To achieve high mannitol production by Lactococcus lactis, the mannitol 1-phosphatase gene of Eimeria tenella and the mannitol 1-phosphate dehydrogenase gene mtlD of Lactobacillus plantarum were cloned in the nisin-dependent L. lactis NICE overexpression system. As predicted by a kinetic L. lactis glycolysis model, increase in mannitol 1-phosphate dehydrogenase and mannitol 1-phosphatase activities resulted in increased mannitol production. Overexpression of both genes in growing cells resulted in glucose-mannitol conversions of 11, 21, and 27% by the L. lactis parental strain, a strain with reduced phosphofructokinase activity, and a lactate dehydrogenase-deficient strain, respectively. Improved induction conditions and increased substrate concentrations resulted in an even higher glucose-to-mannitol conversion of 50% by the lactate dehydrogenase-deficient L. lactis strain, close to the theoretical mannitol yield of 67%. Moreover, a clear correlation between mannitol 1-phosphatase activity and mannitol production was shown, demonstrating the usefulness of this metabolic engineering approach.
- controlled gene-expression
- complete genome sequence
- acid bacteria
Wisselink, H. W., Moers, A. P. H. A., Mars, A. E., Hoefnagel, M. H. N., de Vos, W. M., & Hugenholtz, J. (2005). Overproduction of heterologous mannitol 1-phosphatase : a key factor for engineering mannitol production by Lactococcus lactis. Applied and Environmental Microbiology, 71(3), 1507-1514. https://doi.org/10.1128/AEM.71.3.1507-1514.2005