title: Enzyme catalysed production of sialylated human milk oligosaccharides and galactooligosaccharides by Trypanosoma cruzi trans-sialidase.
abstract: A Trypanosoma cruzi trans-sialidase (E.C. 3.2.1.18) was cloned into Pichia pastoris and expressed. The pH and temperature optimum of the enzyme was determined as pH 5.7 and 30°C. Using casein glycomacropeptide (CGMP) and lactose as sialyl-donor and acceptor respectively, the optimal donor/acceptor ratio for the trans-sialidase catalysed 3'-sialyllactose production was found to be 1:4. Quantitative amounts of 3'-sialyllactose were produced from CGMP and lactose at a yield of 40mg/g CGMP. The 3'-sialyllactose obtained exerted a stimulatory effect on selected probiotic strains, including different Bifidobacterium strains in single culture fermentations. The trans-sialidase also catalysed the transfer of sialic acid from CGMP to galacto-oligosaccharides (GOS) and to the human milk oligosaccharide (HMO) backbone lacto-N-tetraose (LNT) to produce 3'-sialyl-GOS, including doubly sialylated GOS products, and 3'-sialyl-LNT, respectively. This work thus provides proof of the concept of producing 3'-sialyllactose and potentially other sialylated HMOs as well as sialylated GOS enzymatically by trans-sialidase activity, while at the same time providing valorisation of CGMP, a co-processing product from cheese manufacture.
authors: Centre for BioProcess Engineering, Department of Chemical and Biochemical Engineering, Building 229, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.
authors: Jesper
authors: J
authors: Holck
authors: Centre for BioProcess Engineering, Department of Chemical and Biochemical Engineering, Building 229, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.
authors: Dorte M
authors: DM
authors: Larsen
authors: Centre for BioProcess Engineering, Department of Chemical and Biochemical Engineering, Building 229, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.
authors: Malwina
authors: M
authors: Michalak
authors: Department of Biochemistry and Molecular Biology, University of Southern Denmark, 5230 Odense M, Denmark.
authors: Haiying
authors: H
authors: Li
authors: Department of Chemistry, Building 201, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.
authors: Louise
authors: L
authors: Kjærulff
authors: Department of Biochemistry and Molecular Biology, University of Southern Denmark, 5230 Odense M, Denmark.
authors: Finn
authors: F
authors: Kirpekar
authors: Department of Chemistry, Building 201, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.
authors: Charlotte H
authors: CH
authors: Gotfredsen
authors: DuPont Nutrition and Health, Active Nutrition, Sokeritehtaantie 20, 02460 Kantvik, Finland.
authors: Sofia
authors: S
authors: Forssten
authors: DuPont Nutrition and Health, Active Nutrition, Sokeritehtaantie 20, 02460 Kantvik, Finland.
authors: Arthur C
authors: AC
authors: Ouwehand
authors: Centre for BioProcess Engineering, Department of Chemical and Biochemical Engineering, Building 229, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark.
authors: Jørn D
authors: JD
authors: Mikkelsen
authors: Centre for BioProcess Engineering, Department of Chemical and Biochemical Engineering, Building 229, Technical University of Denmark, 2800 Kgs. Lyngby, Denmark. Electronic address: am@kt.dtu.dk.
authors: Anne S
authors: AS
authors: Meyer
mesh_terms: D002364:Caseins / Q000737:chemistry; D005260:Female; D006023:Glycoproteins / Q000737:chemistry* / Q000235:genetics; D006801:Humans; D008895:Milk, Human / Q000737:chemistry*; D019158:N-Acetylneuraminic Acid / Q000737:chemistry*; D009439:Neuraminidase / Q000737:chemistry* / Q000235:genetics; D009844:Oligosaccharides / Q000737:chemistry*; D015800:Protozoan Proteins / Q000737:chemistry* / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D014349:Trypanosoma cruzi / Q000201:enzymology* / Q000235:genetics
chemical_list: D002364:Caseins; D006023:Glycoproteins; D009844:Oligosaccharides; D015800:Protozoan Proteins; D011994:Recombinant Proteins; C013084:lacto-N-neotetraose; C069715:trans-sialidase; D009439:Neuraminidase; D019158:N-Acetylneuraminic Acid