Autocatalytic processing of the potyvirus helper component proteinase in Escherichia coli and in vitro.
The virus-encoded proteins of tobacco etch virus (TEV), a plant potyvirus, arise by proteolytic processing of a large polyprotein precursor. The TEV genome codes for two proteinases, a 49-kilodalton proteinase and helper component proteinase (HC-Pro), which cleave the polyprotein at specific sites. The only known cleavage event catalyzed by HC-Pro occurs at the HC-Pro carboxyl terminus. The proteolytic activity of HC-Pro was analyzed by expression of the enzyme in bacterial and cell-free systems. The carboxyl-terminal domain of HC-Pro exhibited proteolytic activity in Escherichia coli with a processing half-time of approximately 100 s. The processing kinetics of HC-Pro expressed in vitro by cell-free transcription and translation was variable, depending on the presence or absence of TEV polypeptide sequences at the amino terminus of the proteolytic domain. Cleavage of the HC-Pro carboxyl terminus appeared to proceed exclusively by an autocatalytic mechanism; the proteinase synthesized in vitro exhibited little or no proteolytic activity when reacted with the HC-Pro cleavage site in trans or biomolecular reactions.
Carrington
J C
JC
Department of Biology, Texas A&M University, College Station 77843-3258.
Freed
D D
DD
Sanders
T C
TC
D002474:Cell-Free System; D010450:Endopeptidases / Q000096:biosynthesis*; D004926:Escherichia coli / Q000201:enzymology*; D006378:Helper Viruses / Q000201:enzymology*; D010942:Plant Viruses / Q000201:enzymology*; D014176:Protein Biosynthesis
D010450:Endopeptidases