Table 2. Kinetic Parameters Determined for the NADH-tNAD+ Transhydrogenase Reaction Catalyzed by Mtb NDH-2 and Inhibition Constants Produced by Addition of NAD+ to Each Reaction Mixturea.
| NADH-UQ0 reductase reaction | NADH-tNAD+ transhydrogenase reaction | |
|---|---|---|
| KMNADH (mM) | 0.13–0.15b | 0.01 ± 0.001c |
| KMtNAD+ (mM) | – | 0.02 ± 0.003c |
| KitNAD+ (mM) | 1.0 ± 0.13d | – |
| KsitNAD+ (mM) | – | 0.90 ± 0.35c |
| Ki′NAD+ (mM) | 0.54, 1.37e | 1.4 ± 0.34 |
| Ki″NAD+ (mM) | – | 0.03–0.05f |
a
Reaction steps represented by constants are noted on Schemes 3–5 in Figures 4 and 5. Values were obtained from global fits using eq 4 or 5 as described in Experimental Procedures. Analyses of NAD+ inhibitory activity were performed with each substrate varied (see Figure 4); hence, two values are reported for these constants.
b
Data taken from Table 1.
c
Fitted parameters obtained from a global fit of data in Figure 3B to a modified eq 3, assuming the kcat value (0.25 s–1) for the transhydrogenase reaction determined from data in Figure 3A.
d
Value obtained from progress curves in Figure 5.
e
Fitted parameter obtained from data in panels C and D of Figure 4.
f
Fitted parameters obtained from data in panels A and B of Figure 4.