Redox Biocatalysis: Quantitative Comparisons of Nicotinamide Cofactor Regeneration Methods

Abstract

Enzymatic processes, particularly those capable of performing redox reactions, have recently been of growing research interest. Substrate specificity, optimal activity at mild temperatures, high selectivity, and yield are among the desirable characteristics of these oxidoreductase catalyzed reactions. Nicotinamide adenine dinucleotide (phosphate) or NAD(P)H‐dependent oxidoreductases have been extensively studied for their potential applications like biosynthesis of chiral organic compounds, construction of biosensors, and pollutant degradation. One of the main challenges associated with making these processes commercially viable is the regeneration of the expensive cofactors required by the enzymes. Numerous efforts have pursued enzymatic regeneration of NAD(P)H by coupling a substrate reduction with a complementary enzyme catalyzed oxidation of a co‐substrate. While offering excellent selectivity and high total turnover numbers, such processes involve complicated downstream product separation of a primary product from the coproducts and impurities. Alternative methods comprising chemical, electrochemical, and photochemical regeneration have been developed with the goal of enhanced efficiency and operational simplicity compared to enzymatic regeneration. Despite the goal, however, the literature rarely offers a meaningful comparison of the total turnover numbers for various regeneration methodologies. This comprehensive Review systematically discusses various methods of NAD(P)H cofactor regeneration and quantitatively compares performance across the numerous methods. Further, fundamental barriers to enhanced cofactor regeneration in the various methods are identified, and future opportunities are highlighted for improving the efficiency and sustainability of commercially viable oxidoreductase processes for practical implementation.

Biocatalytic Furfuryl Alcohol Production with Ethanol as the Terminal Reductant Using a Single Enzyme

“…Such coupled redox strategies have been commonly used in NAD­(P)­H-dependent dehydrogenase enzyme-catalyzed reactions based on single or multi-enzymes, as evident from previous studies. , Nicotinamide cofactors are expensive; hence, it is essential to optimize the cofactor concentrations to make the process economically viable. As a recent review indicates, enzymatic regeneration has proven to be the most productive based on turnover numbers …”

Crosslinked Aggregates of Fusion Enzymes in Microaqueous Organic Media

“…Alternatively, these multienzymatic cascades can be classified into four modes, i) linear, ii) orthogonal, iii) parallel, and iv) cyclic [23] . The most straightforward design is the linear cascade, where a single substrate is converted to a single product via an intermediate [24–26] . This kind of cascade largely promotes efficient and safe processes by circumventing the downstream processing, transforming intermediate spontaneously, and driving reversible reactions to completion [27–29] .…”

“…[23] The most straightforward design is the linear cascade, where a single substrate is converted to a single product via an intermediate. [24][25][26] This kind of cascade largely promotes efficient and safe processes by circumventing the downstream processing, transforming intermediate spontaneously, and driving reversible reactions to completion. [27][28][29] Consequently, linear cascades have been established for various enzymes in aqueous systems including oxygenases.…”

Enzymatic Electrosynthesis of Glycine from CO2 and NH3

“…However, to deal with energy supply issue in vitro, efficient reductive regeneration of NAD(P)H from NAD(P) + is needed. Recently, alternative methods including chemical, [14] photochemical, [15] and electrochemical [16] regeneration have been developed, providing easier downstream operation and improving catalytic yield as compared to enzymatic regeneration [17] …”

“…Recently, alternative methods including chemical, [14] photochemical, [15] and electrochemical [16] regeneration have been developed, providing easier downstream operation and improving catalytic yield as compared to enzymatic regeneration. [17] Inspired by the reductive glycine pathway, in this study we constructed an enzymatic electrocatalytic system for the one-pot synthesis of glycine, the smallest amino acid, by the simultaneous utilization of CO 2 and NH 3 . The NAD(P)H required in the system can be efficiently regenerated by a Cu foam electrode as reported previously by our group.…”