• Journal of Microbiology and Biotechnology
• /
• 제31권1호
• /
• pp.154-162
• /
• 2021

L-5-methyltetrahydrofolate (5-MTHF) is one of the biological active forms of folate, which is widely used as a nutraceutical. However, low yield and serious pollution associated with the chemical synthesis of 5-MTHF hampers its sustainable supply. In this study, 5-MTHF production was improved by engineering the 5-MTHF biosynthesis pathway and NADPH supply in Lactococcus lactis for developing a green and sustainable biosynthesis approach. Specifically, overexpressing the key rate-limiting enzyme methylenetetrahydrofolate reductase led to intracellular 5-MTHF accumulation, reaching 18 ㎍/l. Next, 5-MTHF synthesis was further enhanced by combinatorial overexpression of 5-MTHF synthesis pathway enzymes with methylenetetrahydrofolate reductase, resulting in 1.7-fold enhancement. The folate supply pathway was strengthened by expressing folE encoding GTP cyclohydrolase I, which increased 5-MTHF production 2.4-fold to 72 ㎍/l. Furthermore, glucose-6-phosphate dehydrogenase was overexpressed to improve the redox cofactor NADPH supply for 5-MTHF biosynthesis, which led to a 60% increase in intracellular NADPH and a 35% increase in 5-MTHF production (97 ㎍/l). To reduce formation of the by-product 5-formyltetrahydrofolate, overexpression of 5-formyltetrahydrofolate cyclo-ligase converted 5-formyltetrahydrofolate to 5,10-methyltetrahydrofolate, which enhanced the 5-MTHF titer to 132 ㎍/l. Finally, combinatorial addition of folate precursors to the fermentation medium boosted 5-MTHF production, reaching 300 ㎍/l. To the best of our knowledge, this titer is the highest achieved by L. lactis. This study lays the foundation for further engineering of L. lactis for efficient 5-MTHF biosynthesis.

• 한국축산식품학회지
• /
• 제34권1호
• /
• pp.65-72
• /
• 2014

This study is executed to develop probiotics which produce S-adenosyl-L-methionine (SAM), a methyl group donor of the 5-methyltetrahydrofolate methylation reaction within the animal cell. SAM is an essential substance for the synthesis, activation, and metabolism of hormones, neurotransmitters, nucleic acids, phospholipids, and cell membranes of animals. The SAM is also known as a nutritional supplement to improve brain functions of the human. In this study, the SAM-producing strains are identified in 18 types of salted fish, and then, the strains with excellent SAM productions are being identified, with 1 strain in the Enterococcus genus and 9 strains in the Bacillus genus. Strains with a large amount of SAM production include the lactic acid bacteria such as En. faecium and En. durans, En. sanguinicola, as well as various strains in the Bacillus genus. The SAM-overproducing strains show antibacterial activities with certain harmful microbes in addition to the weak acid resistances and strong bile resistances, indicating characteristics of probiotics. It is possible that the jeotgal-originated beneficial strains with overproducing SAM can be commercially utilized in order to manufacture SAM enriched foods.