Mobley H L, Rosen B P
Proc Natl Acad Sci U S A. 1982 Oct;79(20):6119-22. doi: 10.1073/pnas.79.20.6119.
Plasmid R773, which codes for resistances to arsenate, arsenite, and antimony, was introduced into Escherichia coli strain AN120, a mutant deficient in the H+-translocating ATPase of oxidative phosphorylation. Cultures depleted of endogenous energy reserves were loaded with 74AsO3-4, and arsenate efflux was measured after dilution into medium containing various energy sources and inhibitors. Rapid extrusion of arsenate occurred when glucose was added. Arsenate was extruded both against and down a concentration gradient. In this strain glucose allows formation of both ATP via substrate-level phosphorylation and an electrochemical proton gradient (or protonmotive force) via oxidation of the products of glycolysis. When oxidation was inhibited by cyanide, glucose metabolism still produced arsenate efflux. Energy sources such as succinate, which supplies a protonmotive force but not ATP, did not result in efflux. Measurement of intracellular ATP concentration under each set of conditions demonstrated a direct correlation between the rate of efflux and ATP levels. Osmotically shocked cells lost the ability to extrude arsenate; however, no arsenate-binding activity was detected in osmotic shock fluid from induced cells. These results suggest that the arsenate efflux system is coupled to cellular ATP rather than an electrochemical proton gradient, possibly by an arsenate-translocating ATPase.
编码对砷酸盐、亚砷酸盐和锑抗性的质粒R773被导入大肠杆菌菌株AN120，该菌株是氧化磷酸化过程中H⁺转运ATP酶缺陷的突变体。耗尽内源性能量储备的培养物用⁷⁴AsO₃⁻⁴加载，在稀释到含有各种能量来源和抑制剂的培养基后测量砷酸盐流出。添加葡萄糖时会迅速挤出砷酸盐。砷酸盐既可以逆浓度梯度也可以顺浓度梯度挤出。在该菌株中，葡萄糖既可以通过底物水平磷酸化形成ATP，也可以通过糖酵解产物的氧化形成电化学质子梯度（或质子动力）。当氧化被氰化物抑制时，葡萄糖代谢仍会产生砷酸盐流出。诸如琥珀酸盐等提供质子动力但不提供ATP的能量来源不会导致流出。在每组条件下测量细胞内ATP浓度表明，流出速率与ATP水平之间存在直接相关性。经渗透压休克处理的细胞失去了挤出砷酸盐的能力；然而，在诱导细胞的渗透压休克液中未检测到砷酸盐结合活性。这些结果表明，砷酸盐流出系统与细胞ATP偶联，而不是与电化学质子梯度偶联，可能是通过砷酸盐转运ATP酶实现的。
