Open AccessProduction and characterization of a monoclonal antibody to a human interferon-induced double-stranded RNA-binding Mr 68,000 protein kinase.
Author(s) -
Linda Penn,
Bryan R.G. Williams
Publication year - 1985
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.82.15.4959
Subject(s) - monoclonal antibody , immunoprecipitation , microbiology and biotechnology , protein kinase a , protein kinase r , interferon , biology , rna , protein kinase inhibitor , affinity chromatography , biochemistry , antibody , phosphorylation , cyclin dependent kinase 2 , virology , enzyme , gene , immunology
One of the interferon-induced proteins thought to be involved in the antiviral effects of interferon is a double-stranded RNA-dependent protein kinase. This paper reports the development of a monoclonal antibody, 10A5, that recognizes a protein that co-migrates with the double-stranded RNA-dependent protein kinase at an approximate molecular weight of 68,000. Levels of this protein and of the protein kinase activity increase 3-fold on interferon treatment of T98G cells. The specificity of the monoclonal antibody was determined by ELISA, immunoblotting, and immunoprecipitation procedures. Furthermore, immunoaffinity chromatography of an interferon-induced T98G cell extract previously phosphorylated in the presence of double-stranded RNA and radiolabeled ATP resulted in the specific elution of a phosphorylated Mr 68,000 protein from the monoclonal antibody 10A5-Sepharose column. Monoclonal antibody 10A5 recognizes both native and denatured protein kinase, independent of double-stranded RNA binding or phosphorylation, and should therefore serve as a useful tool in analyzing the role of the double-stranded RNA-dependent protein kinase in the mechanism of interferon action.