Table 2. Dissociation constants of DHFR inhibitors measured by SNAP-based TR-FRET competition assay.

	Enzymatic assay IC50 (nM)WT	Kd (nM)WT	Kd (nM)L54I	Kd (nM)L54G
MTX	0.071±0.007	0.040±0.004	0.13±0.01	6.9±0.4
Trimetrexate	0.27±0.03	0.15±0.01	0.10±0.01	0.64±0.04
Trimethoprim	0.53±0.09	0.30±0.02	0.10±0.01	0.38±0.03
Pyrimethamine	1900±220	260±19	14±1	39±2
WR99210	54000±16000	12000±950	2600±210	15000±790

IC50 and Kd values are the mean±standard error (N = 3 and 4, respectively).