title: Increased production of folate by metabolic engineering of Lactococcus lactis.
abstract: The dairy starter bacterium Lactococcus lactis is able to synthesize folate and accumulates large amounts of folate, predominantly in the polyglutamyl form. Only small amounts of the produced folate are released in the extracellular medium. Five genes involved in folate biosynthesis were identified in a folate gene cluster in L. lactis MG1363: folA, folB, folKE, folP, and folC. The gene folKE encodes the biprotein 2-amino-4-hydroxy-6-hydroxymethyldihydropteridine pyrophosphokinase and GTP cyclohydrolase I. The overexpression of folKE in L. lactis was found to increase the extracellular folate production almost 10-fold, while the total folate production increased almost 3-fold. The controlled combined overexpression of folKE and folC, encoding polyglutamyl folate synthetase, increased the retention of folate in the cell. The cloning and overexpression of folA, encoding dihydrofolate reductase, decreased the folate production twofold, suggesting a feedback inhibition of reduced folates on folate biosynthesis.
authors.affiliation: Wageningen Centre for Food Sciences, NIZO Food Research, 6718 ZB Ede, The Netherlands.
authors.forename: Wilbert
authors.initials: W
authors.lastname: Sybesma
authors.forename: Marjo
authors.initials: M
authors.lastname: Starrenburg
authors.forename: Michiel
authors.initials: M
authors.lastname: Kleerebezem
authors.forename: Igor
authors.initials: I
authors.lastname: Mierau
authors.forename: Willem M
authors.initials: WM
authors.lastname: de Vos
authors.forename: Jeroen
authors.initials: J
authors.lastname: Hugenholtz
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D003470:Culture Media; D005492:Folic Acid / Q000096:biosynthesis*; D015964:Gene Expression Regulation, Bacterial; D005818:Genetic Engineering / Q000379:methods*; D013294:Lactococcus lactis / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D005810:Multigene Family; D017422:Sequence Analysis, DNA; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D003470:Culture Media; D005492:Folic Acid; D013762:Tetrahydrofolate Dehydrogenase