Rice tungro spherical virus polyprotein processing: identification of a virus-encoded protease and mutational analysis of putative cleavage sites.
Rice tungro spherical virus encodes a large polyprotein containing motifs with sequence similarity to viral serine-like proteases and RNA polymerases. Polyclonal antisera raised against domains of the putative protease and polymerase in fusion with glutathione S-transferase detected a protein of about 35 kDa and, in very low amounts, a protein of about 70 kDa, respectively, in extracts from infected plants. In in vitro transcription/translation systems and in Escherichia coli we demonstrated a proteolytic activity in the C-terminal region of the polyprotein. This protease rapidly cleaved its polyprotein precursors in vitro. Mutating a potential cleavage site located N-terminal to the protease domain, Gln2526-Asp2527, diminished processing. The transversion mutation at the putative C-terminal cleavage site of the protease, at Gln2852-Ala2853, led to a delayed and partial processing.
Department of Virus Research, John Innes Centre, Norwich Research Park, United Kingdom.
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D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D012321:DNA-Directed RNA Polymerases / Q000235:genetics / Q000378:metabolism; D010450:Endopeptidases / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D012275:Oryza / Q000821:virology*; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D019192:Sequivirus / Q000235:genetics* / Q000378:metabolism*; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
D017931:DNA Primers; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; D012321:DNA-Directed RNA Polymerases; D010450:Endopeptidases