大鼠内髓集合管中由钠钾ATP酶介导的铵根离子转运
NH+4 transport mediated by Na(+)-K(+)-ATPase in rat inner medullary collecting duct.
作者信息
Wall S M, Koger L M
机构信息
Division of Nephrology, University of Texas Medical School at Houston 77030.
出版信息
Am J Physiol. 1994 Oct;267(4 Pt 2):F660-70. doi: 10.1152/ajprenal.1994.267.4.F660.
The interaction of K+ and NH+4 on the basolateral membrane of cultured rat inner medullary collecting duct cells (IMCD) was explored using 86Rb+ as a K+ congener. Ouabain addition decreased Rb+ uptake to 22.0 +/- 2.6% of control. Possible ouabain-sensitive NH+4 transport was therefore explored. Replacement of N-methyl-D-glucamine chloride with NH4Cl or KCl decreased both total and ouabain-sensitive Rb+ uptake. This inhibition of Rb+ uptake by NH+4 was not due to changes in intracellular pH or differences in cell viability. We conclude that NH+4 competes with K+ for binding on the Na(+)-K(+)-adenosinetriphosphatase (ATPase). An inhibitory constant (Ki) for NH+4 of 11.0 +/- 1.6 and a Michaelis constant (Km) for K+ of 1.9 +/- 0.7 mM were measured. To extend these observations, NH+4 was tested as a substrate for ouabain-sensitive ATPase activity in native permeabilized IMCD cells. With 20 mM K+, activity was 202 +/- 73 nmol ATP hydrolyzed.min-1.mg protein-1, and with 20 mM NH+4, activity was 259 +/- 81 nmol ATP hydrolyzed.min-1.mg protein-1. Thus NH+4 substitution for K+ on the Na(+)-K(+)-ATPase is a likely mechanism for NH+4 uptake by the IMCD.
以⁸⁶Rb⁺作为K⁺的同类物，研究了K⁺和NH₄⁺在培养的大鼠内髓集合管细胞（IMCD）基底外侧膜上的相互作用。加入哇巴因后，Rb⁺摄取量降至对照的22.0±2.6%。因此，对可能存在的哇巴因敏感的NH₄⁺转运进行了研究。用NH₄Cl或KCl替代N - 甲基 - D - 葡萄糖胺氯化物会降低总的和哇巴因敏感的Rb⁺摄取量。NH₄⁺对Rb⁺摄取的这种抑制作用并非由于细胞内pH值的变化或细胞活力的差异。我们得出结论，NH₄⁺与K⁺竞争结合在钠钾腺苷三磷酸酶（ATP酶）上。测得NH₄⁺的抑制常数（Ki）为11.0±1.6，K⁺的米氏常数（Km）为1.9±0.7 mM。为了扩展这些观察结果，在天然通透的IMCD细胞中测试了NH₄⁺作为哇巴因敏感的ATP酶活性的底物。在20 mM K⁺存在下，活性为202±73 nmol ATP水解·min⁻¹·mg蛋白⁻¹，在20 mM NH₄⁺存在下，活性为259±81 nmol ATP水解·min⁻¹·mg蛋白⁻¹。因此，NH₄⁺替代K⁺与钠钾ATP酶结合可能是IMCD摄取NH₄⁺的一种机制。
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