title: Release of a 22-kDa protein derived from the amino-terminal domain of the 49-kDa NIa of turnip mosaic potyvirus in Escherichia coli.
abstract: The coding region for the precursor 6K-small nuclear inclusion a (NIa) protein and for the NIa protein of turnip mosaic potyvirus (TuMV) were introduced into the plasmid pET-11d for high-level expression in Escherichia coli. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblot analyses of E. coli proteins showed that the NIa protein underwent endoproteolysis and released a 22-kDa polypeptide. NH2-terminal amino acid sequencing of the recombinant 22-kDa protein was performed and was identical to the predicted amino end of the NIa protein. Site-directed mutagenesis confirmed that the hydrolysis was associated with the NIa proteolytic activity and that the proteinase recognized a Glu residue within an amino acid sequence found in the NIa protein which fitted the TuMV consensus cleavage site sequence. Fusion of the 6K protein with the NIa protein partially inhibited the hydrolytic reaction. The recombinant 22-kDa protein is likely the VPg of TuMV.
authors.affiliation: Centre de recherche en virologie, Institut Armand-Frappier, Ville de Laval, Québec, Canada.
authors.forename: J F
authors.initials: JF
authors.lastname: Laliberté
authors.forename: O
authors.initials: O
authors.lastname: Nicolas
authors.forename: H
authors.initials: H
authors.lastname: Chatel
authors.forename: C
authors.initials: C
authors.lastname: Lazure
authors.forename: R
authors.initials: R
authors.lastname: Morosoli
mesh_terms: D001483:Base Sequence; D004279:DNA, Viral; D010450:Endopeptidases; D004926:Escherichia coli / Q000235:genetics; D008969:Molecular Sequence Data; D009029:Mosaic Viruses / Q000737:chemistry* / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D011994:Recombinant Proteins / Q000235:genetics; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D004279:DNA, Viral; D011994:Recombinant Proteins; D014764:Viral Proteins; D010450:Endopeptidases; C059992:nuclear inclusion protein a, mosaic viruses