Abstract: This study aimed to explore the feasibility of synthesizing trehalose using cellobiose as substrate. Firstly, the otsA/otsB gene from Pseudomonas stutzeri A1501 was cloned and exogenously introduced into Escherichia coli BL21(DE3) to construct an OtsAB pathway to synthesize trehalose using glucose as the substrate. Subsequently, the content of uridine diphosphate (UDP)-glucose, the key precursor for biosynthesis of trehalose was increased by overexpressing the galU gene of E. coli itself, resulting in a 4-fold increase in trehalose production. By adding validamycin, the degradation of trehalose was suppressed to increase its yield. Furthermore, by overexpressing the cellobiose phosphorylase gene (cepA) from the naturally occurring cellulolytic bacterium Saccharophagus degradans, the recombinant E. coli was rendered capable of producing trehalose from cellobiose. After 48 h culture with the addition of 0.05 mmol/L validamycin against trehalose degradation, whole recombinant cells catalyzed the transformation of 20 g/L cellobiose to 1.3 g/L trehalose. This study confirms the feasibility of synthesizing trehalose using cellobiose as substrate and provides new ideas for producing fine chemicals from cellobiose.

Key words: cellobiose, cellobiose phosphorylase, trehalose, UDP-glucose, OtsAB pathway