Application of a coupled enzyme assay to characterize nicotinamide riboside kinases.
Analytical biochemistry
confidence
Key findings
Describes a coupled enzyme assay to characterize Nrk kinetic properties; no clinical or biological endpoints reported.
View source on PubMed (PMID 19027704) ↗
- Sample size
- Not reported
- Population
- In vitro enzyme assay (human Nrk isoforms)
- Dosing
- Not reported
- Duration
- Not reported
- Route
- Not applicable
- Blinding
- not_reported
- Controls
- none
- Drug class
- coenzyme
Full abstract
The recently identified nicotinamide riboside kinases (Nrks) constitute a distinct pathway of nicotinamide adenine dinucleotide (NAD) biosynthesis. Here we present the combination of an established optical adenosine triphosphatase (ATPase) test, the pyruvate kinase/lactate dehydrogenase system, with the Nrk-catalyzed reaction to determine kinetic properties of these enzymes, in particular affinities for ATP. The assay allows variation of both nucleoside and phosphate donor substrates, thereby providing major advantages for the characterization of these enzymes. We confirm previously established kinetic parameters and identify differences in substrate selectivity between the two human Nrk isoforms. The proposed assay is inexpensive and may be applied for high-throughput screening.