Journal
GLYCOBIOLOGY
Volume 12, Issue 4, Pages 235-240Publisher
OXFORD UNIV PRESS INC
DOI: 10.1093/glycob/12.4.235
Keywords
glycosyltransferase; high-cell-density cultivation; metabolic engineering; oligosaccharide
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When fed to a beta-galactosidase-negative (lacZ(-)) Escherichia coli strain that was grown on an alternative carbon source (such as glycerol), lactose accumulated intracellularly on induction of the lactose permease. We showed that intracellular lactose was efficiently glycosylated when genes of glycosyltransferase that use lactose as acceptor were expressed. High-cell-density cultivation of lacZ- strains that overexpressed the beta1,3 N acetyl glucosaminyltransferase lgtA gene of Neisseria meningitidis resulted in the synthesis of 6 g . L-1 of the expected trisaccharide (GlcNAcbeta1-3Galbeta1-4Glc). When the beta1,4 galactosyltransferase lgtB gene of N. meningitidis was coexpressed with lgtA, the trisaccharide was further converted to lacto-N-neotetraose (Galbeta1-4GlcNAcbeta1-3Galbeta1-4Glc) and lacto-N-neoheaxose with a yield higher than 5 g . L-1. In a similar way, the nanA(-) E. coli strain that was devoid of NeuAc aldolase activity accumulated NeuAc on induction of the NanT permease and the lacZ- nanA- strain that overexpressed the N. meningitidis genes of the alpha2,3 sialyltransferase and of the CMP-NeuAc synthase efficiently produced sialyllactose (NeuAcalpha2-3Galbeta1-4Glc) from exogenous NeuAc and lactose.
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