KCl的浓度来操纵膜电位的大小，通过改变外部pH来操纵pH梯度。通过添加质子载体羰基氰化物-p-三氟甲氧基苯腙（FCCP），可同时影响这两个组分。在具有缺陷ATP合酶的突变菌株中证明了加工过程受到抑制，在该菌株中，质子动力可以消散而不改变细胞内ATP水平，这表明观察到的抑制不是ATP浓度降低的结果。当质子动力水平降至约150 mV时，在所有情况下均观察到β-内酰胺酶前体的半最大积累。在这些条件下，膜电位在65至140 mV之间变化（内部为负），pH梯度在95至25 mV之间变化（内部为碱性）。因此，总质子动力满足能量需求，对膜电位或pH梯度均无特异性。
