Two classes of ouabain binding sites in ferret heart and two forms of Na+-K+-ATPase

Journal Article · Fri May 01 04:00:00 EDT 1987 · Am. J. Physiol.; (United States)

OSTI ID:6501466

Ng, Y C; Akera, T

In partially purified Na+-K+-adenosinetriphosphatase (ATPase) obtained from ferret heart, ouabain produced a monophasic inhibition curve; however, the curve spanned over 5 logarithmic units, indicating the presence of more than one classes of enzyme. (/sup 3/H)ouabain binding studies revealed high-and low-affinity binding sites in approximately equal abundance, with apparent dissociation constants of 10 and 230 nM, respectively. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of phosphoenzyme formed from (gamma-/sup 32/P)ATP showed two distinct K+-sensitive bands of approximately 100,000 molecular weight. Phosphoenzyme formation from the high-molecular-weight alpha(+) form was selectively inhibited by N-ethylmaleimide. Ouabain caused a 50% inhibition of phosphorylation of the alpha(+) form at 40 nM and the lower-molecular-weight alpha form at 300 nM. In papillary muscle preparations, 1-30 nM ouabain produced a modest positive inotropic effect that reached an apparent plateau at 30 nM. Further increases in ouabain concentrations, however, produced additional and prominent inotropic effects at 0.1-10 microM. These results indicate for the first time in cardiac muscle that the high- and low-affinity ouabain binding sites are associated with the alpha(+) and alpha forms of the Na+-K+-ATPase, respectively, and that binding of ouabain to either of these sites causes enzyme inhibition and the positive inotropic effect.

• Research Organization: Michigan State Univ., East Lansing
• OSTI ID: 6501466
• Journal Information: Am. J. Physiol.; (United States), Journal Name: Am. J. Physiol.; (United States); ISSN AJPHA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ACID ANHYDRASES
AFFINITY
ANIMALS
ATP
ATP-ASE
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOCHEMICAL REACTION KINETICS
BODY
CARBOHYDRATES
CARDIAC GLYCOSIDES
CARDIOTONICS
CARDIOVASCULAR AGENTS
CARDIOVASCULAR SYSTEM
CHEMICAL REACTIONS
CONTRACTION
DAYS LIVING RADIOISOTOPES
DRUGS
ELECTROPHORESIS
ENZYMES
GLYCOSIDES
HEART
HYDROLASES
IN VITRO
ISOENZYMES
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
LABELLED COMPOUNDS
LIGHT NUCLEI
MAMMALS
MEMBRANE PROTEINS
MOLECULAR WEIGHT
MUSCLES
MYOCARDIUM
NUCLEI
NUCLEOTIDES
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
ORGANS
OUABAIN
PHOSPHOHYDROLASES
PHOSPHORUS 32
PHOSPHORUS ISOTOPES
PHOSPHORYLATION
PROTEINS
RADIOISOTOPES
REACTION KINETICS
RECEPTORS
STROPHANTHINS
TRACER TECHNIQUES
TRITIUM COMPOUNDS
VERTEBRATES