Table 1.

Apparent steady-state parameters of the human d-amino acid oxidase reaction with d-alanine

Sample	kcat, S−1	Km (d-ala), mM	Kd (FAD), μM
Wild type	10.0*	1*	11
Mutant enzyme	1.5*	8*	90
FAD-S-mutant enzyme	2.6	13	n/a

The initial rates were measured at 436 nm and room temperature by using a coupled o-dianisidine/peroxidase assay (see Materials and Methods). The steady-state values that were obtained for the samples preincubated with a 100-fold excess of FAD are indicated by asterisks. Apparent K_d values for flavin binding were determined when either wild-type apoprotein (7 μM) or mutant protein (12 μM) were preincubated with various (1 μM to 2 mM range) concentrations of FAD for 20 min at room temperature followed by the assay. n/a = not applicable.