Related Experiment Video

Updated: Aug 15, 2026

Isolation of Fidelity Variants of RNA Viruses and Characterization of Virus Mutation Frequency
18:10

Isolation of Fidelity Variants of RNA Viruses and Characterization of Virus Mutation Frequency

Published on: June 16, 2011

Effects of mutations in poliovirus 3Dpol on RNA polymerase activity and on polyprotein cleavage

C C Burns1, M A Lawson, B L Semler

  • 1Department of Biochemistry, University of Utah School of Medicine, Salt Lake City 84132.

Journal of Virology
|November 1, 1989
PubMed

Insights

Introducing mutations into the poliovirus 3Dpol gene resulted in nonviable viruses. Mutant 3Dpol proteins showed defective RNA polymerase activity, impacting viral replication.

Area of Science:

  • Virology
  • Molecular Biology
  • Biochemistry

Background:

  • Poliovirus replication relies on viral-encoded proteins, including RNA-dependent RNA polymerase (3Dpol).
  • Proper processing of viral proteins is essential for infectivity.

Purpose of the Study:

  • To investigate the functional impact of mutations within the poliovirus 3Dpol gene.
  • To determine how sequence alterations affect 3Dpol's enzymatic activities and overall viral viability.

Main Methods:

  • Introduction of insertion mutations into the poliovirus 3Dpol gene.
  • Testing mutant viral cDNA for infectivity.
  • Expression and in vitro analysis of mutant 3CD (protease-polymerase) and released 3Dpol proteins in bacteria (Escherichia coli).

Main Results:

  • All 3Dpol mutants rendered the virus nonviable.
  • Mutant 3Dpol proteins exhibited defective RNA-dependent RNA polymerase activity in vitro.
  • Some mutant 3CD proteins showed impaired protease activity essential for capsid precursor processing.

Conclusions:

  • Alterations in the poliovirus 3Dpol gene significantly disrupt its RNA polymerase function.
  • Specific mutations affect both polymerase and protease activities, highlighting the multifunctional nature of 3CD.
  • The integrity of 3Dpol is critical for poliovirus replication and viability.

More Related Videos

Analysis of RNA Processing Reactions Using Cell Free Systems: 3' End Cleavage of Pre-mRNA Substrates in vitro09:16

Analysis of RNA Processing Reactions Using Cell Free Systems: 3' End Cleavage of Pre-mRNA Substrates in vitro

Published on: May 3, 2014

Analysis of Group IV Viral SSHHPS Using In Vitro and In Silico Methods10:40

Analysis of Group IV Viral SSHHPS Using In Vitro and In Silico Methods

Published on: December 21, 2019

Related Concept Videos

Proofreading01:43

Proofreading

Synthesis of new DNA molecules starts when DNA polymerase links nucleotides together in a sequence that is complementary to the template DNA strand. DNA polymerase has a higher affinity for the correct base to ensure fidelity in DNA replication. The DNA polymerase furthermore proofreads during replication, using an exonuclease domain that cuts off incorrect nucleotides from the nascent DNA strand.Errors during Replication Are Corrected by the DNA Polymerase EnzymeGenomic DNA is synthesized in...
Viral Mutations00:36

Viral Mutations

A mutation is a change in the sequence of bases of DNA or RNA in a genome. Some mutations occur during replication of the genome due to errors made by the polymerase enzymes that replicate DNA or RNA. Unlike DNA polymerase, RNA polymerase is prone to errors because it is not capable of “proofreading” its work. Viruses with RNA-based genomes, like HIV, therefore accrue mutations faster than viruses with DNA-based genomes. Because mutation and recombination provide the raw material for adaptive...
Translesion DNA Polymerases02:10

Translesion DNA Polymerases

Translesion (TLS) polymerases rescue stalled DNA polymerases at sites of damaged bases by replacing the replicative polymerase and installing a nucleotide across the damaged site. Doing so, TLS allows additional time for the cell to repair the damage before resuming regular DNA replication.
TLS polymerases are found in all three domains of life - archaea, bacteria, and eukaryotes. Of the different classes of TLS polymerases, members of the Y family are fitted with specialized structures that...
Leaky Scanning02:28

Leaky Scanning

During most eukaryotic translation processes, the small 40S ribosome subunit scans an mRNA from its 5' end until it encounters the first start AUG codon. The large 60S ribosomal subunit then joins the smaller one to initiate protein synthesis. The location of the translation initiation is largely determined by the nucleotides near the start codon as there may be multiple translation initiation sites present on the mRNA.  Marilyn Kozak discovered that the sequence RCCAUGG (where R stands for...
Proofreading01:31

Proofreading

Synthesis of new DNA molecules is carried out by the enzyme DNA polymerase, which adds nucleotides on the daughter strand complementary to the template DNA strand. DNA polymerase has a higher affinity to add the correct base and ensures fidelity during DNA replication. Furthermore,  it exhibits proofreading activity during replication, using an exonuclease domain that cuts off incorrect nucleotides from the nascent DNA strand.
Errors During Replication are Corrected by the DNA Polymerase Enzyme
Poliomyelitis01:17

Poliomyelitis

Poliomyelitis is caused by poliovirus, a small, non-enveloped, positive-sense RNA virus of the Picornaviridae family and Enterovirus genus. Transmission occurs primarily via the fecal-oral route, often through ingestion of contaminated water or food. The virus initially replicates in the oropharynx and intestinal mucosa, particularly in lymphoid tissues such as the tonsils, Peyer’s patches, and regional lymph nodes. Primary viremia follows, allowing dissemination throughout the body.In most...