title: Recombinant production of Epstein-Barr virus BZLF1 trans-activator and characterization of its DNA-binding specificity.
abstract: This paper describes the recombinant production of a biologically active Epstein-Barr virus BZLF1 trans-activator, i.e., Z-encoded broadly reactive activator (ZEBRA), that recognized specific DNA motifs. We used auto-induction for histidine-tagged BZLF1 expression in Escherichia coli and immobilized cobalt affinity membrane chromatography for protein purification under native conditions. We obtained the purified BZLF1 at a yield of 5.4mg per gram of wet weight cells at 75% purity, in which 27% of the recombinant BZLF1 remained biologically active. The recombinant BZLF1 bound to oligonucleotides containing ZEBRA response elements, either AP-1 or ZIIIB, but not a ZIIIB mutant. The recombinant BZLF1 showed a specific DNA-binding activity which could be useful for functional studies.
authors.affiliation: Institute of Biological Sciences, Faculty of Science, University of Malaya, 50603 Kuala Lumpur, Malaysia.
authors.forename: Chun Shen
authors.initials: CS
authors.lastname: Lim
authors.affiliation: Institute of Biological Sciences, Faculty of Science, University of Malaya, 50603 Kuala Lumpur, Malaysia.
authors.forename: Siang Ling
authors.initials: SL
authors.lastname: Goh
authors.affiliation: Department of Otorhinolaryngology, Faculty of Medicine, University of Malaya, 50603 Kuala Lumpur, Malaysia.
authors.forename: Gopala
authors.initials: G
authors.lastname: Krishnan
authors.affiliation: Institute of Biological Sciences, Faculty of Science, University of Malaya, 50603 Kuala Lumpur, Malaysia. Electronic address: ccng@um.edu.my.
authors.forename: Ching Ching
authors.initials: CC
authors.lastname: Ng
mesh_terms: D002846:Chromatography, Affinity; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004926:Escherichia coli; D009841:Oligonucleotides / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D015534:Trans-Activators / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: C063676:BZLF1 protein, Herpesvirus 4, Human; D009841:Oligonucleotides; D011994:Recombinant Proteins; D015534:Trans-Activators; D004247:DNA
keywords: Auto-induction; EBV; IMAC; Membrane adsorber; ZEBRA; ZREs