title: Integration requires a specific interaction of the donor DNA terminal 5'-cytosine with glutamine 148 of the HIV-1 integrase flexible loop.
abstract: Integration is essential for retroviral replication and gene therapy using retroviral vectors. Human immunodeficiency virus, type 1 (HIV-1), integrase specifically recognizes the terminal sequences of each long terminal repeat (LTR) and cleaves the 3'-end terminal dinucleotide 5'-GT. The exposed 3'-hydroxyl is then positioned for nucleophilic attack and subsequent strand transfer into another DNA duplex (target or chromosomal DNA). We report that both the terminal cytosine at the protruding 5'-end of the long terminal repeats (5'-C) and the integrase residue Gln-148 are critical for strand transfer. Proximity of the 5'-C and Gln-148 was demonstrated by disulfide cross-linking. Cross-linking is inhibited by the inhibitor 5CITEP 1-(5-chloroindol-3-yl)-3-hydroxy-3-(2H-tetrazol-5-yl)-propenone. We propose that strand transfer requires a conformational change of the integrase-viral (donor) DNA complex with formation of an H-bond between the N-3 of the 5'-C and the amine group of Gln-148. These findings have implications for the molecular mechanisms coupling 3'-processing and strand transfer as well as for the molecular pharmacology of integrase inhibitors.
authors.affiliation: Laboratory of Molecular Pharmacology, Center for Cancer Research, NCI, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Allison A
authors.initials: AA
authors.lastname: Johnson
authors.forename: Webster
authors.initials: W
authors.lastname: Santos
authors.forename: Godwin C G
authors.initials: GC
authors.lastname: Pais
authors.forename: Christophe
authors.initials: C
authors.lastname: Marchand
authors.forename: Ronak
authors.initials: R
authors.lastname: Amin
authors.forename: Terrence R
authors.initials: TR
authors.lastname: Burke
authors.forename: Gregory
authors.initials: G
authors.lastname: Verdine
authors.forename: Yves
authors.initials: Y
authors.lastname: Pommier
mesh_terms: D017124:Conserved Sequence; D003596:Cytosine / Q000737:chemistry; D005973:Glutamine / Q000737:chemistry; D019427:HIV Integrase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016325:HIV Long Terminal Repeat / Q000502:physiology*; D015497:HIV-1 / Q000201:enzymology* / Q000235:genetics* / Q000254:growth & development; D006860:Hydrogen Bonding; D016296:Mutagenesis; D017433:Protein Structure, Secondary; D013379:Substrate Specificity; D016662:Virus Integration; D014779:Virus Replication
chemical_list: D005973:Glutamine; D003596:Cytosine; D019427:HIV Integrase