组氨酸特异性试剂对肾微绒毛膜钠氢交换体的失活作用。

Inactivation of the renal microvillus membrane Na+-H+ exchanger by histidine-specific reagents.

作者信息

Grillo F G, Aronson P S

出版信息

J Biol Chem. 1986 Jan 25;261(3):1120-5.

Abstract

We examined the effect of histidine-specific reagents on the transport activity of the Na+-H+ exchanger in microvillus (brush-border) membrane vesicles isolated from the rabbit renal cortex. Rose bengal-catalyzed photo-oxidation caused irreversible inhibition of the rate of Na+-H+ exchange but also caused significant loss of vesicle integrity. Treatment of the membrane vesicles with diethylpyrocarbonate caused inactivation of Na+-H+ exchange that could not be attributed to vesicle disruption or collapse of transmembrane H+ gradients. Inactivation of Na+-H+ exchange by diethylpyrocarbonate followed pseudo-first order kinetics to below 10% residual activity, could be reversed by hydroxylamine, was reflected by a decreased Vmax with no change in the Km for Na+, was dependent on external pH but not internal pH, was blocked by amiloride, and was enhanced by Na+. These data are consistent with the hypothesis that a diethylpyrocarbonate-sensitive imidazolium residue is the titratable group found in kinetic studies to bind H+ at the external transport site of the Na+-H+ exchanger.

摘要

我们研究了组氨酸特异性试剂对从兔肾皮质分离的微绒毛（刷状缘）膜囊泡中Na⁺-H⁺交换体转运活性的影响。孟加拉玫瑰红催化的光氧化导致Na⁺-H⁺交换速率的不可逆抑制，但也导致囊泡完整性的显著丧失。用焦碳酸二乙酯处理膜囊泡导致Na⁺-H⁺交换失活，这不能归因于囊泡破裂或跨膜H⁺梯度的崩溃。焦碳酸二乙酯使Na⁺-H⁺交换失活遵循伪一级动力学至残留活性低于10%，可被羟胺逆转，表现为Vmax降低而Na⁺的Km不变，依赖于外部pH而非内部pH，被阿米洛利阻断，并被Na⁺增强。这些数据与以下假设一致：焦碳酸二乙酯敏感的咪唑残基是在动力学研究中发现的可滴定基团，在Na⁺-H⁺交换体的外部转运位点结合H⁺。