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首页> 外文期刊>American Journal of Physiology >Functional characterization of human NBC4 as an electrogenic Na+-HCO cotransporter (NBCe2).
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Functional characterization of human NBC4 as an electrogenic Na+-HCO cotransporter (NBCe2).

机译：人类NBC4作为电性Na + -HCO共转运蛋白（NBCe2）的功能表征。

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We have functionally characterized Na+-driven bicarbonate transporter (NBC)4, originally cloned from human heart by Pushkin et al. (Pushkin A, Abuladze N, Newman D, Lee I, Xu G, and Kurtz I. Biochem Biophys Acta 1493: 215-218, 2000). Of the four NBC4 variants currently present in GenBank, our own cloning efforts yielded only variant c. We expressed NBC4c (GenBank accession no. AF293337) in Xenopus laevis oocytes and assayed membrane potential (Vm) and pH regulatory function with microelectrodes. Exposing an NBC4c-expressing oocyte to a solution containing 5% CO2 and 33 mM HCO elicited a large hyperpolarization, indicating that the transporter is electrogenic. The initial CO2-induced decrease in intracellular pH (pH(i)) was followed by a slow recovery that was reversed by removing external Na+. Two-electrode voltage clamp of NBC4c-expressing oocytes revealed large HCO- and Na+-dependent currents. When we voltage clamped V(m) far from NBC4c's estimated reversal potential (E(rev)), the pH(i) recovery rateincreased substantially. Both the currents and pH(i) recovery were blocked by 200 microM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS). We estimated the transporter's HCO:Na+ stoichiometry by measuring E(rev) at different extracellular Na+ concentration ([Na+]o) values. A plot of E(rev) against log[Na+]o was linear, with a slope of 54.8 mV/log[Na+]o. This observation, as well as the absolute E(rev) values, are consistent with a 2:1 stoichiometry. In conclusion, the behavior of NBC4c, which we propose to call NBCe2-c, is similar to that of NBCe1, the first electrogenic NBC.
机译：我们具有功能上由Na +驱动的碳酸氢根转运蛋白（NBC）4的功能特征，最初是由普希金等人从人的心脏中克隆的。 （Pushkin A，Abuladze N，Newman D，Lee I，Xu G和Kurtz I.Biochem Biophys Acta 1493：215-218，2000）。目前在GenBank中存在的四个NBC4变体中，我们自己的克隆工作仅产生了变体c。我们在非洲爪蟾卵母细胞中表达了NBC4c（GenBank登录号AF293337），并通过微电极测定了膜电位（Vm）和pH调节功能。将表达NBC4c的卵母细胞暴露于含有5％CO2和33 mM HCO的溶液中会引起大的超极化现象，表明转运蛋白是电动的。最初由CO2引起的细胞内pH（pH（i））下降是缓慢的恢复，随后通过去除外部Na +逆转。表达NBC4c的卵母细胞的两电极电压钳显示出大量的HCO和Na +依赖性电流。当我们用电压钳位V（m）使其远离NBC4c的估计反转电位（E（rev））时，pH（i）的回收率将大大提高。 200 microM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid（DIDS）阻止了电流和pH（i）的恢复。我们通过在不同的细胞外Na +浓度（[Na +] o）值下测量E（rev）来估算转运蛋白的HCO：Na +化学计量。 E（rev）对log [Na +] o的图是线性的，斜率为54.8 mV / log [Na +] o。该观察结果以及绝对E（rev）值与化学计量比为2：1一致。总之，我们建议将NBC4c的行为称为NBCe2-c，它与第一个电动NBC的NBCe1相似。

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