title: Designing protein dimerizers: the importance of ligand conformational equilibria.
abstract: In an effort to elucidate the role of ligand conformation in induced protein dimerization, we synthesized a flexible methotrexate (MTX) dimer, demonstrated its ability to selectively dimerize Escherichia coli dihydrofolate reductase (DHFR), and evaluated the factors regulating its ability to induce cooperative dimerization. Despite known entropic barriers, bis-MTX proved to possess substantial conformational stability in aqueous solution (-3.8 kcal/mol >/= DeltaG(fold) >/= -4.9 kcal/mol), exerting a dominant influence on the thermodynamics of dimerization. To dimerize DHFR, bis-MTX must shift from a folded to an extended conformation. From this conclusion, the strength of favorable protein-protein interactions in bis-MTX-E. coli DHFR dimers (-3.1 kcal/mol >/= DeltaG(c) >/= -4.2 kcal/mol), and the selectivity of dimerization for E. coli DHFR relative to mouse DHFR (>10(7)) could be determined. The crystal structure of bis-MTX in complex with E. coli DHFR confirms the feasibility of a close-packed dimerization interface and suggests a possible solution conformation for the induced protein dimers. Consequently, the secondary structure of this minimal foldamer regulates its ability to dimerize dihydrofolate reductase in solution, providing insight into the complex energy landscape of induced dimerization.
authors.affiliation: Department of Medicinal Chemistry, College of Pharmacy, University of Minnesota, Minneapolis, Minnesota 55455, USA.
authors.forename: Jonathan C T
authors.initials: JC
authors.lastname: Carlson
authors.forename: Aaron
authors.initials: A
authors.lastname: Kanter
authors.forename: Guruvasuthevan R
authors.initials: GR
authors.lastname: Thuduppathy
authors.forename: Vivian
authors.initials: V
authors.lastname: Cody
authors.forename: Pamela E
authors.initials: PE
authors.lastname: Pineda
authors.forename: R Scott
authors.initials: RS
authors.lastname: McIvor
authors.forename: Carston R
authors.initials: CR
authors.lastname: Wagner
mesh_terms: D000818:Animals; D019281:Dimerization; D004926:Escherichia coli / Q000201:enzymology; D007700:Kinetics; D008024:Ligands; D008727:Methotrexate / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry; D008958:Models, Molecular; D008968:Molecular Conformation; D019906:Nuclear Magnetic Resonance, Biomolecular; D011487:Protein Conformation; D012996:Solutions; D013762:Tetrahydrofolate Dehydrogenase / Q000737:chemistry* / Q000378:metabolism; D013816:Thermodynamics
chemical_list: D008024:Ligands; D012996:Solutions; D013762:Tetrahydrofolate Dehydrogenase; D008727:Methotrexate