Chloride transport across vesicle and cell membranes by steroid-based receptors
Chloride transport across vesicle and cell membranes by steroid-based receptors
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DOI：
10.1002/anie.200351957
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发表时间：
2003-01-01
影响因子：
16.6
通讯作者：
Davis, AP
中科院分区：
文献类型：
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作者：
Koulov, AV;Lambert, TN;Davis, AP
众所周知，跨细胞膜转运阳离子的分子（阳离子载体）可能具有有效的生物学效应。[1]阴离子通量对细胞也很重要，相应地，阴离子载体可能具有生物活性。事实上，氯离子转运蛋白具有治疗囊性纤维化和其他由缺陷通道蛋白引起的疾病的直接医学潜力。[2]尽管有这样的动机，有相对较少的报道阴离子载体天然产品[3]或阴离子运输的合成系统。[4]此外，大多数都涉及阳离子中心，[5]这可以帮助阴离子通过形成离子对。[4a-g]纯电中性系统的阴离子传输仍然非常罕见。最近描述的“Cholapod”阴离子受体1本质上是亲脂性的，可能是膜溶性的，并且可调节至非常高的亲和力。[6]某些胆足类已被证明具有“翻转酶”活性，即它们可以将极性头部基团传送穿过磷脂膜。[7]因此，我们推测，电中性的胆足可能通过“穿梭”机制运输无机阴离子，如阳离子载体缬氨霉素的方式。我们现在报道，胆足2确实能够跨脂质体膜和跨作为极化上皮生长的活细胞转运氯离子。受体2由胆酸3通过转化为二胺4 [8]并用适当的异氰酸酯处理制备。最初，他们的阴离子转运性能进行了研究，以下氯离子流出单层囊泡使用氯离子选择性电极。更具体地，通过挤出1-棕榈酰基-2-油酰基-sn-甘油基-3-磷酸胆碱（POPC）和胆固醇[9]在NaCl水溶液（500 mm）中的7：3混合物来制备30 mm单层囊泡（200 nm平均直径）。用NaNO 3水溶液（500 mM）透析该储液囊泡分散体，以用硝酸盐代替外部氯离子，然后用500 mM NaNO 3稀释，得到1 mM总脂质浓度。如图1所示，加入2a-e的THF溶液（最终胆足浓度为4.0 μm）导致氯离子从囊泡内相流出。本文测定了不同浓度的Et_4N +Cl ~-和Et_4N + NO_3 ~-的Cl ~-流出初始速率和缔合常数2a-e，
It is well-established that molecules which transport cations across cell membranes (cationophores) can have potent biological effects.[1] Anion flux is also important to the cell and, correspondingly, anion carriers may be capable of biological activity. Indeed, chloride transporters have direct medical potential as treatments for cystic fibrosis and other diseases caused by defective channel proteins.[2] Despite this motivation, there have been relatively few reports of anionophore natural products [3] or of anion transport by synthetic systems.[4] Most, moreover, have involved cationic centers,[5] which can assist anion passage through the formation of an ion pair.[4a–g] Anion transport by purely electroneutral systems is still quite rare.[4j–l] The recently described “cholapod” anion receptors 1 are intrinsically lipophilic, potentially membranesoluble, and tuneable to very high affinities.[6] Certain cholapods have proved capable of “flippase” activity, that is, they can convey polar head groups across phospholipid membranes.[7] We therefore supposed that electroneutral cholapods might transport inorganic anions by a “shuttle” mechanism, in the manner of cationophores such as valinomycin. We now report that cholapods 2 are indeed capable of transporting chloride ions across liposomal membranes and also across live cells grown as polarized epithelia.Receptors 2 were prepared from cholic acid 3 by conversion into diamine 4 [8] and treatment with appropriate isocyanates. Initially, their anion-transport properties were studied by following chloride efflux from unilamellar vesicles using a chloride-selective electrode. More specifically, 30 mm unilamellar vesicles (200 nm mean diameter) were prepared by extruding a 7: 3 mixture of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and cholesterol [9] in aqueous NaCl (500 mm). This stock vesicle dispersion was dialyzed against aqueous NaNO3 (500 mm) to replace the external chloride ions with nitrate and then diluted with 500 mm NaNO3 to give a 1mm total lipid concentration. As shown in Figure1, addition of a solution of 2a–e in THF (4.0 μm final cholapod concentration) caused chloride efflux from the inner phase of the vesicles. The initial rates of chloride efflux and the association constants of 2a–e for Et4N+Cl¿ and Et4N+NO3¿ in