AJP - Cell Physiology, Vol 262, Issue 6 C1491-C1499, Copyright © 1992 by American Physiological Society
Determination of Na(+)-K(+)-ATPase alpha- and beta-isoforms and kinetic properties in mammalian liver
Y. Sun and W. J. Ball Jr
Department of Pharmacology, College of Medicine, University of Cincinnati, Ohio 45267-0575.
While Western blot analysis clearly revealed the presence of the alpha- and
beta-subunits of Na(+)-K(+)-ATPase in a variety of rat tissues, beta was
not readily detectable in liver. This observation was consistent with a
previous report indicating that Na(+)-K(+)-ATPase immunoprecipitated from
rat liver gives no clear evidence for the presence of a beta-subunit
(Hubert et al. Biochemistry 25: 4156-4163, 1986). However, Western blot
analysis of density gradient-purified lamb and rat liver microsomes showed
the presence of a protein with an approximate molecular mass of 42 kDa that
was immunoreactive with beta-specific polyclonal antibodies as well as
beta-directed monoclonal antibodies. Deglycosylation of this protein by
N-glycosidase F generated a core protein (beta c, M(r) approximately
32,000) that had the identical electrophoretic mobility as the beta c
protein of the purified kidney enzyme. Isoform-specific monoclonal and
synthetic peptide-directed polyclonal antibodies were used to demonstrate
the presence of only the alpha 1- and beta 1-proteins in the liver and the
presence of beta 2 in rat brain. Functional studies then showed that
although both rat and lamb liver enzymes had sensitivities to cardiac
glycoside inhibition similar to that of their corresponding kidney enzyme,
the lamb liver enzyme had higher affinities for Na+, K+, and ATP than the
kidney enzyme.
