1887

Abstract

Monopartite members of the family utilize three virus-encoded proteinases to cleave the viral polyprotein into mature proteins. The amino-terminal region of the viral polyprotein is autolytically cleaved by the P1 proteinase. A domain required for P1 proteinase activity of (WSMV) was mapped using a series of templates with nested 3′-truncations or 5′-deletions to program transcription–translation reactions. The WSMV P1 proteinase cleavage site was mapped to a position downstream of amino acid residue 348 and upstream of amino acid residue 353, with the peptide bond between amino acid residues Y and G the most probable site of hydrolysis. An alignment of potyvirus polyprotein sequences in the carboxy-terminal region of the P1 domain revealed WSMV P1 contained conserved H, D, S and FIVXG residues upstream of the cleavage site that are typical of serine proteinases and shown by others to be required for P1 proteolysis in . Insertion of the GUS reporter gene immediately downstream of the P1 cleavage site in a full-length clone of WSMV resulted in systemic infection and GUS expression upon inoculation of plants with transcripts. When cleaved by P1 at the amino terminus and NIa proteinase at a site engineered in the carboxy-terminus, active GUS protein expressed by WSMV in infected wheat had electrophoretic mobility similar to wild-type GUS protein.

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2002-02-01
2020-07-15
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