Issue 3, Volume 3 – 1 articles

Rapid Production of High-Titer d-Mannitol and Gluconate Catalyzed by a Combination of Whole-Cell and an Enzyme at High Temperatures

d-Mannitol and d-gluconate are value-added biobased chemicals with diverse applications in food, medical, and chemical industries. d-Mannitol can be hydrogenated from hexoses (e.g., d-fructose) catalyzed by microbial fermentation, whole-cell biocatalysis, and purified-enzyme cascade biocatalysis. Here we designed a cell–enzyme system comprised of the whole cells co-expressing both hyperthermophilic mannitol dehydrogenase (MDH) and glucose dehydrogenase (GDH) as well as a hyperthermophilic xylose isomerase (XI). The whole cells have its inherent NAD enabled to implement NAD-self sufficient coupled redox reactions without externally-added NAD and aeration. Four cases of whole cells co-expressed MDH and GDH in E. coli BL21(DE3) were compared and optimized by expressing two genes separately or in tandem and changing gene alignment. Also, two-step biotransformation was developed to convert 300 g/L glucose to 129 g/L mannitol and 161 g/L gluconate in a pH-controlled bioreactor at 70 °C. This cell–enzyme system had a high volumetric productivity (10.7 g/L/h mannitol and 13.4 g/L/h gluconate) and a high product yield (91.7%). This study implied that using hyperthermophilic enzymes and cell–enzyme system could open great opportunities for industrial biomanufacturing.