title: A molecular docking strategy identifies Eosin B as a non-active site inhibitor of protozoal bifunctional thymidylate synthase-dihydrofolate reductase.
abstract: Protozoal parasites are unusual in that their thymidylate synthase (TS) and dihydrofolate reductase (DHFR) enzymes exist on a single polypeptide. In an effort to probe the possibility of substrate channeling between the TS and DHFR active sites and to identify inhibitors specific for bifunctional TS-DHFR, we used molecular docking to screen for inhibitors targeting the shallow groove connecting the two active sites. Eosin B is a 100 microm non-active site inhibitor of Leishmania major TS-DHFR identified by molecular docking. Eosin B slows both the TS and DHFR reaction rates. When Arg-283, a key residue to which eosin B is predicted to bind, is mutated to glutamate, however, eosin B only minimally inhibits the TS-DHFR reaction. Additionally, eosin B was found to be a 180 microm inhibitor of Toxoplasma gondii in both biochemical and cell culture assays.
authors.affiliation: Department of Pharmacology, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
authors.forename: Chloé E
authors.initials: CE
authors.lastname: Atreya
authors.forename: Eric F
authors.initials: EF
authors.lastname: Johnson
authors.forename: John J
authors.initials: JJ
authors.lastname: Irwin
authors.forename: Antonia
authors.initials: A
authors.lastname: Dow
authors.forename: Kristen M
authors.initials: KM
authors.lastname: Massimine
authors.forename: Isabelle
authors.initials: I
authors.lastname: Coppens
authors.forename: Valeska
authors.initials: V
authors.lastname: Stempliuk
authors.forename: Stephen
authors.initials: S
authors.lastname: Beverley
authors.forename: Keith A
authors.initials: KA
authors.lastname: Joiner
authors.forename: Brian K
authors.initials: BK
authors.lastname: Shoichet
authors.forename: Karen S
authors.initials: KS
authors.lastname: Anderson
mesh_terms: D000818:Animals; D000981:Antiprotozoal Agents / Q000494:pharmacology*; D001120:Arginine / Q000737:chemistry; D001665:Binding Sites; D016466:CHO Cells; D002851:Chromatography, High Pressure Liquid; D006224:Cricetinae; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D010697:Eosine I Bluish; D005452:Fluoresceins / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D018698:Glutamic Acid / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D020128:Inhibitory Concentration 50; D007700:Kinetics; D018320:Leishmania major / Q000378:metabolism; D008958:Models, Molecular; D009097:Multienzyme Complexes / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D013762:Tetrahydrofolate Dehydrogenase / Q000737:chemistry* / Q000378:metabolism; D013940:Thymidylate Synthase / Q000737:chemistry* / Q000378:metabolism; D013997:Time Factors; D014122:Toxoplasma / Q000378:metabolism
chemical_list: D000981:Antiprotozoal Agents; D004791:Enzyme Inhibitors; D005452:Fluoresceins; D009097:Multienzyme Complexes; C056930:thymidylate synthase-dihydrofolate reductase; D018698:Glutamic Acid; D001120:Arginine; D013762:Tetrahydrofolate Dehydrogenase; D013940:Thymidylate Synthase; D010697:Eosine I Bluish