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Designing Protein Dimerizers: The Importance of Ligand Conformational Equilibria
journal contribution
posted on 2003-01-17, 00:00 authored by Jonathan C. T. Carlson, Aaron Kanter, Guruvasuthevan R. Thuduppathy, Vivian Cody, Pamela E. Pineda, R. Scott McIvor, Carston R. WagnerIn 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 ≥ ΔGfold ≥ −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 ≥ ΔGc ≥ −4.2 kcal/mol), and the selectivity of dimerization for E. coli
DHFR relative to mouse DHFR (>107) 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.