title: Mouse interferon regulatory factor-2: expression, purification and DNA binding activity.
abstract: Interferon regulatory factor-2 (IRF-2) is a mammalian transcription factor for Interferon and Interferon inducible genes; biologically, plays an important role in cell growth regulation and has been shown to be a potential oncogene. We have expressed, purified recombinant Murine IRF-2 (349 amino acid) as a GST (Glutathione-S-Transferase)-IRF-2 soluble fusion protein in E. coli XL-1 blue cells. Recombinant GST-IRF-2 was biologically active in terms of its DNA binding activity with IRF-E oligo (GAAAGT)4. GST-alone expressed from the vector did not bind to it. We observed five different molecular mass complexes of GST-IRF-2/DNA (1-5) with IRF-E, which were competed out by 100×-fold molar excess of IRF-E, suggesting that the complexes were specific for IRF-2. Such GAAANN (N=any nucleotide) hexa nucleotides occur in the promoters of many virus and interferon-inducible mammalian genes. Multimeric GAAAGT/C sequences are inducible by virus, IFN, dsRNA and IRF-1/2. Multiple GST-IRF-2/DNA complexes may be helpful to understand the mechanism of DNA binding activity of IRF-2.
authors.affiliation: Molecular Biology Laboratory, School of Life Sciences, Jawaharlal Nehru University, New Delhi, 110067, India. kpmolbio@gmail.com
authors.forename: Krishna
authors.initials: K
authors.lastname: Prakash
authors.forename: Pramod C
authors.initials: PC
authors.lastname: Rath
mesh_terms: D000818:Animals; D015153:Blotting, Western; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D024202:Electrophoretic Mobility Shift Assay; D004926:Escherichia coli; D040461:Gene Components; D005822:Genetic Vectors / Q000235:genetics; D005982:Glutathione Transferase / Q000378:metabolism*; D050836:Interferon Regulatory Factor-2 / Q000235:genetics* / Q000378:metabolism*; D051379:Mice; D009841:Oligonucleotides / Q000235:genetics; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D012394:Rosaniline Dyes
chemical_list: D017931:DNA Primers; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D050836:Interferon Regulatory Factor-2; D009841:Oligonucleotides; D011993:Recombinant Fusion Proteins; D012394:Rosaniline Dyes; D005982:Glutathione Transferase; C004692:coomassie Brilliant Blue