High-Level Production of the Low-Calorie Sugar Sorbitol by Lactobacillus plantarum through Metabolic Engineering

Abstract

Sorbitol is a low-calorie sugar alcohol that is largely used as an ingredient in the food industry, based on its sweetness and its high solubility. Here, we investigated the capacity of Lactobacillus plantarum, a lactic acid bacterium found in many fermented food products and in the gastrointestinal tract of mammals, to produce sorbitol from fructose-6-phosphate by reverting the sorbitol catabolic pathway in a mutant strain deficient for both l- and d-lactate dehydrogenase activities. The two sorbitol-6-phosphate dehydrogenase (Stl6PDH) genes (srlD1 and srlD2) identified in the genome sequence were constitutively expressed at a high level in this mutant strain. Both Stl6PDH enzymes were shown to be active, and high specific activity could be detected in the overexpressing strains. Using resting cells under pH control with glucose as a substrate, both Stl6PDHs were capable of rerouting the glycolytic flux from fructose-6-phosphate toward sorbitol production with a remarkably high efficiency (61 to 65% glucose conversion), which is close to the maximal theoretical value of 67%. Mannitol production was also detected, albeit at a lower level than the control strain (9 to 13% glucose conversion), indicating competition for fructose-6-phosphate rerouting by natively expressed mannitol-1-phosphate dehydrogenase. By analogy, low levels of this enzyme were detected in both the wild-type and the lactate dehydrogenase-deficient strain backgrounds. After optimization, 25% of sugar conversion into sorbitol was achieved with cells grown under pH control. The role of intracellular NADH pools in the determination of the maximal sorbitol production is discussed.

Sugar alcohols—their role in the modern world of sweeteners: a review

“…However, it can be also produced by electrochemical reduction of dextrose in alkaline conditions [21,22]. Although there are known several industrial processes which are used for the production of sorbitol, only a few microorganisms, including three yeast strains and bacteria such as Zymomonas mobilis and Candida boidini, have been suggested as potential sorbitol producers [17,73,87,[90][91][92].…”

“…As it prevents loss of moisture content, it is used in the production of confectionery, baked goods and chocolate, which have a tendency to become dry or harden during storage. Sorbitol is also used as an important precursor of the vitamin C production, sorbose as well as surfactants [5,87,92]. The worldwide production of sorbitol is estimated to be higher than 500,000 tons/year, and the market is continuously increasing, and about 25 % of that production is used for the synthesis of vitamin C [73,85,87].…”

Control of acute, chronic, and constitutive hyperammonemia by wild-type and genetically engineered Lactobacillus plantarum in rodents

“…[41][42][43] Using this approach, we developed a L. plantarum strain overproducing alanine dehydrogenase and consuming in vitro higher amounts of ammonia than its wild-type counterpart. When given at doses of 10 9 CFU in vivo to mice suffering from acute liver failure with HA, this modified strain had the same ability to decrease blood ammonia, to decrease mortality, and to consume gut ammonia as that of wild-type L. plantarum.…”

Genetically Engineered Lactobacilli for Technological and Functional Food Applications

“…Fed-batch fermentations using whey permeate, a waste product from the dairy industry with high concentration of lactose, resulted in a conversion rate of 9.4% of lactose into sorbitol (De Boeck et al, 2010). L. plantarum has also been metabolically engineered to produce sorbitol by constitutive overexpression of either srlD1 or srlD2 genes that encode S6PDH activities in a mutant strain deficient in LDH activity (Ladero et al, 2007). Using non-growing or growing cells under pH control resulted in a very efficiency conversion rate of about 65% and 25%, respectively, of sugar into sorbitol.…”