Mutational analyses of the core domain of Avian Leukemia and Sarcoma Viruses integrase: critical residues for concerted integration and multimerization

• Published in: Virology (New York, N.Y.) Vol. 318; no. 2; pp. 566 - 581
• Main Authors: Moreau, Karen, Faure, Claudine, Violot, Sébastien, Gouet, Patrice, Verdier, Gérard, Ronfort, Corinne
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 20.01.2004; Elsevier
• Subjects: Alpharetrovirus - enzymology; Alpharetrovirus - physiology; ALSV; Amino Acid Sequence; Avian sarcoma and leukosis virus; Biochemistry, Molecular Biology; Catalytic Domain - genetics; Concerted integration; Integrase; Integrases - genetics; Integrases - metabolism; Life Sciences; Models, Molecular; Molecular Sequence Data; Mutation; Retrovirus; Virus Integration; Virus Replication; ALSV; Integrase; Retrovirus; Concerted integration
• ISSN: 0042-6822; 1096-0341
• DOI: 10.1016/j.virol.2003.09.037

During replicative cycle of retroviruses, the reverse-transcribed viral DNA is integrated into the cell DNA by the viral integrase (IN) enzyme. The central core domain of IN contains the catalytic site of the enzyme and is involved in binding viral ends and cell DNA as well as dimerization. We previously performed single amino acid substitutions in the core domain of an Avian Leukemia and Sarcoma Virus (ALSV) IN [Arch. Virol. 147 (2002) 1761]. Here, we modeled the resulting IN mutants and analyzed the ability of these mutants to mediate concerted DNA integration in an in vitro assay, and to form dimers by protein–protein cross-linking and size exclusion chromatography. The N197C mutation resulted in the inability of the mutant to perform concerted integration that was concomitant with a loss of IN dimerization. Surprisingly, mutations Q102G and A106V at the dimer interface resulted in mutants with higher efficiencies than the wild-type IN in performing two-ended concerted integration of viral DNA ends. The G139D and A195V mutants had a trend to perform one-ended DNA integration of viral ends instead of two-ended integration. More drastically, the I88L and L135G mutants preferentially mediated nonconcerted DNA integration although the proteins form dimers. Therefore, these mutations may alter the formation of IN complexes of higher molecular size than a dimer that would be required for concerted integration. This study points to the important role of core domain residues in the concerted integration of viral DNA ends as well as in the oligomerization of the enzyme.