Effects of sulfate and chloride on three separate oxalate transporters reconstituted from rabbit renal cortex.
Effects of sulfate and chloride on three separate oxalate transporters reconstituted from rabbit renal cortex.
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硫酸盐和氯化物对从兔肾皮质重建的三个单独的草酸盐转运蛋白的影响。
DOI：
10.1152/ajprenal.1998.274.1.f189
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发表时间：
1998
期刊：
影响因子：
--
通讯作者：
Karniski,LP
中科院分区：
文献类型：
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作者：
Karniski,LP
了解哺乳动物近端小管中硫酸盐依赖性、草酸盐刺激的氯离子重吸收机制是复杂的，因为存在多种草酸盐和硫酸盐转运途径。因此，我们开发了一种方法，重建功能性草酸盐转运从兔肾皮质，使个别转运可能会被检查。溶解的微绒毛膜蛋白通过羟基磷灰石层析分离，然后重组成脂蛋白体。观察到两个草酸盐/草酸盐交换活性峰。硫酸盐（10 mM）顺式抑制草酸盐运输的早期峰值为93%，后期峰值为41%。相比之下，20 mM氯化物在早期峰值中仅抑制32%的草酸盐/草酸盐交换，但在后期峰值中抑制70%的草酸盐交换。草酸盐刺激的硫酸盐吸收中观察到的早期馏分，但不是在后来的馏分。这些数据与早期羟基磷灰石级分中硫酸盐/草酸盐交换剂和后期级分中氯化物/草酸盐交换剂的回收率一致。基底外侧膜硫酸盐/草酸盐交换器也被重建。重建的基底外侧和顶端膜硫酸盐/草酸盐交换器表现出几乎相同的模式的底物特异性。然而，在室温下暴露于辛基葡萄糖苷后，顶端硫酸盐/草酸盐交换活性丧失了98%，而基底外侧硫酸盐/草酸盐交换活性降低了67%（P< 0.05）。总之，溶解的膜蛋白的功能重建表明，顶端膜氯/草酸盐交换和硫酸盐/草酸盐交换介导的不同的转运蛋白。顶端和基底侧硫酸盐/草酸盐交换也可能代表两个单独的交换器上的转运。
Understanding the mechanism of sulfate-dependent, oxalate-stimulated chloride reabsorption in the mammalian proximal tubule is complicated by the presence of multiple oxalate and sulfate transport pathways. Accordingly, we developed a method of reconstituting functional oxalate transport from the rabbit renal cortex so that the individual transporters might be examined. Solubilized microvillus membrane proteins were separated by hydroxyapatite chromatography and then reconstituted into proteoliposomes. Two peaks of oxalate/oxalate exchange activity were observed. Sulfate (10 mM)cis-inhibits oxalate transport in the early peak by 93% and in the later peak by 41%. In contrast, 20 mM chloride inhibits oxalate/oxalate exchange by only 32% in the early peak but inhibits oxalate exchange by 70% in the later peak. Oxalate-stimulated sulfate uptake was observed in the early fractions but not in the later fractions. These data are consistent with the recovery of the sulfate/oxalate exchanger in the early hydroxyapatite fractions and the chloride/oxalate exchanger in the later fractions. The basolateral membrane sulfate/oxalate exchanger was also reconstituted. The reconstituted basolateral and apical membrane sulfate/oxalate exchangers demonstrate nearly identical patterns of substrate specificities. However, 98% of apical sulfate/oxalate exchange activity is lost following exposure to octylglucoside at room temperature, whereas the basolateral sulfate/oxalate exchange activity was reduced 67% (P< 0.05). In conclusion, functional reconstitution of solubilized membrane proteins demonstrates that apical membrane chloride/oxalate exchange and sulfate/oxalate exchange are mediated by different transport proteins. Apical and basolateral sulfate/oxalate exchange may also represent transport on two separate exchangers.