Harnessing Adaptive Evolution to Achieve Superior Mannitol Production by Using Its Native Metabolism.

Mannitol can be obtained as a by-product of certain heterolactic lactic acid bacteria, when grown on substrates containing fructose. , a homolactic lactic acid bacterium, normally does not form mannitol but can be persuaded into doing so by expressing certain foreign enzyme activities. In this study, we find that has an inherent capacity to form mannitol from glucose. By adaptively evolving or derivatives blocked in NAD regenerating pathways, we manage to accelerate growth on mannitol. When cells of the adapted strains are resuspended in buffer containing glucose, 4-58% of the glucose metabolized is converted into mannitol, in contrast to nonadapted strains. The highest conversion was obtained for a strain lacking all major NAD regenerating pathways. Mannitol had an inhibitory effect on the conversion, which we speculated was due to the mannitol uptake system. After its inactivation, 60% of the glucose was converted into mannitol by cells suspended in glucose buffer. Using a two-stage setup, where biomass first was accumulated by aerated culturing, followed by a nonaerated phase (static conditions), it was possible to obtain 6.1 g/L mannitol, where 60% of the glucose had been converted into mannitol, which is the highest yield reported for .