title: Targeting flavivirus RNA dependent RNA polymerase through a pyridobenzothiazole inhibitor.
abstract: RNA dependent RNA polymerases (RdRp) are essential enzymes for flavivirus replication. Starting from an in silico docking analysis we identified a pyridobenzothiazole compound, HeE1-2Tyr, able to inhibit West Nile and Dengue RdRps activity in vitro, which proved effective against different flaviviruses in cell culture. Crystallographic data show that HeE1-2Tyr binds between the fingers domain and the priming loop of Dengue virus RdRp (Site 1). Conversely, enzyme kinetics, binding studies and mutational analyses suggest that, during the catalytic cycle and assembly of the RdRp-RNA complex, HeE1-2Tyr might be hosted in a distinct binding site (Site 2). RdRp mutational studies, driven by in silico docking analysis, allowed us to locate the inhibition Site 2 in the thumb domain. Taken together, our results provide innovative concepts for optimization of a new class of anti-flavivirus compounds.
authors.affiliation: Dipartimento di Bioscienze, Università di Milano, Via Celoria 26, I-20133, Milano, Italy; CNR-IBF, Istituto di Biofisica, Via Celoria 26, I-20133, Milano, Italy.
authors.forename: Delia
authors.initials: D
authors.lastname: Tarantino
authors.affiliation: Dipartimento di Scienze Farmaceutiche, Università degli Studi di Perugia, Perugia, Italy.
authors.forename: Rolando
authors.initials: R
authors.lastname: Cannalire
authors.affiliation: Dipartimento di Bioscienze, Università di Milano, Via Celoria 26, I-20133, Milano, Italy; CNR-IBF, Istituto di Biofisica, Via Celoria 26, I-20133, Milano, Italy.
authors.forename: Eloise
authors.initials: E
authors.lastname: Mastrangelo
authors.affiliation: Dipartimento di Bioscienze, Università di Milano, Via Celoria 26, I-20133, Milano, Italy.
authors.forename: Romina
authors.initials: R
authors.lastname: Croci
authors.affiliation: UMR "Emergence des Pathologies Virales" (EPV: Aix-Marseille university - IRD 190 - Inserm 1207 - EHESP), & Fondation IHU Méditerranée Infection, APHM Public Hospitals of Marseille, Marseille, France.
authors.forename: Gilles
authors.initials: G
authors.lastname: Querat
authors.affiliation: Dipartimento di Scienze Farmaceutiche, Università degli Studi di Perugia, Perugia, Italy.
authors.forename: Maria Letizia
authors.initials: ML
authors.lastname: Barreca
authors.affiliation: Dipartimento di Bioscienze, Università di Milano, Via Celoria 26, I-20133, Milano, Italy; CNR-IBF, Istituto di Biofisica, Via Celoria 26, I-20133, Milano, Italy.
authors.forename: Martino
authors.initials: M
authors.lastname: Bolognesi
authors.affiliation: Dipartimento di Scienze Farmaceutiche, Università degli Studi di Perugia, Perugia, Italy. Electronic address: giuseppe.manfroni@unipg.it.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Manfroni
authors.affiliation: Dipartimento di Scienze Farmaceutiche, Università degli Studi di Perugia, Perugia, Italy.
authors.forename: Violetta
authors.initials: V
authors.lastname: Cecchetti
authors.affiliation: Dipartimento di Bioscienze, Università di Milano, Via Celoria 26, I-20133, Milano, Italy; CNR-IBF, Istituto di Biofisica, Via Celoria 26, I-20133, Milano, Italy. Electronic address: mario.milani@unimi.it.
authors.forename: Mario
authors.initials: M
authors.lastname: Milani
mesh_terms: D000998:Antiviral Agents / Q000494:pharmacology; D052160:Benzothiazoles / Q000737:chemistry / Q000494:pharmacology*; D001665:Binding Sites; D020134:Catalytic Domain; D003460:Crystallization; D003716:Dengue Virus / Q000187:drug effects / Q000201:enzymology; D055808:Drug Discovery; D005416:Flavivirus / Q000187:drug effects / Q000201:enzymology*; D007700:Kinetics; D008958:Models, Molecular; D062105:Molecular Docking Simulation; D009154:Mutation; D012324:RNA-Dependent RNA Polymerase / Q000037:antagonists & inhibitors* / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D014902:West Nile virus / Q000187:drug effects / Q000201:enzymology
chemical_list: D000998:Antiviral Agents; D052160:Benzothiazoles; D012324:RNA-Dependent RNA Polymerase
keywords: Crystal structure; Flavivirus; Non-competitive inhibition; Polymerase mutants; RNA dependent RNA polymerase; Virus inhibition