IDENTIFICATION OF THE CLEAVAGE SITE RECOGNIZED BY THE TURNIP YELLOW MOSAIC-VIRUS PROTEASE

The noncapsid protein expressed from ORF-206 of turnip yellow mosaic v irus (TYMV) is autocatalytically processed by a papain-like protease, producing N-terminal 150-kDa and C-terminal 70-kDa proteins. By introd ucing two methionine residues near the N-terminus of the 70-kDa protei n, we have obtained N-terminal amino acid sequence of that protein pro duced from [S-35] methionine-labeled in vitro translations. The introd uction of methionine residues was demonstrated to not interfere with v iral replication or proteolysis, as assayed by inoculating mutant RNA transcripts onto whole plants and protoplasts, as well as by translati ng the RNAs in a rabbit reticulocyte lysate. This has allowed us to de termine that the TYMV protease cleaves between alanine(1259) and threo nine(1250) of the precursor protein p206, yielding proteins of calcula ted M(r) 140,618 and 66,037, which will be referred to henceforth as p 141 and pas, respectively. The sequence context around the cleavage si te is LNGA/TP. (C) 1996 Academic Press, Inc.