Long-Term Stability of Nicotinamide Cofactors in Common Aqueous Buffers: Implications for Cell-Free Biocatalysis
The use of nicotinamide cofactors in cell-free biocatalytic systems is necessitated by the high specificity that these enzymes show for their natural redox mediators. Unfortunately, isolation and use of natural cofactors is costly, which suggests that enhancing their stability is key to enabling their use in industrial processes. This study details NAD+ and NADH stability in three buffer systems (sodium phosphate, HEPES, and Tris) at 19 °C and 25 °C and for up to 43 d. In Tris, both NADH and NAD+ were found to be highly stable. NADH degradation rates of 4 μM/d (19 °C) and 11 μM/d (25 °C) were observed in Tris buffer, corresponding to >90% and 75% remaining after 43 d, respectively. Higher degradation rates (up to 34 μM/d) were observed when sodium phosphate or HEPES buffers were used. The effect of a mild increase in temperature was determined to be significant for long-term stability, and it was shown that degradation under these conditions can be easily monitored via UV–Vis, because the degradation proceeds via the oxidation/de-aromatization of the dihydropyridine ring. Overall, this work emphasizes that the choice of buffer system is consequential for bioreactor systems employing natural nicotinamide cofactors for extended periods of time.
- Research Organization:
- National Renewable Energy Laboratory (NREL), Golden, CO (United States)
- Sponsoring Organization:
- USDOE; USDOE Office of Energy Efficiency and Renewable Energy (EERE), Office of Sustainable Transportation. Bioenergy Technologies Office (BETO)
- Grant/Contract Number:
- AC36-08GO28308
- OSTI ID:
- 2478318
- Report Number(s):
- NREL/JA--2700-90741; PII: molecules29225453
- Journal Information:
- Molecules, Journal Name: Molecules Journal Issue: 22 Vol. 29; ISSN MOLEFW; ISSN 1420-3049
- Publisher:
- MDPI AGCopyright Statement
- Country of Publication:
- Switzerland
- Language:
- English
Similar Records
Computer-Aided Design of Stability Enhanced Nicotinamide Cofactor Biomimetics for Cell-Free Biocatalysis
Experimental and Computational Evaluation of Nicotinamide Cofactor Biomimetics