Table 2. Kinetic and inhibitor properties of recombinant human ALDH3B1.
| Substrate | Vmax (nmol NADH/min/mg)a | Km (μM) | Vmax/Kmb |
|---|---|---|---|
| Acetaldehyde | 315 ± 8 | 23300 ± 1500 | 0.01 |
| Malondialdehyde | 3399 ± 377 | 152000 ± 17110 | 0.02 |
| Hexanal | 679 ± 27 | 62 ± 8 | 11.0 |
| Octanal | 1239 ± 59 | 8 ± 2 | 155.0 |
| 4HNE | 653 ± 18 | 52 ± 5 | 12.6 |
| Benzaldehyde | 129 ± 6 | 46 ± 11 | 2.8 |
| p-Nitrophenylacetate Cofactorc | 1319 ± 80 | 3600 ± 250 | 0.4 |
| NAD+ | 2987 ± 218 | 316 ± 54 | 9.5 |
| NADP+ | 5121 ± 740 | 1041 ± 311 | 4.9 |
|
| |||
| Inhibitorc | Concentration | % Uninhibited ALDH3B1 activity | P value |
|
| |||
| None | 100 ± 3 | ||
| Disulfiram | 100 μM | 100 ± 3 | no effect |
| 500 μM | 103 ± 1 | no effect | |
| Cyanamided | 400 μM | 106 ± 4 | no effect |
| 2000 μM | 83 ± 1 | *P<0.05 | |
a
Apparent Vmax and Km values were determined by fitting the data to the Michaelis-Menten equation using SigmaPlot.
b
Vmax/Km represents aldehyde (or esterase, in the case of p-nitrophenylacetate) oxidizing capacity (expressed as nmol NADH produced/min per mg protein per nmol substrate per ml).
c
Cofactor and inhibitor kinetics were determined using octanal as substrate
d
Cyanamide requires metabolic activation; thus, ALDH3B1-infected Sf9 cell lysates as opposed to purified protein were used to determine cyanamide-mediated enzyme inhibition.
Data represent means ± SEM from triplicate experiments.