Formate dehydrogenase from Rhodococcus jostii (RjFDH) – A high‐performance tool for NADH regeneration

Research output: Contribution to journalResearch articleContributedpeer-review



Abstract. The use of formate dehydrogenases (FDHs) for regeneration of the important cofactor NADH in enzyme-catalysed synthetic reactions has several advantages over alternative systems. However, a major bottleneck for broad industrial applications is the low specific activity of the currently used FDHs. In this study, we introduce a novel NAD-dependent formate dehydrogenase from Rhodococcus jostii (RjFDH) with both high specific activity and stability. The enzyme was identified in a targeted database research and recombinantly obtained from Escherichia coli. RjFDH is a homodimer with a monomeric molecular mass of 44.7 kDa. The homology model shows that all amino acid residues of the NAD-dependent formate dehydrogenases are usually concerned with catalytic activity, substrate acceptance, and cofactor binding. The only substrate oxidised by these enzymes is formate. RjFDH had a specific activity of 19.9 U mg-1 at 22 °C along with unimpaired activity and high stability over a broad pH range. The Km values for formate and NAD+ were 7.3 and 0.098 mmol L-1, respectively.
The optimum temperature was found to be 50 °C, at which the enzyme activity increased to about 318%. Both activity and thermal stability were higher than those of the FDH from Candida boidinii (CbFDH), which is the standard enzyme currently in use for cofactor regeneration. Different solvents roughly had the same impact on the activity and stability of both RjFDH and CbFDH. The superior performance of RjFDH over CbFDH as a regeneration system for NADH was demonstrated for the synthesis of L-tert-leucine as well as (S)-1-phenylethanol. In both systems, the concentration of RjFDH used was only one-third of the concentration of CbFDH required to achieve comparable conversion rates. Rational designing provided a promising NADP-accepting variant. Thus, RjFDH has a great potential to serve as an alternative system for NADH regeneration in enzyme-catalysed synthetic reactions.


Original languageEnglish
Pages (from-to)4109-4118
JournalAdvanced synthesis & catalysis
Issue number362
Publication statusPublished - 2020

External IDs

Scopus 85088592429
ORCID /0000-0002-2912-546X/work/142238748