Sodium-potassium-activated adenosine triphosphatase of electrophorus electric organ. X. Immunochemical properties of the Lubrol-solubilized enzume and its constituent polypeptides

• Published in: The Journal of biological chemistry Vol. 250; no. 3; pp. 1035 - 1040
• Main Authors: Jean, D H, Albers, R W, Koval, G J
• Format: Journal Article
• Language: English
• Published: United States American Society for Biochemistry and Molecular Biology 10-02-1975
• Subjects: Adenosine Triphosphatases - immunology; Adenosine Triphosphatases - metabolism; Adenosine Triphosphatases - pharmacology; Animals; Antigen-Antibody Reactions; Detergents; Electric Organ - enzymology; Electrophoresis, Polyacrylamide Gel; Electrophorus; Enzyme Activation - drug effects; Immunodiffusion; Kinetics; Macromolecular Substances; Molecular Weight; Peptides - immunology; Rabbits - immunology; Sodium - pharmacology; Solubility; Time Factors
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(19)41888-7

Detergent (Lubrol WX)-solubilized sodium-potassium-activated adenosine triphosphatase ((Na+ + K+)-ATPase) of electrophorus
electric organ contains two major constituent polypeptides with molecular weights of 96,000 and 58,000 which can be readily
demonstrated by sodium dodecyl sulfate polyacrylamide gel electrophoresis. These two polypeptides can be clearly separated
and can be obtained in milligram quantities by preparative sodium dodecyl sulfate gel electrophoresis. The separated polypeptides,
after removal of sodium dodecyl sulfate, and Lubrol-solubilized (Na+ + K+)-ATPase activity to some degree. Moreover, the degree
of inhibition is directly proportional to the increasing amounts of antisera. The inhibition is maximal 4 weeks after the
first injection. Immunodiffusion in 1% agar gel indicated that only Lubrol-solubilized enzyme antiserum, but not 58,000-dalton
or 96,00-dalton polypeptide antiserum, gives one major precipitin band. However, specific complex formation between each polypeptide
antiserum and Lubrol-solubilized enzyme occurs. This was demonstrated indirectly. After incubating Lubrol-solubilized enzyme
with increasing amounts of polypeptide antisera at 37 degrees for 15 min, they were placed in the side wells of an immunodiffusion
plate with antiserum against Lubrol-solubilized enzyme in the central well. The intensity of the precipitin band decreased
with increasing amounts of polypeptide antisera. Thus, the results indicate that both 96,000-dalton and 58,000-dalton polypeptides
are integral subunits of (Na+ + K+)-ATPase.