Passive water and ion transport by cotransporters
Passive water and ion transport by cotransporters
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DOI：
10.1111/j.1469-7793.1999.0195r.x
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发表时间：
1999-07-01
影响因子：
5.5
通讯作者：
Wright, EM
中科院分区：
文献类型：
--
作者：
Loo, DDF;Hirayama, BA;Wright, EM
1.在非洲爪蟾卵母细胞中表达了兔Na+-葡萄糖（SGLT 1）和人Na+-Cl--GABA（GAT 1）共转运蛋白，并利用电学和光学技术研究了Na+和水的被动转运。被动水渗透性（L-p）的协同转运蛋白测定卵母细胞体积的变化，响应渗透梯度。通过在存在和不存在阻断剂（根皮苷和SKF 89976 A）的情况下测量L-p，获得特异性SGLT 1和GAT 1 L-p值。在存在阻断剂的情况下，表达SGLT 1和GAT 1的卵母细胞的L-p值与对照卵母细胞的L-p值无法区分。被动Na+转运（Na+泄漏），从阻断剂敏感的Naf电流的情况下的底物（葡萄糖和GABA）。根皮苷以相同的敏感性（抑制常数（Ki），3-5 μ M）阻断了通过SGLT 1的被动Na+和水转运。当Na+被Li+取代时，根皮苷也抑制Li+和水的运输，但具有较低的亲和力（Ki，100 μ M）。当Na+被胆碱替代时，SGLT 1 L-p与Na+或Li+中的L-p没有区别，但在这种情况下，水转运对根皮苷的敏感性较低. Na+和水通过SGLT 1被动转运的活化能（E-a）分别为21和5 kcal mol（-1）。Na+转运的高E-a与Na+-葡萄糖共转运的E-a相当，表明该过程依赖于蛋白质的构象变化，而水转运的低E-a与水通道（水通道蛋白）的E-a相似. GAT 1也表现为SKF 89976 A敏感的水通道。我们没有观察到通过GAT1.5的被动Na+转运。我们得出结论，被动水和Na+运输通过cotransporters取决于不同的机制：Na+运输发生饱和的单端口机制，和水渗透是通过一个低电导的水通道。在SGLT 1的情况下，我们认为水通道和水的共转运可能有助于等渗液体转运通过肠刷状缘膜。
1. The rabbit Na+-glucose (SGLT1) and the human Na+-Cl--GABA (GAT1) cotransporters were expressed in Xenopus laevis oocytes, and passive Na+ and water transport were studied using electrical and optical techniques. Passive water permeabilities (L-p) of the cotransporters were deter mined from the changes in oocyte volume in response to osmotic gradients. The specific SGLT1 and GAT1 L-p values were obtained by measuring L-p in the presence and absence of blockers (phlorizin and SKF89976A). In the presence of the blockers, the L-p values of oocytes expressing SGLT1 and GAT1 were indistinguishable from the L-p of control oocytes,2. Passive Na+ transport (Na+ leak) was obtained from the blocker-sensitive Naf currents in the absence of substrates (glucose and GABA). Passive Na+ and water transport through SGLT1 were blocked by phlorizin with the same sensitivity (inhibitory constant (K-i), 3-5 mu M). When Na+ was replaced with Li+, phlorizin also inhibited Li+ and water transport, but with a lower affinity (K-i, 100 mu M). When Na+ was replaced by choline, which is not transported, the SGLT1 L-p was indistinguishable from that in Na+ or Li+, but in this case water transport was less sensitive to phlorizin.3. The activation energies (E-a) for passive Na+ and water transport through SGLT1 were 21 and 5 kcal mol(-1), respectively. The high E-a for Na+ transport is comparable to that of Na+-glucose cotransport and indicates that the process is dependent on conformational changes of the protein, while the low E-a for water transport is similar to that of water channels (aquaporins).4. GAT1 also behaved as an SKF89976A-sensitive water channel. We did not observe passive Na+ transport through GAT1.5. We conclude that passive water and Na+ transport through cotransporters depend on different mechanisms: Na+ transport occurs by a saturable uniport mechanism, and water permeation is through a low conductance water channel. In the case of SGLT1, we suggest that both the water channel and water cotransport could contribute to isotonic fluid transport across the intestinal brush border membrane.