Overexpression and purification of enzymatically active recombinant integrase protein of rous sarcoma virus

Ilona Marczinovits, J. Molnár, J. Sóki, István Fodor

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

The carboxy-terminal domain of polymerase gene of Rous sarcoma virus was cloned into an expression vector under the control of lac regulatory elements, resulting in the plasmid pMF1413. Upon isopropyl-β-D-thiogalactopyranoside induction, viral integration (IN) protein was expressed in large quantity in Escherichia coli. The expressed recombinant protein was prepurified by successive washing of the bacterial pellet with 0.1 M NaCl and detergents. Further purification was performed in high yield by standard chromatography methods. The purified enzyme revealed selective DNA cleaving activity on supercoiled plasmid with the LTR-LTR junction fragment. The reaction was metal ion dependent, with a preference for Mn2+ over Mg2+, and showed substrate specificity at 1 mM MnCl2.

Original languageEnglish
Pages (from-to)301-306
Number of pages6
JournalVirus Genes
Volume6
Issue number3
DOIs
Publication statusPublished - Aug 1992

Fingerprint

Rous sarcoma virus
Integrases
Recombinant Proteins
Plasmids
Thiogalactosides
Virus Integration
Viral Proteins
Substrate Specificity
Detergents
Chromatography
Metals
Ions
Escherichia coli
DNA
Enzymes
Genes
manganese chloride

Keywords

  • enzyme purification
  • gene expression
  • integrase protein
  • molecular cloning
  • Rous sarcoma virus

ASJC Scopus subject areas

  • Virology
  • Immunology
  • Applied Microbiology and Biotechnology
  • Molecular Biology
  • Genetics

Cite this

Overexpression and purification of enzymatically active recombinant integrase protein of rous sarcoma virus. / Marczinovits, Ilona; Molnár, J.; Sóki, J.; Fodor, István.

In: Virus Genes, Vol. 6, No. 3, 08.1992, p. 301-306.

Research output: Contribution to journalArticle

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