Modeling Dihydrofolate Reductase

Abstract

Dihydrofolate reductase (DHFR) is a ubiquitous enzyme that catalyzes the reduction of dihydrofolate (DHF) to tetrahydrofolate (THF). THF is crucial for purine and thymidylate synthesis, precursors to DNA and RNA. Inhibition of DHFR leads to cessation of cell proliferation because the cell is unable to build DNA and RNA. DHFR is a small protein with an α/β structure, consisting of a central eight‐stranded beta sheet surrounded by four alpha helices. The protein has two subdomains, the adenosine binding subdomain (residues 36–108), the binding site for the adenosine moiety of the cofactor, and the loop subdomain, which consists of three loops, the most important being the Met20 loop (residues 9–24). The active site is in the hydrophobic cleft between the two subdomains. The Met20 loop lies above the active site and protects substrates from the solvent. The Met 20 loop assumes two main conformations (“closed” and “occluded”) correlated with the five distinct intermediates of the DHFR catalytic cycle. It is “closed” in the E:NADPH intermediate and the model Michaelis intermediate, E:NADP+:folate. It is “occluded” in the three product complexes, E:NADP+:THF, E:THF and E:NADPH:THF. The Francis Parker SMART Team (Students Modeling A Research Topic) used 3D printing to create models of DHFR in both formations. Supported by a grant from the HHMI Pre‐College Program.