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Epitope‐Tagged Constructs of the Yeast Plasma‐Membrane H+‐ATPase
Author(s) -
Santos Carlos F. Tourinho dos,
Dewitt Natalie D.,
Gupta Soma Sen,
Allen Kenneth E.,
Slayman Carolyn W.
Publication year - 2000
Publication title -
iubmb life
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.132
H-Index - 113
eISSN - 1521-6551
pISSN - 1521-6543
DOI - 10.1080/15216540050022494
Subject(s) - vanadate , epitope , atpase , yeast , chemistry , biochemistry , vesicle , v atpase , secretion , secretory vesicle , membrane , microbiology and biotechnology , enzyme , biology , antigen , genetics
In this study, two different epitope tags (HA, c‐myc) were introduced near the N terminus of the yeast PMA1 H+‐ATPase. The resulting proteins were indistinguishable from the wild‐type ATPase in their ability to travel through the secretory pathway, as judged by quantitative immunoblotting of isolated secretory vesicles. Furthermore, there were no significant abnormalities in ATPase activity (including Km for MgATP, Vmax, pH optimum, and IC50 for inhibition by vanadate) or in ATP‐dependent proton pumping. Finally, the epitope‐tagged ATPases could support normal growth and displayed the expected activation by glucose.