title: A dynamic checkpoint in oxidative lesion discrimination by formamidopyrimidine-DNA glycosylase.
abstract: In contrast to proteins recognizing small-molecule ligands, DNA-dependent enzymes cannot rely solely on interactions in the substrate-binding centre to achieve their exquisite specificity. It is widely believed that substrate recognition by such enzymes involves a series of conformational changes in the enzyme-DNA complex with sequential gates favoring cognate DNA and rejecting nonsubstrates. However, direct evidence for such mechanism is limited to a few systems. We report that discrimination between the oxidative DNA lesion, 8-oxoguanine (oxoG) and its normal counterpart, guanine, by the repair enzyme, formamidopyrimidine-DNA glycosylase (Fpg), likely involves multiple gates. Fpg uses an aromatic wedge to open the Watson-Crick base pair and everts the lesion into its active site. We used molecular dynamics simulations to explore the eversion free energy landscapes of oxoG and G by Fpg, focusing on structural and energetic details of oxoG recognition. The resulting energy profiles, supported by biochemical analysis of site-directed mutants disturbing the interactions along the proposed path, show that Fpg selectively facilitates eversion of oxoG by stabilizing several intermediate states, helping the rapidly sliding enzyme avoid full extrusion of every encountered base for interrogation. Lesion recognition through multiple gating intermediates may be a common theme in DNA repair enzymes.
authors.affiliation: Department of Chemistry, Stony Brook University, Stony Brook, NY 11794, USA.
authors.forename: Haoquan
authors.initials: H
authors.lastname: Li
authors.affiliation: SB RAS Institute of Chemical Biology and Fundamental Medicine, 8 Lavrentieva Ave., Novosibirsk 630090, Russia Novosibirsk State University, 2 Pirogova St., Novosibirsk 630090, Russia.
authors.forename: Anton V
authors.initials: AV
authors.lastname: Endutkin
authors.affiliation: Department of Chemistry, Stony Brook University, Stony Brook, NY 11794, USA.
authors.forename: Christina
authors.initials: C
authors.lastname: Bergonzo
authors.affiliation: Department of Chemistry, Stony Brook University, Stony Brook, NY 11794, USA.
authors.forename: Arthur J
authors.initials: AJ
authors.lastname: Campbell
authors.affiliation: Department of Pharmacological Sciences, Stony Brook University, Stony Brook, NY 11794, USA.
authors.forename: Carlos
authors.initials: C
authors.lastname: de los Santos
authors.affiliation: Department of Pharmacological Sciences, Stony Brook University, Stony Brook, NY 11794, USA.
authors.forename: Arthur
authors.initials: A
authors.lastname: Grollman
authors.affiliation: SB RAS Institute of Chemical Biology and Fundamental Medicine, 8 Lavrentieva Ave., Novosibirsk 630090, Russia Novosibirsk State University, 2 Pirogova St., Novosibirsk 630090, Russia dzharkov@niboch.nsc.ru.
authors.forename: Dmitry O
authors.initials: DO
authors.lastname: Zharkov
authors.affiliation: Department of Chemistry, Stony Brook University, Stony Brook, NY 11794, USA Department of Chemistry, Stony Brook University, Stony Brook, NY 11794, USA carlos.simmerling@stonybrook.edu.
authors.forename: Carlos
authors.initials: C
authors.lastname: Simmerling
mesh_terms: D001120:Arginine / Q000737:chemistry / Q000378:metabolism; D020134:Catalytic Domain; D003596:Cytosine / Q000737:chemistry / Q000378:metabolism; D043382:DNA-Formamidopyrimidine Glycosylase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001411:Geobacillus stearothermophilus / Q000737:chemistry; D006147:Guanine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D009154:Mutation; D011487:Protein Conformation; D013379:Substrate Specificity
chemical_list: D029968:Escherichia coli Proteins; C024829:8-hydroxyguanine; D006147:Guanine; D003596:Cytosine; D001120:Arginine; D043382:DNA-Formamidopyrimidine Glycosylase; C468610:DNA-formamidopyrimidine glycosylase, E coli