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Mutational analysis of the encephalomyocarditis virus primary cleavage
1Institute for Molecular Virology, University of Wisconsin, Madison 53706, USA.
Journal of Virology
|October 1, 1996
Summary
Mutations in the encephalomyocarditis virus DvExNPGP sequence severely impaired viral polyprotein processing and were lethal. This highlights the critical role of this sequence in viral replication and capsid precursor maturation.
Area of Science:
- Virology
- Molecular Biology
- Biochemistry
Background:
- The conserved DvExNPGP motif is crucial for primary polyprotein cleavage in cardioviruses and aphthoviruses.
- Encephalomyocarditis virus (EMCV) polyprotein processing is essential for viral replication.
Purpose of the Study:
- To investigate the role of the DvExNPGP sequence in EMCV polyprotein processing.
- To determine the impact of mutations within this motif on viral replication.
Main Methods:
- Site-directed mutagenesis was used to create sixteen substitution mutations in the EMCV DvExNPGP sequence.
- Mutated viral cDNA was expressed and characterized for processing activity in cell-free systems.
- Representative mutations were tested in full-length genomic contexts to assess their effect on viral viability.
Main Results:
- Nearly all DvExNPGP mutations significantly reduced the efficiency of primary polyprotein cleavage.
- Mutations in the DvExNPGP sequence were lethal when tested in full-length viral genomes, with no observed revertants.
- Both primary cleavage and subsequent processing of the P1 capsid precursor by 3C protease were impaired by these mutations.
Conclusions:
- The natural DvExNPGP sequence is strictly required for efficient primary polyprotein cleavage in EMCV.
- Primary cleavage is essential for the proper processing of the viral capsid precursor.
- Disruption of the DvExNPGP motif leads to lethal viral phenotypes, underscoring its critical role in the viral life cycle.