Expression and characterization of a recombinant murine coronavirus 3C-like proteinase.
The replication of coronaviruses involves proteolytic processing of the gene 1 translation products, pp1a and pp1ab. One of the key enzymes in this process is predicted to be a virus-encoded 3C-like proteinase. In this report, we describe a bacterial system that has allowed us to express and characterize a recombinant murine coronavirus (MHV-JHM) 3C-like proteinase. The partially purified protein has been shown to exhibit proteolytic activity in trans and mutation analysis has been used to demonstrate the indispensability of Cys-3495 for enzymatic activity. Finally, the effect of class-specific proteinase inhibitors on the trans cleavage activity of the MHV 3C-like proteinase has been used to demonstrate the functional and structural homology of this enzyme to the picornavirus 3C proteinases.
Seybert
A
A
Institute of Virology, University of Würzburg, Germany.
Ziebuhr
J
J
Siddell
S G
SG
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D000086782:Coronavirus 3C Proteases; D003546:Cysteine Endopeptidases / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers; D007700:Kinetics; D051379:Mice; D008969:Molecular Sequence Data; D006517:Murine hepatitis virus / Q000201:enzymology* / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D016366:Open Reading Frames; D016133:Polymerase Chain Reaction; D011480:Protease Inhibitors / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000378:metabolism; D013379:Substrate Specificity
D017931:DNA Primers; D011480:Protease Inhibitors; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D003546:Cysteine Endopeptidases; D000086782:Coronavirus 3C Proteases