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Home / Equine Research Bank / J Virol / Biochemical characterization of equine herpesvirus type 3-in...
Journal of virology1980; 34(2); 474-483; doi: 10.1128/JVI.34.2.474-483.1980

Biochemical characterization of equine herpesvirus type 3-induced deoxythymidine kinase purified from lytically infected horse embryo dermal fibroblasts.

Abstract: Infection of horse KyED cells with equine herpesvirus type 3 (EHV-3) resulted in a sevenfold increase in cytosol deoxythymidine kinase (dTK) activity. The EHV-3 dTK was purified from KyED cytosol dTK by affinity chromatography on deoxythymidine-Sepharose and characterized with respect to its electrophoretic mobility, molecular weight, substrate specificity, phosphate donor specificity, and immunological specificity. The purified EHV-3 dTK migrated in polyacrylamide gels with an Rf of 0.30 and sedimented in glycerol gradients with an S value of 5.13, corresponding to a molecular weight of 83,000. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis yielded a single band with a molecular weight of 38,000 to 40,000. Antiserum prepared against the EHV-3 dTK induced in KyED cells neutralized the EHV-3-induced enzyme activity but not the dTK purified from uninfected cells. EHV-3 dTK was less sensitive to feedback inhibition to dTTP and had a lower Ki for the antiviral compound 1-beta-D-arabinofuranyosylthymine and a lower Km for the substrate deoxythymidine. These results indicate that infection of cells with EHV-3 results in the induction of a new virus-coded dTK activity which meets the criteria of Jensen for an evolutionary primitive enzyme.
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Publication Date: 1980-05-01 PubMed ID: 7373717PubMed Central: PMC288726DOI: 10.1128/JVI.34.2.474-483.1980Google Scholar: Lookup
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  • U.S. Gov't
  • P.H.S.

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

This research investigates how infection by equine herpesvirus type 3 (EHV-3) in horse cells leads to a significant increase in a particular enzyme activity, resulting in the production of a new virus-coded enzyme. This enzyme exhibits unique characteristics which may convey evolutionary advantages to the virus.

About the Research

The primary focus of this research was on a specific enzyme known as deoxythymidine kinase (dTK) that is produced in horse cells upon infection by Equine Herpesvirus 3 (EHV-3).

  • The scientists monitored the activity of this enzyme in cells infected with EHV-3 and found that its activity increased sevenfold.
  • The enzyme was then purified from the cells and subjected to detailed analysis to understand its characteristics that could potentially add an evolutionary advantage to the virus.

Procedure

The researchers used sophisticated analysis methods to study the properties of the EHV-3 dTK enzyme.

  • For the purification of the EHV-3 dTK from the cells, affinity chromatography was used. This involves a biochemical technique to separate and purify a specific molecule, the dTK, from a mix of molecules, in this case, the cytosol from infected horse cells.
  • After the enzyme was purified, its molecular structure, weight, substrate specificity, immunological specificity and other properties were examined.
  • Electrophoretic mobility study, conducted using polyacrylamide gels, and sedimentation analysis in glycerol gradients were used to determine the approximate molecular weight of the enzyme.

Results

The results of the study bring forward various unique characteristics of the EHV-3 dTK enzyme.

  • It was found that the molecular weight of the enzyme was between 38,000 and 40,000, which was determined by Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE).
  • The antiserum prepared against EHV-3 dTK was able to neutralize the dTK induced enzyme activity, but not the dTK from uninfected cells. This indicates that this particular dTK is a result of EHV-3 infection.
  • The enzyme was also found to be less susceptible to feedback inhibition, had a lower Ki for the antiviral compound, and a lower Km for the substrate deoxythymidine, indicating its evolutionary advantage.

Conclusion

The results of this research indicate that EHV-3 infection results in a new virus-coded dTK activity which displays characteristics of an evolutionary primitive enzyme. Understanding this enzyme can help reveal further insights about the virus, its infection process, and potential ways to counter it.

Cite This Article

APA
McGowan JJ, Allen GP, Barnett JM, Gentry GA. (1980). Biochemical characterization of equine herpesvirus type 3-induced deoxythymidine kinase purified from lytically infected horse embryo dermal fibroblasts. J Virol, 34(2), 474-483. https://doi.org/10.1128/JVI.34.2.474-483.1980

Publication

Journal of virology
ISSN: 0022-538X
NlmUniqueID: 0113724
Country: United States
Language: English
Volume: 34
Issue: 2
Pages: 474-483

Researcher Affiliations

McGowan, J J
    Allen, G P
      Barnett, J M
        Gentry, G A

          MeSH Terms

          • Animals
          • Cations, Divalent / pharmacology
          • Cell Line
          • Enzyme Induction
          • Fibroblasts
          • Herpesviridae / enzymology
          • Herpesviridae / growth & development
          • Horses
          • Kinetics
          • Molecular Weight
          • Nucleotides / metabolism
          • Substrate Specificity
          • Thymidine Kinase / isolation & purification
          • Thymidine Kinase / metabolism

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          Citations

          This article has been cited 3 times.
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