title: Diketo acid inhibitor mechanism and HIV-1 integrase: implications for metal binding in the active site of phosphotransferase enzymes.
abstract: The process of integrating the reverse-transcribed HIV-1 DNA into the host chromosomal DNA is catalyzed by the virally encoded enzyme integrase (IN). Integration requires two metal-dependent reactions, 3' end processing and strand transfer. Compounds that contain a diketo acid moiety have been shown to selectively inhibit the strand transfer reaction of IN in vitro and in infected cells and are effective as inhibitors of HIV-1 replication. To characterize the molecular basis of inhibition, we used functional assays and binding assays to evaluate a series of structurally related analogs. These studies focused on investigating the role of the conserved carboxylate and metal binding. We demonstrate that an acidic moiety such as a carboxylate or isosteric heterocycle is not required for binding to the enzyme complex but is essential for inhibition and confers distinct metal-dependent properties on the inhibitor. Binding requires divalent metal and resistance is metal dependent with active site mutants displaying resistance only when the enzymes are evaluated in the context of Mg(2+). The mechanism of action of these inhibitors is therefore likely a consequence of the interaction between the acid moiety and metal ion(s) in the IN active site, resulting in a functional sequestration of the critical metal cofactor(s). These studies thus have implications for modeling active site inhibitors of IN, designing and evaluating analogs with improved efficacy, and identifying inhibitors of other metal-dependent phosphotransferases.
authors: Department of Biological Chemistry, Merck Research Laboratories, P.O. Box 4, West Point, PA 19486, USA.
authors: Jay A
authors: JA
authors: Grobler
authors: Kara
authors: K
authors: Stillmock
authors: Binghua
authors: B
authors: Hu
authors: Marc
authors: M
authors: Witmer
authors: Peter
authors: P
authors: Felock
authors: Amy S
authors: AS
authors: Espeseth
authors: Abigail
authors: A
authors: Wolfe
authors: Melissa
authors: M
authors: Egbertson
authors: Michele
authors: M
authors: Bourgeois
authors: Jeffrey
authors: J
authors: Melamed
authors: John S
authors: JS
authors: Wai
authors: Steve
authors: S
authors: Young
authors: Joseph
authors: J
authors: Vacca
authors: Daria J
authors: DJ
authors: Hazuda
mesh_terms: D019380:Anti-HIV Agents / Q000737:chemistry* / Q000494:pharmacology; D001667:Binding, Competitive; D019427:HIV Integrase / Q000737:chemistry* / Q000187:drug effects; D019428:HIV Integrase Inhibitors / Q000737:chemistry* / Q000494:pharmacology; D016325:HIV Long Terminal Repeat; D015497:HIV-1 / Q000201:enzymology*; D006801:Humans; D008024:Ligands; D008274:Magnesium; D008345:Manganese; D008956:Models, Chemical; D015394:Molecular Structure; D010770:Phosphotransferases / Q000737:chemistry; D019809:Streptavidin; D013343:Styrenes
chemical_list: D019380:Anti-HIV Agents; D019428:HIV Integrase Inhibitors; D008024:Ligands; D013343:Styrenes; C027142:vinyltoluene; D008345:Manganese; D019809:Streptavidin; D010770:Phosphotransferases; D019427:HIV Integrase; D008274:Magnesium