Characterization of human herpesvirus 6A/B U94 as ATPase,helicase, exonuclease and DNA-binding proteins |
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Authors: | Frédéric Trempe Annie Gravel Isabelle Dubuc Nina Wallaschek Vanessa Collin Shella Gilbert-Girard Guillaume Morissette Benedikt B Kaufer Louis Flamand |
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Institution: | 1Division of Infectious Disease and Immunity, CHU de Québec Research Center, Quebec city, Quebec G1V 4G2, Canada;2Institut für Virologie, Freie Universität Berlin, Berlin 14163, Germany;3Department of microbiology, infectious disease and immunology, Faculty of Medicine, Université Laval, Quebec city, Québec,G1V 0A6 Canada |
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Abstract: | Human herpesvirus-6A (HHV-6A) and HHV-6B integrate their genomes into the telomeres of human chromosomes, however, the mechanisms leading to integration remain unknown. HHV-6A/B encode a protein that has been proposed to be involved in integration termed U94, an ortholog of adeno-associated virus type 2 (AAV-2) Rep68 integrase. In this report, we addressed whether purified recombinant maltose-binding protein (MBP)-U94 fusion proteins of HHV-6A/B possess biological functions compatible with viral integration. We could demonstrate that MBP-U94 efficiently binds both dsDNA and ssDNA containing telomeric repeats using gel shift assay and surface plasmon resonance. MBP-U94 is also able to hydrolyze adenosine triphosphate (ATP) to ADP, providing the energy for further catalytic activities. In addition, U94 displays a 3′ to 5′ exonuclease activity on dsDNA with a preference for 3′-recessed ends. Once the DNA strand reaches 8–10 nt in length, the enzyme dissociates it from the complementary strand. Lastly, MBP-U94 compromises the integrity of a synthetic telomeric D-loop through exonuclease attack at the 3′ end of the invading strand. The preferential DNA binding of MBP-U94 to telomeric sequences, its ability to hydrolyze ATP and its exonuclease/helicase activities suggest that U94 possesses all functions required for HHV-6A/B chromosomal integration. |
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