Department of Cell Biology, Emory University School of Medicine, Atlanta, GA 30322, USA.
Int J Mol Sci. 2022 Jan 4;23(1):532. doi: 10.3390/ijms23010532.
The Na/HCO cotransporter NBCe1 is a member of SLC4A transporters that move HCO across cell membranes and regulate intracellular pH or transepithelial HCO transport. NBCe1 is highly selective to HCO and does not transport other anions; the molecular mechanism of anion selectivity is presently unclear. We previously reported that replacing Asp with a Glu (D555E) in NBCe1 induces increased selectivity to other anions, including Cl. This finding is unexpected because all SLC4A transporters contain either Asp or Glu at the corresponding position and maintain a high selectivity to HCO. In this study, we tested whether the Cl transport in D555E is mediated by an interaction between residues in the ion binding site. Human NBCe1 and mutant transporters were expressed in oocytes, and their ability to transport Cl was assessed by two-electrode voltage clamp. The results show that the Cl transport is induced by a charge interaction between Glu and Lys. The bond length between the two residues is within the distance for a salt bridge, and the ionic strength experiments confirm an interaction. This finding indicates that the HCO selectivity in NBCe1 is established by avoiding a specific charge interaction in the ion binding site, rather than maintaining such an interaction.
钠/碳酸氢盐协同转运蛋白 NBCe1 是 SLC4A 转运蛋白家族的成员，能跨膜转运 HCO3-并调节细胞内 pH 值或跨上皮 HCO3-转运。NBCe1 对 HCO3-具有高度选择性，不转运其他阴离子；阴离子选择性的分子机制目前尚不清楚。我们之前的研究报道，用 Glu（D555E）取代 NBCe1 中的 Asp 会诱导对其他阴离子（包括 Cl-）的选择性增加。这一发现出乎意料，因为所有 SLC4A 转运蛋白在相应位置都含有 Asp 或 Glu，对 HCO3-保持高选择性。在这项研究中，我们测试了 D555E 中的 Cl-转运是否由离子结合位点中残基之间的相互作用介导。将人 NBCe1 和突变体转运蛋白在卵母细胞中表达，并通过双电极电压钳评估它们转运 Cl-的能力。结果表明，Cl-的转运是由 Glu 和 Lys 之间的电荷相互作用诱导的。两个残基之间的键长在盐桥的距离范围内，离子强度实验证实了这种相互作用。这一发现表明，NBCe1 中的 HCO3-选择性是通过避免离子结合位点中特定的电荷相互作用而建立的，而不是维持这种相互作用。
