PROTEOLYTIC MATURATION OF THE 206-KDA NONSTRUCTURAL PROTEIN ENCODED BY TURNIP YELLOW MOSAIC-VIRUS RNA

The longest open reading frame of turnip yellow mosaic virus genomic RNA (ORF-206) encodes a 206-kDa nonstructural protein. The most prominentin vitro translation products of ORF-206 are the full-length p206 and a shorter N-coterminal 150-kDa protein. We have confirmed these assignments by immunoprecipitation ofin vitro translation products with antisera raised to N-terminal and C-terminal regions encoded by ORF-206. The mechanism by which the 150-kDa protein arises from ORF-206 was investigated byin vitro translation of deletion and substitution derivatives transcribed from pTYMC, a cDNA clone of TYMV RNA. The following observations demonstrate that the 150-kDa protein and a C-terminal 70-kDa protein arise from ORF-206 by autoproteolysis: (1) Two regions encoded by ORF-206 were necessary for the formation of the 150-kDa protein: a domain between amino acids 555 and 1051, postulated to encode a protease, and the region between amino acids 1253 and 1261, thought to constitute the protease recognition and/or cleavage site. (2) Mutants with substitutions between amino acids 1253 and 1261 that produce low levels of the 150-kDa protein inin vitro translations also have high levels of p206 and low levels of the 70-kDa protein. (3) The rate of formation of the 150-kDa protein is dilution insensitive, suggesting that proteolysis occurs mainlyin cis. © 1991 Academic Press, Inc.