title: Characterization of the DNA-binding activity of HIV-1 integrase using a filter binding assay.
abstract: Based on the selective binding of proteins and DNA to distinct filter materials a double-layered dot blot radio assay was developed to evaluate the binding of DNA to HIV-1 integrase. In this assay the DNA-binding was found to be independent of Mn2+ concentration, inhibited by concentrations of Mg2+ above 5 mM, abolished by zinc chelation and inhibited by monoclonal antibodies reacting with either the N-terminal or C-terminal regions of integrase. Atomic absorption spectroscopy revealed the molar ratio between integrase and zinc to be close to 1. It is concluded that both the N-terminal and the C-terminal regions of integrase are involved in DNA-binding and that the reported double-layered dot blot radio assay is well suited for further characterization of the integrase.
authors.affiliation: Laboratory of Biotechnology, University of Bergen, Norway.
authors.forename: I R
authors.initials: IR
authors.lastname: Haugan
authors.forename: B M
authors.initials: BM
authors.lastname: Nilsen
authors.forename: S
authors.initials: S
authors.lastname: Worland
authors.forename: L
authors.initials: L
authors.lastname: Olsen
authors.forename: D E
authors.initials: DE
authors.lastname: Helland
mesh_terms: D001483:Base Sequence; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004706:Endodeoxyribonucleases / Q000378:metabolism*; D019427:HIV Integrase; D015497:HIV-1 / Q000201:enzymology*; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D016662:Virus Integration; D015032:Zinc / Q000502:physiology; D016335:Zinc Fingers
chemical_list: D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D009838:Oligodeoxyribonucleotides; D019427:HIV Integrase; D004706:Endodeoxyribonucleases; C058608:endonuclease-integrase; D015032:Zinc