Proteolytic processing of the MHV polymerase polyprotein. Identification of the P28 cleavage site and the adjacent protein, P65
- PMID: 8830520
Proteolytic processing of the MHV polymerase polyprotein. Identification of the P28 cleavage site and the adjacent protein, P65
Abstract
The polymerase gene of Mouse Hepatitis Virus strain JHM (MHV-JHM) encodes a polyprotein larger than 750 kilodaltons. This polyprotein is proposed to be processed by several viral proteinases into functional subunits. The amino-terminal subunit is p28, which is cleaved by the first viral papain-like proteinase domain. In this study, we identified the cleavage site of this papain-like cysteine proteinase by amino acid sequencing of radiolabeled polypeptide adjacent to p28. Proteolysis occurs between the glycine-247 and valine-248 dipeptide bond. To determine which amino acid residues are critical for proteolysis, we preformed site-directed mutagenesis on the coding sequences surrounding the cleavage site and assayed for the efficiency of cleavage of p28 in an in vitro transcription and translation system. We report that glycine-247 and arginine-246 are the most critical residues for efficient processing of p28.
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