A novel open reading frame in tobacco mosaic virus genome coding for a putative small, positively charged protein

• Published in: Biochimie Vol. 75; no. 8; pp. 659 - 665
• Main Authors: Morozov, S.Yu, Denisenko, O.N., Zelenina, D.A., Fedorkin, O.N., Solovyev, A.G., Maiss, E., Casper, R., Atabekov, J.G.
• Format: Journal Article
• Language: English
• Published: France Elsevier Masson SAS 1993
• Subjects: ADN RECOMBINADO; ADN RECOMBINE; Amino Acid Sequence; Base Sequence; Cell-Free System; DNA, Viral; DNA-Directed RNA Polymerases; Electrochemistry; GENETICA MOLECULAR; GENETIQUE MOLECULAIRE; GENOMAS; GENOME; GENOMES; in vitro translation; MOLECULAR GENETICS; Molecular Sequence Data; Mutagenesis, Site-Directed; NUCLEOTIDE SEQUENCE; Open Reading Frames; plant virus; PLANT VIRUSES; Protein Biosynthesis; PROTEINAS; PROTEINE; PROTEINS; RECOMBINANT DNA; RNA, Messenger - genetics; RNA, Messenger - metabolism; RNA, Viral - genetics; RNA, Viral - metabolism; SECUENCIA NUCLEICA; SEQUENCE NUCLEIQUE; TOBACCO MOSAIC TOBAMOVIRUS; tobacco mosaic virus; Tobacco Mosaic Virus - genetics; TOBAMOVIRUS DEL MOSAICO DEL TABACO; TOBAMOVIRUS MOSAIQUE DU TABAC; Viral Proteins - genetics; VIRUS DE LAS PLANTAS; VIRUS DES VEGETAUX; recombinant DNA; tobacco mosaic virus; in vitro translation; plant virus
• ISSN: 0300-9084; 1638-6183
• DOI: 10.1016/0300-9084(93)90096-B

From sequence comparisons between the tobamovirus genomes an open reading frame (ORF-X) potentially encoding a small, positively charged protein (33- to 45-amino-acids long) was found to overlap the immediate 3′ and 5′ sides of the transport protein gene and coat protein gene, respectively. In vitro translation of the monocistronic artificial transcripts generated with T7 RNA polymerase yielded a protein of M r 4000 (p4) and an unexpected trypsin-sensitive complex of M r 54000 that was resistant to reduction with 2-mercaptoethanol but could be dissociated by 8 M urea. Assembly of this complex was inhibited completely by site-directed mutagenesis within a conserved, positively charged 5-amino-acid long segment of the ORF-X protein. After centrifugation in low salt buffer the 54-kDa complex remained mostly associated with ribosomes. Apparently this complex represents a specific aggregate of the p4 product of ORF-X with a protein of approximate M r 50000 that is a component of the translation apparatus.