Virus-specified protease in poliovirus-infected HeLa cells

Area of Science:

• Virology
• Molecular Biology
• Biochemistry

Background:

• Picornavirus replication involves complex protein processing.
• Cellular proteases perform initial cleavages of viral polyproteins.
• The enzymes responsible for secondary cleavages remained largely uncharacterized.

Purpose of the Study:

• To characterize the enzyme responsible for secondary cleavages of picornavirus precursors in infected cells.
• To determine the origin and function of this protease in viral capsid polypeptide production.

Main Methods:

• Enzyme kinetics analysis (pH and temperature optima).
• Inhibition studies using guanidine and cycloheximide.
• Amino acid analog incorporation and protease inhibitor assays.
• Copurification with viral polypeptides.

Main Results:

• Protease activity kinetics correlated with viral protein synthesis.
• The enzyme exhibited specific pH and temperature optima.
• Protease production was inhibited by guanidine and cycloheximide.
• The enzyme was synthesized late in infection, independent of host protein synthesis, and copurified with a 40 kDa viral polypeptide.
• Viral mutants defective in coat protein synthesis still produced wild-type protease activity.

Conclusions:

• A non-structural poliovirus gene product is responsible for secondary cleavages of viral precursors.
• This viral protease is essential for producing mature viral coat proteins.
• The findings elucidate a key step in picornavirus assembly and replication.