Cloning and characterization of two distinct water-forming NADH oxidases from Lactobacillus pentosus for the regeneration of NAD.
Bioprocess and biosystems engineering
confidence
Key findings
Two water-forming NADH oxidases (LpNox1, LpNox2) from Lactobacillus pentosus cloned and characterized; no clinical or biological endpoints reported.
View source on PubMed (PMID 26801669) ↗
- Sample size
- Not applicable
- Population
- Not applicable (in vitro enzyme characterization)
- Dosing
- Not applicable
- Duration
- Not applicable
- Route
- Not applicable
- Blinding
- not_reported
- Controls
- none
- Drug class
- coenzyme
Full abstract
Two uncharacterized nicotinamide adenine dinucleotide (NADH) oxidases (named as LpNox1, LpNox2) from Lactobacillus pentosus ATCC 8041 were cloned and overexpressed in Escherichia coli BL21 (DE3). The sequence analysis revealed that the two enzymes are water-forming Noxs with 64 % and 52 % identity to LbNox from Lactobacillus brevis DSM 20054. The optimal pH and temperature of the purified LpNox1 and LpNox2 were 7.0 and 8.0 and 35 and 40 °C, respectively, with K M of 99.0 μM (LpNox1) and 27.6 μM (LpNox2), and yielding catalytic efficiency k cat/K M of 1.0 and 0.2 μM(-1) s(-1), respectively. Heat inactivation studies revealed that the two enzymes are relatively instable. The application of LpNox1 for the regeneration of NAD(+) was demonstrated by coupling with a glycerol dehydrogenase-catalyzed oxidation of glycerol to 1,3-dihydroxyacetone. The characteristics of the LpNox1 could prove to be of interest in industrial application such as NAD(+) regeneration in dehydrogenase-catalyzed oxidations.