DMSO tolerant NAD(P)H recycler enzyme from a pathogenic bacterium, Burkholderia dolosa PC543: effect of N-/C-terminal His Tag extension on protein solubility and activity

Abstract

NAD(P)+ dependent formate dehydrogenase (FDH) is an oxidoreductase used as a biocatalyst to regenerate NAD(P)H in reductase-mediated chiral synthesis reactions. Solvent stability and the need to reduce NADP+ to NADPH, due to the high cost of NADPH, are required features in the industrial usage of FDHs. Therefore, we aimed to identify a novel, robust NADP+ dependent FDH and evaluate the effect of N- and C- terminus His tag extensions on protein solubility and activity. Herein, we report a novel, DMSO tolerant formate dehydrogenase (BdFDH), which has dual coenzyme specificity and tolerance to acidic pH, from Burkholderia dolosa PC543. N- and C-terminus His-tagged BdFDHs were expressed separately in Escherichia coli BL21 (DE3). The C-terminal His-tagged BdFDH was soluble and active whereas the N-terminal version was not. The enzyme displays dual coenzyme specificity and resistance to some organic solvents, particularly DMSO, and is able to tolerate acidic pH conditions. The apparent KM values for NADP+, NAD+ and sodium formate (with NADP+), are 1.17, 14.7 and 5.66 mM, respectively. As a result, due to its DMSO tolerance and coenzyme preference, this enzyme can be utilized as an NAD(P)H recycler in several biotransformations particularly when carried out under acidic conditions. Moreover, it can be said that the position of the His tag extension may affect the enzyme solubility and functionality.