Department of Medicine, Division of Nephrology, David Geffen School of Medicine, University of California, Los Angeles, California.
Department of Neurobiology, University of California, Los Angeles, California.
Am J Physiol Cell Physiol. 2020 Feb 1;318(2):C392-C405. doi: 10.1152/ajpcell.00425.2019. Epub 2019 Nov 27.
Whether SLC4A11 transports ammonia and its potential mode of ammonia transport (, NH, or NH-2H transport have been proposed) are controversial. In the absence of ammonia, whether SLC4A11 mediates significant conductive H(OH) transport is also controversial. The present study was performed to determine the mechanism of human SLC4A11 ammonia transport and whether the transporter mediates conductive H(OH) transport in the absence of ammonia. We quantitated H flux by monitoring changes in intracellular pH (pH) and measured whole cell currents in patch-clamp studies of HEK293 cells expressing the transporter in the absence and presence of NHCl. Our results demonstrate that SLC4A11 mediated conductive H(OH) transport that was stimulated by raising the extracellular pH (pH). Ammonia-induced HEK293 whole cell currents were also stimulated by an increase in pH. In studies using increasing NHCl concentrations with equal extracellular and intracellular concentrations, the shift in the reversal potential () due to the addition of ammonia was compatible with NH-H transport competing with H(OH) rather than NH-nH ( ≥ 2) transport. The increase in equivalent H(OH) flux observed in the presence of a transcellular H gradient was also compatible with SLC4A11-mediated NH-H flux. The NH versus data fit a theoretical model suggesting that NH-H and H(OH) competitively interact with the transporter. Studies of mutant SLC4A11 constructs in the putative SLC4A11 ion coordination site showed that both H(OH) transport and ammonia-induced whole cell currents were blocked suggesting that the H(OH) and NH-H transport processes share common features involving the SLC4A11 transport mechanism.
SLC4A11 是否转运氨及其潜在的氨转运方式（提出了 NH3 或 NH-2H 转运）存在争议。在没有氨的情况下，SLC4A11 是否介导显著的导电 H（OH）转运也存在争议。本研究旨在确定人 SLC4A11 氨转运的机制，以及该转运体在没有氨的情况下是否介导导电 H（OH）转运。我们通过监测细胞内 pH 值（pH）的变化来定量 H 通量，并在表达转运体的 HEK293 细胞的膜片钳研究中测量全细胞电流，在不存在和存在 NH4Cl 的情况下。我们的结果表明，SLC4A11 介导的导电 H（OH）转运受细胞外 pH 值升高的刺激（pH）。氨诱导的 HEK293 全细胞电流也受到 pH 值升高的刺激。在使用增加的 NH4Cl 浓度的研究中，具有相等的细胞外和细胞内浓度，由于添加氨而导致的反转电位（）的变化与 NH-H 转运与 H（OH）竞争而不是 NH-nH（≥2）转运兼容。在存在跨细胞 H 梯度的情况下观察到的等效 H（OH）通量的增加也与 SLC4A11 介导的 NH-H 通量兼容。NH 与的关系数据符合一个理论模型，表明 NH-H 和 H（OH）与转运体竞争性相互作用。在假定的 SLC4A11 离子配位位点突变 SLC4A11 构建体的研究中，H（OH）转运和氨诱导的全细胞电流均被阻断，表明 H（OH）和 NH-H 转运过程共享涉及 SLC4A11 转运机制的共同特征。
