Bacterial expression of human respiratory syncytial viral phosphoprotein P and identification of Ser237 as the site of phosphorylation by cellular casein kinase II.
The phosphoprotein P gene of human respiratory syncytial virus has been cloned and the protein expressed in Escherichia coli. The expressed protein was soluble, unphosphorylated, and constituted approximately 10% of the total bacterial protein. Electrophoretic and antigenic analyses demonstrated the identity of the recombinant protein with viral P protein and P protein synthesized in reticulocyte lysates. Purified recombinant P protein was efficiently phosphorylated in vitro by purified native as well as recombinant casein kinase II (CKII) or by the CKII activity in uninfected cell extracts. Through deletions and site-directed mutagenesis, the site of CKII phosphorylation was mapped to a single serine residue (Ser237) near the C-terminal end of the P protein.
Mazumder
B
B
Department of Biochemistry and Molecular Biology, College of Medicine, University of South Alabama, Mobile 36688-0002.
Adhikary
G
G
Barik
S
S
D000595:Amino Acid Sequence; D001483:Base Sequence; D047390:Casein Kinase II; D002384:Catalysis; D003001:Cloning, Molecular; D017931:DNA Primers; D004926:Escherichia coli / Q000235:genetics*; D015707:HN Protein; D006801:Humans; D008969:Molecular Sequence Data; D010710:Phosphates; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012136:Respiratory Syncytial Viruses / Q000378:metabolism*; D012694:Serine / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D014759:Viral Envelope Proteins; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism
D017931:DNA Primers; D015707:HN Protein; D010710:Phosphates; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins; D014764:Viral Proteins; C116204:attachment protein G; D012694:Serine; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases; D010744:Phosphoric Monoester Hydrolases