Dihydrofolate reductase (DHFR) catalyzes the reduction of dihydrofolate (DHF) to tetrahydrofolate (THF) through a hydride transfer from NADPH in all dividing cells. Tetrahydrofolate is essential for DNA synthesis, which makes DHFR a main target in pharmaceutical development. DHFR’s importance for <i>in vitro</i> screening of antifolates as potential therapeutic agents in chemotherapy, rheumatoid arthritis treatment, and antibacterial drugs makes it essential to better define its kinetic behavior in the cell. Biochemical studies have shown that macromolecule concentrations in a cell range from 200-400 g/L, which lead to thermodynamic and kinetic deviations from dilute solutions. <i>In vitro</i> studies have been conducted to simulate crowding effects on human dihydrofolate reductase using bovine serum albumin (BSA), ovalbumin and Ficoll-70 as model crowding agents. DHFR shows remarkable sensitivity to crowding effects as reflected by substantial changes in the enzyme’s steady state kinetics at low concentrations of model crowder.

Effects of macromolecular crowding on the kinetics of human dihydrofolate reductase

255th National Meeting (2018)

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