These tools will no longer be maintained as of December 31, 2024. Archived website can be found here. PubMed4Hh GitHub repository can be found here. Contact NLM Customer Service if you have questions.
Pubmed for Handhelds
PUBMED FOR HANDHELDS
Search MEDLINE/PubMed
Title: Functional consequences of mutation Asn326-->Leu in the 4th transmembrane segment of the alpha-subunit of the rat kidney Na+, K(+)-ATPase. Author: Vilsen B. Journal: FEBS Lett; 1995 Apr 17; 363(1-2):179-83. PubMed ID: 7729543. Abstract: Site-specific mutagenesis was used to replace Asn326 in transmembrane segment M4 of the ouabain-insensitive alpha 1-isoform of rat kidney Na+, K(+)-ATPase. Mutant Asn326-->Leu was functional as demonstrated by the ability of COS cells expressing the mutant enzyme to grow in the presence of ouabain. In three independent assays encompassing Na+ titrations of Na+,K(+)-ATPase activity, Na(+)-ATPase activity, and phosphorylation from ATP, the Asn326-->Leu mutant displayed a reduced apparent affinity for Na+. By contrast, this mutant exhibited a slightly increased apparent affinity for K+ relative to the wild-type enzyme. In the presence of Na+ without K+, the Asn326-->Leu mutant hydrolyzed ATP at a high rate corresponding to 32% of the maximal Na+,K(+)-ATPase activity, and the rate of dephosphorylation of the phosphoenzyme intermediate was enhanced in the mutant relative to that of the wild-type enzyme. Oligomycin, known to stabilize the Na(+)-occluded phosphoenzyme intermediate, reduced the dephosphorylation rate of the mutant and increased the steady-state phosphoenzyme level formed by the mutant at least 3-fold, whereas an increase in the steady-state phosphoenzyme level of only 10-15% was determined for the wild-type enzyme. The molecular turnover number for the Na+,K(+)-ATPase reaction, calculated when the steady-state phosphoenzyme level obtained in the presence of oligomycin was taken as a measure of the concentration of active sites, was slightly reduced relative to that of the wild-type enzyme. The data are discussed in terms of a role for Asn326 in binding of cytoplasmic Na+ and in mediation of inhibition of dephosphorylation.[Abstract] [Full Text] [Related] [New Search]