title: Human dihydrofolate reductase and thymidylate synthase form a complex in vitro and co-localize in normal and cancer cells.
abstract: Enzymes involved in thymidylate biosynthesis, thymidylate synthase (TS), and dihydrofolate reductase (DHFR) are well-known targets in cancer chemotherapy. In this study, we demonstrated for the first time, that human TS and DHFR form a strong complex in vitro and co-localize in human normal and colon cancer cell cytoplasm and nucleus. Treatment of cancer cells with methotrexate or 5-fluorouracil did not affect the distribution of either enzyme within the cells. However, 5-FU, but not MTX, lowered the presence of DHFR-TS complex in the nucleus by 2.5-fold. The results may suggest the sequestering of TS by FdUMP in the cytoplasm and thereby affecting the translocation of DHFR-TS complex to the nucleus. Providing a strong likelihood of DHFR-TS complex formation in vivo, the latter complex is a potential new drug target in cancer therapy. In this paper, known 3D structures of human TS and human DHFR, and some protozoan bifunctional DHFR-TS structures as templates, are used to build an in silico model of human DHFR-TS complex structure, consisting of one TS dimer and two DHFR monomers. This complex structure may serve as an initial 3D drug target model for prospective inhibitors targeting interfaces between the DHFR and TS enzymes.
authors.affiliation: a Faculty of Chemistry , Warsaw University of Technology , Noakowskiego 3, 00-664 Warsaw , Poland.
authors.forename: Anna
authors.initials: A
authors.lastname: Antosiewicz
authors.affiliation: b Nencki Institute of Experimental Biology , Polish Academy of Sciences , Pasteura 3, 02-093 , Warsaw , Poland.
authors.forename: Adam
authors.initials: A
authors.lastname: Jarmuła
authors.affiliation: b Nencki Institute of Experimental Biology , Polish Academy of Sciences , Pasteura 3, 02-093 , Warsaw , Poland.
authors.forename: Dorota
authors.initials: D
authors.lastname: Przybylska
authors.affiliation: b Nencki Institute of Experimental Biology , Polish Academy of Sciences , Pasteura 3, 02-093 , Warsaw , Poland.
authors.forename: Grażyna
authors.initials: G
authors.lastname: Mosieniak
authors.affiliation: b Nencki Institute of Experimental Biology , Polish Academy of Sciences , Pasteura 3, 02-093 , Warsaw , Poland.
authors.forename: Joanna
authors.initials: J
authors.lastname: Szczepanowska
authors.affiliation: a Faculty of Chemistry , Warsaw University of Technology , Noakowskiego 3, 00-664 Warsaw , Poland.
authors.forename: Anna
authors.initials: A
authors.lastname: Kowalkowska
authors.affiliation: b Nencki Institute of Experimental Biology , Polish Academy of Sciences , Pasteura 3, 02-093 , Warsaw , Poland.
authors.forename: Wojciech
authors.initials: W
authors.lastname: Rode
authors.affiliation: a Faculty of Chemistry , Warsaw University of Technology , Noakowskiego 3, 00-664 Warsaw , Poland.
authors.forename: Joanna
authors.initials: J
authors.lastname: Cieśla
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000737:chemistry* / Q000494:pharmacology; D001665:Binding Sites; D002453:Cell Cycle / Q000187:drug effects / Q000235:genetics; D002460:Cell Line; D018360:Crystallography, X-Ray; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D005472:Fluorouracil / Q000737:chemistry* / Q000494:pharmacology; D045325:HCT116 Cells; D006801:Humans; D007700:Kinetics; D008727:Methotrexate / Q000737:chemistry* / Q000494:pharmacology; D062105:Molecular Docking Simulation; D056004:Molecular Dynamics Simulation; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D000072756:Protein Conformation, alpha-Helical; D000072757:Protein Conformation, beta-Strand; D054730:Protein Interaction Domains and Motifs; D013379:Substrate Specificity; D013762:Tetrahydrofolate Dehydrogenase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013816:Thermodynamics; D013940:Thymidylate Synthase / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D000964:Antimetabolites, Antineoplastic; D009097:Multienzyme Complexes; C056930:thymidylate synthase-dihydrofolate reductase; D004247:DNA; D013762:Tetrahydrofolate Dehydrogenase; D013940:Thymidylate Synthase; D005472:Fluorouracil; D008727:Methotrexate
keywords: molecular dynamics; protein–protein complex model; protein–protein interaction; protein–protein interface; thymidylate synthesis