title: Galactose-limited fed-batch cultivation of Escherichia coli for the production of lacto-N-tetraose.
abstract: Lacto-N-tetraose (Gal(β1-3)GlcNAc(β1-3)Gal(β1-4)Glc) is one of the most abundant oligosaccharide structures in human milk. We recently described the synthesis of lacto-N-tetraose by a whole-cell biotransformation with recombinant Escherichia coli cells. However, only about 5% of the lactose was converted into lacto-N-tetraose by this approach. The major product obtained was the intermediate lacto-N-triose II (GlcNAc(β1-3)Gal(β1-4)Glc). In order to improve the bioconversion of lactose to lacto-N-tetraose, we have investigated the influence of the carbon source on the formation of lacto-N-tetraose and on the intracellular availability of the glycosyltransferase substrates, UDP-N-acetylglucosamine and UDP-galactose. By growth of the recombinant E. coli cells on D-galactose, the yield of lacto-N-tetraose (810.8 mg L(-1) culture) was 3.6-times higher compared to cultivation on D-glucose. Using fed-batch cultivation with galactose as sole energy and carbon source, a large-scale synthesis of lacto-N-tetraose was demonstrated. During the 26 h feeding phase the growth rate (μ = 0.05) was maintained by an exponential galactose feed. In total, 16 g L(-1) lactose were fed and resulted in final yields of 12.72 ± 0.21 g L(-1) lacto-N-tetraose and 13.70 ± 0.10 g L(-1) lacto-N-triose II. In total, 173 g of lacto-N-tetraose were produced with a space-time yield of 0.37 g L(-1) h(-1).
authors.affiliation: Institute of Microbiology, University of Stuttgart, Allmandring 31, 70569 Stuttgart, Germany.
authors.forename: Florian
authors.initials: F
authors.lastname: Baumgärtner
authors.affiliation: Institute of Microbiology, University of Stuttgart, Allmandring 31, 70569 Stuttgart, Germany.
authors.forename: Georg A
authors.initials: GA
authors.lastname: Sprenger
authors.affiliation: Institute of Microbiology, University of Stuttgart, Allmandring 31, 70569 Stuttgart, Germany. Electronic address: imbca@imb.uni-stuttgart.de.
authors.forename: Christoph
authors.initials: C
authors.lastname: Albermann
mesh_terms: D019149:Bioreactors / Q000382:microbiology; D004926:Escherichia coli / Q000378:metabolism*; D005285:Fermentation; D005690:Galactose / Q000378:metabolism*; D006801:Humans; D007218:Industrial Microbiology; D007700:Kinetics; D053858:Metabolic Networks and Pathways; D008895:Milk, Human / Q000737:chemistry; D009844:Oligosaccharides / Q000096:biosynthesis*; D014539:Uridine Diphosphate Sugars / Q000378:metabolism
chemical_list: D009844:Oligosaccharides; D014539:Uridine Diphosphate Sugars; C013084:lacto-N-neotetraose; D005690:Galactose
keywords: Fed-batch cultivation; Human milk oligosaccharides; Lacto-N-tetraose; UDP-d-galactose