Production of alpha-galactosyl epitopes via combined use of two recombinant whole cells harboring UDP-galactose 4-epimerase and alpha-1,3-galactosyltransferase.
alpha-Galactosyl epitopes (or alpha-Gal, oligosaccharides with a terminal Galalpha1,3Gal sequence) are a class of biologically important oligosaccharides in great demand in bulk quantities for basic and clinical studies on preventing hyperacute rejection in pig-to-primate organ xenotransplantation. A truncated bovine alpha-1, 3-galactosyltransferase, the key enzyme responsible for the biosynthesis of the terminal structure of alpha-Gal, was cloned and overexpressed previously. The acceptor specificity was further studied in the present paper, and lactose and galactose derivatives were found to be good acceptors. To develop a more proficient reaction process, we report herein an example of an efficient enzymatic synthesis of alpha-Gal oligosaccharides catalyzed by the combination of two recombinant Escherichia coli whole cells harboring the genes of a UDP-galactose 4-epimerase and the alpha-1, 3-galactosyltransferase, respectively. Using lactosyl azide (LacN(3)) as the acceptor for the glycosyltransferase, the combined use of the two recombinant cells efficiently produced alpha-Gal epitope Gal alpha1,3LacN(3) in 60-68% yield.
Department of Chemistry, Wayne State University, Detroit, Michigan 48202, USA.
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D002240:Carbohydrate Sequence; D003470:Culture Media; D000939:Epitopes / Q000737:chemistry / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D005690:Galactose / Q000737:chemistry*; D005700:Galactosyltransferases / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D010957:Plasmids; D011995:Recombination, Genetic; D014534:UDPglucose 4-Epimerase / Q000737:chemistry*
D003470:Culture Media; D000939:Epitopes; D005700:Galactosyltransferases; D014534:UDPglucose 4-Epimerase; D005690:Galactose