The bifunctional fusion protein systems consisting of Rhodococcus erythropolis chiral alcohol dehydrogenase (READH)

Candida boidinii formate dehydrogenase (CbFDH) or maltose binding protein (MBP) were constructed to regenerate the cofactors for biocatalysis.READH originated from Rhodococcus erythropolis is an (S)-specific nicotinamide adenine dinucleotide (NADH)dependent alcohol dehydrogenase

meanwhile

CbFDH originated from Candida boidinii is an NADH-dependent formate dehydrogenaseThe strategies of the different fusion protein systems included: (1) fusion of the N terminus of READH to the C terminus of MBP

(2) fusion of the N terminus of CbFDH to the C terminus of MBP

(3) fusion of the N terminus of READH to the C terminus of CbFDH

(4) fusion of the C terminus of READH to the N terminus of CbFDHThe activities of READHs were depressed in all fusion strategiesWhen the N terminus of READH was fused to the C terminus of CbFDH

READH reached the highest activity

but CbFDH had no activity.In contrast

when the C terminus of READH was fused to the N terminus of CbFDH

CbFDH showed the highest activity

and both moieties displayed activities.From this study

the authors suggest that the rational design of the bifunctional fusion protein system may improve the biocatalysis efficiency by the simultaneous cofactor regeneration.