Electrogenic Cation Binding in the Electroneutral Na+/H+ Antiporter of Pyrococcus abyssi.
Electrogenic Cation Binding in the Electroneutral Na+/H+ Antiporter of Pyrococcus abyssi.
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DOI：
10.1074/jbc.m116.761080
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发表时间：
2016-12-23
期刊：
影响因子：
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通讯作者：
Fendler K
中科院分区：
文献类型：
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作者：
Călinescu O;Linder M;Wöhlert D;Yildiz Ö;Kühlbrandt W;Fendler K
阳离子质子逆向转运蛋白家族CPA1分支中的Na+/H+逆向转运蛋白驱动真核生物和原核生物中H+与Na+离子的电子中和交换，并确保pH动态平衡。虽然它们的运输周期是整体的电子中和，但特定的部分反应是电生的。在这里，我们提出了一个电生理研究的PaNhaP的Na+/H+逆向转运蛋白从深海热球重组为脂质体。当Na+加入到PaNhaP蛋白脂质体中时，记录到正的瞬变电流，表明正电荷以k>；200 S−1的速率迅速转移到蛋白脂质体中。我们将记录到的电流归因于包括Na+结合和可能的闭合的电化学反应。随后，正电荷被输送到与H+结合相关的细胞外，因此整个反应是电子中和的。我们发现PaNhaP和相关的Jannaschii甲烷球菌MjNhaP1的pH谱和Na+亲和力的差异可以归因于PaNhaP中额外的带负电荷的谷氨酸残基。这些结果在PaNhaP和其他微生物Na+/H+交换器的生理功能的背景下进行了讨论。我们认为，无论是电子中和型还是生电型Na+/H+逆向转运体，都代表着一个精心调整的自我调节系统，在任何偏离后，它都会将细胞质的pH恢复到中性。
Na+/H+ antiporters in the CPA1 branch of the cation proton antiporter family drive the electroneutral exchange of H+ against Na+ ions and ensure pH homeostasis in eukaryotic and prokaryotic organisms. Although their transport cycle is overall electroneutral, specific partial reactions are electrogenic. Here, we present an electrophysiological study of the PaNhaP Na+/H+ antiporter from Pyrococcus abyssi reconstituted into liposomes. Positive transient currents were recorded upon addition of Na+ to PaNhaP proteoliposomes, indicating a reaction where positive charge is rapidly displaced into the proteoliposomes with a rate constant of k >200 s−1. We attribute the recorded currents to an electrogenic reaction that includes Na+ binding and possibly occlusion. Subsequently, positive charge is transported out of the cell associated with H+ binding, so that the overall reaction is electroneutral. We show that the differences in pH profile and Na+ affinity of PaNhaP and the related MjNhaP1 from Methanocaldococcus jannaschii can be attributed to an additional negatively charged glutamate residue in PaNhaP. The results are discussed in the context of the physiological function of PaNhaP and other microbial Na+/H+ exchangers. We propose that both, electroneutral and electrogenic Na+/H+ antiporters, represent a carefully tuned self-regulatory system, which drives the cytoplasmic pH back to neutral after any deviation.