Mutations in the C-terminal domain of ALSV (Avian Leukemia and Sarcoma Viruses) integrase alter the concerted DNA integration process in vitro

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Abstract

Integrase (IN) is the retroviral enzyme responsible for the integration of the DNA copy of the retroviral genome into the host cell DNA. The C-terminal domain of IN is involved in DNA binding and enzyme multimerization. We previously performed single amino acid substitutions in the C-terminal domain of the avian leukemia and sarcoma viruses (ALSV) IN [Moreau et al. (2002). Arch. Virol. 147, 1761-1778]. Here, we modelled these IN mutants and analysed their ability to mediate concerted DNA integration (in an in vitro assay) as well as to form dimers (by size exclusion chromatography and protein-protein cross-linking). Mutations of residues located at the dimer interface (V239, L240, Y246, V257 and K266) have the greatest effects on the activity of the IN. Among them: (a) the L240A mutation resulted in a decrease of integration efficiency that was concomitant with a decrease of IN dimerization; (b) the V239A, V249A and K266A mutants preferentially mediated non-concerted DNA integration rather than concerted DNA integration although they were found as dimers. Other mutations (V260E and Y246W/ΔC25) highlight the role of the C-terminal domain in the general folding of the enzyme and, hence, on its activity. This study points to the important role of residues at the IN C-terminal domain in the folding and dimerization of the enzyme as well as in the concerted DNA integration of viral DNA ends.

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Moreau, K., Faure, C., Violot, S., Verdier, G., & Ronfort, C. (2003). Mutations in the C-terminal domain of ALSV (Avian Leukemia and Sarcoma Viruses) integrase alter the concerted DNA integration process in vitro. European Journal of Biochemistry, 270(22), 4426–4438. https://doi.org/10.1046/j.1432-1033.2003.03833.x

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