Gene expression by a hypovirulence-associated virus of the chestnut blight fungus involves two papain-like protease activities. Essential residues and cleavage site requirements for p48 autoproteolysis

Proteolytic processing plays a fundamental role in gene expression of a recently characterized viral-like double-stranded RNA associated with biological control of the chestnut blight fungus. Polypeptide p29, a papain-like protease, was shown to autocatalytically release itself from the NH2 terminus...

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Bibliographic Details
Published in:The Journal of biological chemistry 1991-10, Vol.266 (29), p.19419-19425
Main Authors: Shapira, R. (Roche Institute of Molecular Biology, Nutley, NJ), Nuss, D.L
Format: Article
Language:English
Subjects:
Amino Acid Sequence
ARN
Ascomycota - metabolism
Base Sequence
Biological and medical sciences
Catalysis
CONTROL BIOLOGICO
CRYPHONECTRIA PARASITICA
Electrophoresis, Polyacrylamide Gel
Endopeptidases - metabolism
EXPRESION GENICA
EXPRESSION DES GENES
Fundamental and applied biological sciences. Psychology
Gene Expression
Genes, Viral
Hydrolysis
LUTTE BIOLOGIQUE
Molecular and cellular biology
Molecular genetics
Molecular Sequence Data
Mutation
Papain - metabolism
PODER PATOGENO
Polymerase Chain Reaction
POUVOIR PATHOGENE
PROTEASAS
PROTEASE
Protein Biosynthesis
RNA Viruses - genetics
RNA Viruses - metabolism
RNA, Double-Stranded - genetics
RNA, Viral - genetics
Sequence Alignment
Transcription, Genetic
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Summary:Proteolytic processing plays a fundamental role in gene expression of a recently characterized viral-like double-stranded RNA associated with biological control of the chestnut blight fungus. Polypeptide p29, a papain-like protease, was shown to autocatalytically release itself from the NH2 terminus of the polyprotein specified by the first of two encoded open reading frames, ORF A (Choi, G. H., Shapira, R., and Nuss, D. L. (1991) Proc. Natl. Acad. Sci. U. S. A. 88, 1167-1171; Choi, G. H., Pawlyk, D. M.. and Nuss, D. L. (1991) Virology 183, 747-752). The characterization of a second autocatalytic protease, p48, which is encoded by ORF B, is the subject of this report. Deletion analysis revealed that the catalytic domain resides within the carboxyl-terminal region, while site-specific mutational analysis identified Cys-341 and His-388 residues essential for autoproteolysis. Autoproteolytic processing by p48 was also demonstrated when expressed in Escherichia coli and microsequence analysis of the recovered COOH-terminal cleavage product indicated that cleavage occurred between Gly-418 Ala-419. The requirements for a functional cleavage site, including confirmation of the cleavage dipeptide, were defined by amino acid substitution analysis. Similarities between p29 and p48 suggest that the respective coding domains could have arisen as a result of a gene duplication event
ISSN:0021-9258
1083-351X
DOI:10.1016/s0021-9258(18)55013-4