title: Effects of mutations in residues near the active site of human immunodeficiency virus type 1 integrase on specific enzyme-substrate interactions.
abstract: The phylogenetically conserved catalytic core domain of human immunodeficiency virus type 1 (HIV-1) integrase contains elements necessary for specific recognition of viral and target DNA features. In order to identify specific amino acids that determine substrate specificity, we mutagenized phylogenetically conserved residues that were located in close proximity to the active-site residues in the crystal structure of the isolated catalytic core domain of HIV-1 integrase. Residues composing the phylogenetically conserved DD(35)E active-site motif were also mutagenized. Purified mutant proteins were evaluated for their ability to recognize the phylogenetically conserved CA/TG base pairs near the viral DNA ends and the unpaired dinucleotide at the 5' end of the viral DNA, using disintegration substrates. Our findings suggest that specificity for the conserved A/T base pair depends on the active-site residue E152. The phenotype of IN(Q148L) suggested that Q148 may be involved in interactions with the 5' dinucleotide of the viral DNA end. The activities of some of the proteins with mutations in residues in close proximity to the active-site aspartic and glutamic acids were salt sensitive, suggesting that these mutations disrupted interactions with DNA.
authors.affiliation: Department of Microbiology and Immunology, Stanford University Medical Center, Stanford, California 94305-5428, USA.
authors.forename: J L
authors.initials: JL
authors.lastname: Gerton
authors.forename: S
authors.initials: S
authors.lastname: Ohgi
authors.forename: M
authors.initials: M
authors.lastname: Olsen
authors.forename: J
authors.initials: J
authors.lastname: DeRisi
authors.forename: P O
authors.initials: PO
authors.lastname: Brown
mesh_terms: D001665:Binding Sites / Q000235:genetics; D019427:HIV Integrase / Q000235:genetics* / Q000378:metabolism*; D015497:HIV-1 / Q000201:enzymology*; D006801:Humans; D009154:Mutation; D013329:Structure-Activity Relationship; D013379:Substrate Specificity / Q000235:genetics
chemical_list: D019427:HIV Integrase