Analysis of conserved and non-conserved amino acids critical for ALSV (Avian leukemia and sarcoma viruses) integrase functions in vitro.
Retroviral integrase (IN) is the viral enzyme responsible for the integration of viral DNA into host cellular DNA. In vitro, recombinant IN protein is able to catalyze the 3'-processing, strand transfer and disintegration activities. In order to analyze the importance of specific residues of ALSV (Avian leukemia and sarcoma viruses) IN protein, we introduced 31 amino acid substitutions either in residues previously shown by others to be involved in IN oligomerization or in selected conserved and non-conserved residues through the IN sequence. We tested, in vitro, the three catalytic activities of these mutants as well as their capacity to bind DNA. We found that (i) 88% of the substitutions occurring on well-conserved residues have an effect on IN activities (ii) two mutants (S85T in the central catalytic domain and N197C in the C-terminal domain) present a reduced efficiency of DNA binding compared to the wild type protein. Moreover, all mutations made on the dimer interface of C-terminal domain present reduced activities, suggesting an important role of this part of the protein. Finally, for some mutations, we observed differences between the ALSV and HIV (Human immunodeficiency virus) IN corresponding residues.
Moreau
K
K
Centre de Génétique Moléculaire et Cellulaire, Institut National de la Recherche Agronomique, Université Claude Bernard, Villeurbanne, France.
Faure
C
C
Verdier
G
G
Ronfort
C
C
D001355:Alpharetrovirus / Q000201:enzymology*; D000595:Amino Acid Sequence; D017124:Conserved Sequence; D004247:DNA / Q000378:metabolism; D019427:HIV Integrase / Q000737:chemistry; D019426:Integrases / Q000737:chemistry* / Q000502:physiology; D008969:Molecular Sequence Data; D009154:Mutation; D012995:Solubility; D013329:Structure-Activity Relationship
D004247:DNA; D019427:HIV Integrase; D019426:Integrases