Department of Anesthesiology, Weill Cornell Medical College, New York, New York, USA.
Nat Struct Mol Biol. 2012 Apr 8;19(5):525-31, S1. doi: 10.1038/nsmb.2277.
Active exchangers dissipate the gradient of one substrate to accumulate nutrients, export xenobiotics and maintain cellular homeostasis. Mechanistic studies have suggested that two fundamental properties are shared by all exchangers: substrate binding is antagonistic, and coupling is maintained by preventing shuttling of the empty transporter. The CLC H(+)/Cl(-) exchangers control the homeostasis of cellular compartments in most living organisms, but their transport mechanism remains unclear. We show that substrate binding to CLC-ec1 is synergistic rather than antagonistic: chloride binding induces protonation of a crucial glutamate. The simultaneous binding of H(+) and Cl(-) gives rise to a fully loaded state that is incompatible with conventional transport mechanisms. Mutations in the Cl(-) transport pathway identically alter the stoichiometries of H(+)/Cl(-) exchange and binding. We propose that the thermodynamics of synergistic substrate binding, rather than the kinetics of conformational changes and ion binding, determine the stoichiometry of transport.
活性交换器消耗一种基质的梯度来积累营养物质，排出异源生物，并维持细胞内环境稳定。机制研究表明，所有交换器都具有两个基本特性：基质结合具有拮抗性，并且通过防止空载转运体穿梭来维持偶联。CLC H(+)/Cl(-)交换器控制大多数生物体细胞区室的内环境稳定，但它们的运输机制仍不清楚。我们表明，CLC-ec1 的基质结合是协同的而不是拮抗的：氯离子结合诱导关键谷氨酸的质子化。H(+)和 Cl(-)的同时结合产生了与传统运输机制不兼容的完全加载状态。Cl(-)运输途径中的突变以相同的方式改变 H(+)/Cl(-)交换和结合的计量关系。我们提出，协同基质结合的热力学而不是构象变化和离子结合的动力学决定了运输的计量关系。
