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INTERET THERAPEUTIQUE DE LA PROTEINE A20 DES ORTHOPOXVIRUS COMME CIBLE PERTINENTE D'APTAMERES PEPTIDIQUES ET DE COMPOSES CHIMIQUES BLOQUANT SES INTERACTIONS ESSENTIELLES A L'INTERIEUR DU COMPLEXE DE REPLICATION VIRALE

Abstract : Variola virus (VARV), the etiologic agent of smallpox was responsible of the most devastating infectious disease. Because of successful preventive measures by immunization, the World Health Organization (WHO) declared global eradication of smallpox in 1980. The subsequent discontinuation of vaccination has rendered all children and many adults virtually susceptible to smallpox infection. If variola virus was used in an act of terrorism or warfare it could cause a real catastrophe. So it is essential to develop new antiviral molecules with different mechanisms of action and usable immediately in case of terrorist attack. Here we report the use of two original strategies to identify new effective anti-orthopoxvirus agents targeting specifically the viral replication complex of vaccinia virus (VACV), a valuable surrogate for the smallpox virus. First, through a yeast two-hybrid assay, we have selected peptide aptamers directed against the VACV A20 protein, a central component of the replication complex and shown to form, with the D4 protein, a processivity factor for the viral DNA polymerase. Peptide aptamers are combinatorial protein molecules designed to inhibit the function of target proteins in living cells. We have proved that one selected aptamer interacting with a central region of A20 was able to significantly inhibit viral DNA synthesis and viral production in cell culture. Second, we have performed a high-throughput screening of small molecules to isolate compounds capable of disrupting A20 interaction with either D4, an uracil DNA glycosylase or D5, a DNA-independent nucleoside triphosphatase (NTPase) that contains a helicase domain. The screen is based on an automated dual-luciferase yeast two-hybrid assay, performed in 384-well plates. Among a collection of 27,600 compounds from diverse commercial chemical libraries we have identified two potential inhibitors that exhibit antipoxvirus effect on infected cell culture. These compounds were also found to specifically inhibit DNA replication. Thus, the screening for inhibitors of protein-protein interactions within viral replication complex remains to be a promising strategy for identifying new compounds active against orthopoxvirus infections.
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https://tel.archives-ouvertes.fr/tel-00440047
Contributor : Laurent Saccucci <>
Submitted on : Tuesday, June 1, 2010 - 7:00:17 AM
Last modification on : Tuesday, June 1, 2010 - 7:00:17 AM
Long-term archiving on: : Wednesday, November 30, 2016 - 10:05:19 AM

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  • HAL Id : tel-00440047, version 1

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Laurent Saccucci. INTERET THERAPEUTIQUE DE LA PROTEINE A20 DES ORTHOPOXVIRUS COMME CIBLE PERTINENTE D'APTAMERES PEPTIDIQUES ET DE COMPOSES CHIMIQUES BLOQUANT SES INTERACTIONS ESSENTIELLES A L'INTERIEUR DU COMPLEXE DE REPLICATION VIRALE. Sciences du Vivant [q-bio]. Université Joseph-Fourier - Grenoble I, 2009. Français. ⟨tel-00440047⟩

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