US 11,926,858 B2
Oligosaccharide production
Margit Pedersen, Roskilde (DK); Manos Papadakis, Copenhagen V (DK); Peter Becker, Virum (DK); Eric Samain, Gieres (FR); Pauline Peltier-Pain, Orleans (FR); Katrine Bych, Valby (DK); Ted Johanson, Virum (DK); Elise Champion, Toulouse (FR); and Gyula Dekany, Sinnamon Park (AU)
Assigned to GLYCOM A/S, Hørsholm (DK)
Filed by GLYCOM A/S, Hørsholm (DK)
Filed on Apr. 4, 2022, as Appl. No. 17/657,787.
Application 17/657,787 is a continuation of application No. 16/927,700, filed on Jul. 13, 2020, granted, now 11,293,042.
Application 16/927,700 is a continuation in part of application No. 15/321,996, granted, now 10,731,193, issued on Aug. 4, 2020, previously published as PCT/DK2015/050191, filed on Jun. 29, 2015.
Claims priority of application No. PA 2014 70392 (DK), filed on Jun. 27, 2014.
Prior Publication US 2022/0235388 A1, Jul. 28, 2022
Int. Cl. C12P 19/18 (2006.01); C07K 14/245 (2006.01); C12N 9/10 (2006.01); C12N 9/12 (2006.01); C12N 9/24 (2006.01); C12N 9/38 (2006.01); C12N 15/52 (2006.01); C12P 19/12 (2006.01)
CPC C12P 19/18 (2013.01) [C07K 14/245 (2013.01); C12N 9/1029 (2013.01); C12N 9/1051 (2013.01); C12N 9/1205 (2013.01); C12N 9/2402 (2013.01); C12N 9/2471 (2013.01); C12N 15/52 (2013.01); C12P 19/12 (2013.01); C12Y 203/01018 (2013.01); C12Y 204/01149 (2013.01); C12Y 302/01023 (2013.01); C12Y 302/01026 (2013.01); C12Y 204/01094 (2013.01)] 20 Claims
 
1. A genetically modified microorganism for making an oligosaccharide or a glycoside of said oligosaccharide, comprising:
two or more recombinant genes encoding two or more glycosyl transferases which can transfer a glycosyl residue of an activated sugar nucleotide to a carbohydrate acceptor or a glycoside of said acceptor,
a biosynthetic pathway for making said activated sugar nucleotide from sucrose, and
one or more genes encoding a heterologous PTS-dependent sucrose utilization system, so that said cell is capable to use sucrose as a carbon source for making said activated sugar nucleotide and as energy source for making said oligosaccharide,
wherein the carbohydrate acceptor is not sucrose,
wherein said genetically modified microorganism is an E. coli cell of LacZ− genotype,
said oligosaccharide is lacto-N-tetraose (LNT),
said activated sugar nucleotide is UDP-Gal and UDP-GlcNAc,
said two or more glycosyl transferase comprises β-1,3-N-acetyl glucosaminyl transferase and β-1,3-galactosyl transferase,
and said heterologous PTS-dependent sucrose utilization system comprises proteins encoded by scr genes scrY for a sucrose porin, scrA for a PTS permease, scrB for a sucrose-6-phosphate hydrolase and scrR for a repressor protein.