Residues within transmembrane domains 4 and 6 of the Na,K-ATPase alpha subunit are important for Na+ selectivity.
Residues within transmembrane domains 4 and 6 of the Na,K-ATPase alpha subunit are important for Na+ selectivity.
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Na,K-ATPase α 亚基跨膜域 4 和 6 内的残基对于 Na 选择性很重要。
DOI：
10.1021/bi049484s
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发表时间：
2004
期刊：
影响因子：
--
通讯作者：
Blanco,Gustavo
中科院分区：
文献类型：
--
作者：
Sanchez,Gladis;Blanco,Gustavo
The Na,K- and H,K-ATPases are plasma membrane enzymes responsible for the active exchange of extracellular K+for cytoplasmic Na+or H+, respectively. At present, the structural determinants for the specific function of these ATPases remain poorly understood. To investigate the cation selectivity of these ATPases, we constructed a series of Na,K-ATPase mutants in which residues in the membrane spanning segments of the α subunit were changed to the corresponding residues common to gastric H,K-ATPases. Thus, mutants were created with substitutions in transmembrane domains TM1, TM4, TM5, TM6, TM7, and TM8 independently or together (designated TMAll). The function of each mutant was assessed after coexpression with the β subunit inSf-9 cells using baculoviruses. The enzymatic properties of TM1, TM7, and TM8 mutants were similar to the wild-type Na,K-ATPase, and while TM5 showed modest changes in apparent affinity for Na+, TM4, TM6, and TMAll displayed an abnormal activity. This resulted in a Na+-independent hydrolysis of ATP, a 2-fold higherK0.5for Na+activation, and the ability to function at low pH. These results suggest a loss of discrimination for Na+over H+for the enzymes. In addition, TM4, TM6, and TMAll mutants exhibited a 1.5-fold lower affinity for K+and a 4−5-fold decreased sensitivity to vanadate. Altogether, these results provide evidence that residues in transmembrane domains 4 and 6 of the α subunit of the Na,K-ATPase play an important role in determining the specific cation selectivity of the enzyme and also its E1/E2 conformational equilibrium.
