title: Hepatitis C virus NS5A protein is phosphorylated in vitro by a stably bound protein kinase from HeLa cells and by cAMP-dependent protein kinase A-alpha catalytic subunit.
abstract: Hepatitis C virus (HCV) has a positive-strand RNA genome that codes for a polyprotein precursor, which is processed co- and post-translationally by cellular and viral proteinases into three structural and at least six non-structural (NS) proteins. The NS5A protein, expressed in mammalian cells, exists in two phosphorylated forms of 56-kDa and 58-kDa. In this study, we provide evidence for a stable association between NS5A and a protein kinase from HeLa cells and hepatocellular carcinoma (HepG2) cells by co-immunoprecipitation and by affinity to immobilized glutathione-S-transferase (GST)-NS5A fusion protein produced in E. coli. This protein kinase could phosphorylate in vitro the native NS5A on serine residues, (GST)-NS5A, histone H1, and casein as substrates. In addition, the GST-NS5A was also phosphorylated in vitro by the cAMP-dependent protein kinase A-alpha catalytic subunit.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City 66160-7421, USA.
authors.forename: Y
authors.initials: Y
authors.lastname: Ide
authors.forename: A
authors.initials: A
authors.lastname: Tanimoto
authors.forename: Y
authors.initials: Y
authors.lastname: Sasaguri
authors.forename: R
authors.initials: R
authors.lastname: Padmanabhan
mesh_terms: D001665:Binding Sites; D002364:Caseins / Q000378:metabolism; D002384:Catalysis; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D004800:Enzymes, Immobilized; D004926:Escherichia coli / Q000378:metabolism; D015870:Gene Expression; D005982:Glutathione Transferase / Q000235:genetics; D006367:HeLa Cells; D006657:Histones / Q000378:metabolism; D006801:Humans; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014407:Tumor Cells, Cultured; D017361:Viral Nonstructural Proteins / Q000235:genetics / Q000378:metabolism*; D012324:RNA-Dependent RNA Polymerase
chemical_list: D002364:Caseins; D017868:Cyclic AMP-Dependent Protein Kinases; D004800:Enzymes, Immobilized; D005982:Glutathione Transferase; D006657:Histones; D017346:Protein Serine-Threonine Kinases; D011993:Recombinant Fusion Proteins; D017361:Viral Nonstructural Proteins; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase