Selective Assembly of Na,K-ATPase α2β2 Heterodimers in the Heart: DISTINCT FUNCTIONAL PROPERTIES AND ISOFORM-SELECTIVE INHIBITORS.
The Na,K-ATPase α2subunit plays a key role in cardiac muscle contraction by regulating intracellular Ca2+, whereas α1has a more conventional role of maintaining ion homeostasis. The β subunit differentially regulates maturation, trafficking, and activity of α-β heterodimers. It is not known whether the distinct role of α2in the heart is related to selective assembly with a particular one of the three β isoforms. We show here by immunofluorescence and co-immunoprecipitation that α2is preferentially expressed with β2in T-tubules of cardiac myocytes, forming α2β2heterodimers. We have expressed human α1β1, α2β1, α2β2, and α2β3in Pichia pastoris, purified the complexes, and compared their functional properties. α2β2and α2β3differ significantly from both α2β1and α1β1in having a higher K0.5K+and lower K0.5Na+for activating Na,K-ATPase. These features are the result of a large reduction in binding affinity for extracellular K+and shift of the E1P-E2P conformational equilibrium toward E1P. A screen of perhydro-1,4-oxazepine derivatives of digoxin identified several derivatives (e.g. cyclobutyl) with strongly increased selectivity for inhibition of α2β2and α2β3over α1β1(range 22-33-fold). Molecular modeling suggests a possible basis for isoform selectivity. The preferential assembly, specific T-tubular localization, and low K+affinity of α2β2could allow an acute response to raised ambient K+concentrations in physiological conditions and explain the importance of α2β2for cardiac muscle contractility. The high sensitivity of α2β2to digoxin derivatives explains beneficial effects of cardiac glycosides for treatment of heart failure and potential of α2β2-selective digoxin derivatives for reducing cardiotoxicity.
From the Department of Biomolecular Sciences and.
Michael
M
Habeck
the Department of Physiology, School of Medicine, UCLA and Veterans Affairs Greater Los Angeles Healthcare System, Los Angeles, California 90073.
Elmira
E
Tokhtaeva
From the Department of Biomolecular Sciences and.
Yotam
Y
Nadav
Israel National Centre for Personalized Medicine, Weizmann Institute of Science, Rehovoth 7610001, Israel.
Efrat
E
Ben Zeev
the Department of Biological Chemistry, University of Michigan Medical Center, Ann Arbor, Michigan 48109.
Sean P
SP
Ferris
the Department of Biological Chemistry, University of Michigan Medical Center, Ann Arbor, Michigan 48109.
Randal J
RJ
Kaufman
From the Department of Biomolecular Sciences and.
Elizabeta
E
Bab-Dinitz
the Department of Biochemistry and Molecular Genetics, University of Illinois, Chicago, Illinois 60607, and.
Jack H
JH
Kaplan
the Division of Pulmonary and Critical Care Medicine, Feinberg School of Medicine, Northwestern University, Chicago, Illinois 60611.
Laura A
LA
Dada
From the Department of Biomolecular Sciences and.|the School of Medicine, Tel Aviv University, Tel Aviv 6997801, Israel.
Zvi
Z
Farfel
From the Department of Biomolecular Sciences and.
Daniel M
DM
Tal
From the Department of Biomolecular Sciences and.
Adriana
A
Katz
the Department of Physiology, School of Medicine, UCLA and Veterans Affairs Greater Los Angeles Healthcare System, Los Angeles, California 90073.
George
G
Sachs
the Department of Physiology, School of Medicine, UCLA and Veterans Affairs Greater Los Angeles Healthcare System, Los Angeles, California 90073, olgav@ucla.edu.
Olga
O
Vagin
From the Department of Biomolecular Sciences and Steven.Karlish@weizmann.ac.il.
Steven J D
SJ
Karlish
D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D000818:Animals; D027682:Cation Transport Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D015816:Cell Adhesion Molecules, Neuronal / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D019281:Dimerization; D004791:Enzyme Inhibitors / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D051379:Mice; D009206:Myocardium / Q000737:chemistry / Q000201:enzymology*; D011188:Potassium / Q000737:chemistry / Q000378:metabolism; D012964:Sodium / Q000737:chemistry / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
C504191:Atp1b2 protein, mouse; D027682:Cation Transport Proteins; D015816:Cell Adhesion Molecules, Neuronal; D004791:Enzyme Inhibitors; D007527:Isoenzymes; D012964:Sodium; D000251:Adenosine Triphosphatases; C401838:Atp1a2 protein, mouse; D000254:Sodium-Potassium-Exchanging ATPase; D011188:Potassium
Na+/K+-ATPase; alpha2beta2; cardiomyocyte; excitation-contraction coupling (E-C coupling); isoform assembly; isoform-selective inhibitors; kinetics; medicinal chemistry; molecular modeling