Enzymatic synthesis of homogeneous chondroitin sulfate oligosaccharides
AssigneeThe University of North Carolina at Chapel Hill
InventorsJian Liu, Jine Li, Guowei Su
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Abstract
Methods of synthesizing chondroitin sulfate oligosaccharides are provided. Enzymatic schematic approaches to synthesizing structurally defined homogenous chondroitin sulfate oligosaccharides at high yields are provided. Synthetic chondroitin sulfate oligosaccharides ranging from 3-mers to 15-mers are provided.
Claims22
A method of synthesizing a synthetic chondroitin sulfate oligosaccharide, the method comprising:
- providing a chondroitin backbone;
- and
- performing at least two of the following enzymatic reaction steps:
- an elongation step to add a GalNAc residue using a glycosyltransferase from E. coli K4 (KfoC);
- an elongation step to add a GlcA residue using KfoC;
- a 6-O-sulfation step using chondroitin sulfate 6-O-sulfotransferase (CS6OST) and sulfate donor 3′-phosphoadenosine 5′-phosphosulfate (PAPS);
- and
- a 4-O-sulfation step involving chondroitin sulfate 4-O-sulfotransferase (CS4OST) and PAPS;
- whereby a synthetic chondroitin sulfate oligosaccharide is synthesized.
A synthetic chondroitin sulfate oligosaccharide, comprising a 4-O-sulfated and/or 6-O-sulfated chondroitin oligosaccharide having a size ranging from a 3-mer to a 15-mer.
A synthetic chondroitin sulfate oligosaccharide, wherein the synthetic chondroitin sulfate oligosaccharides comprise a structure selected from the group consisting of:
- where R1=SO3H;
- R2=H, R3=NHAc,
- where R1=H;
- R2=H;
- R3=NHAc,
- where R1=R2=H;
- R3=NHFTA,
- where R1=SO3H;
- R2=H;
- R3=NHFTA,
- where R1=R2=H;
- R3=NH2,
- where R1=SO3H;
- R2=H;
- R3=NH2,
- where R1=R2=H;
- R3=N3, or where R1=SO3H;
- R2=H;
- R3=N3;
- where RB1=RD1=RF1=SO3H;
- R2=H,
- where RB1=RD1=RF1=H;
- R2=SO3H,
- where RB1=SO3H;
- RD1=RF1=R2=H, or
- where RB1=RD1=SO3H;
- RF1=R2=H;
- and
- where R1=SO3H;
- R2=H, or
- where R1=H;
- R2=SO3H,
- wherein R is selected from the group consisting of —H, alkyl (such as but not limited to —CH3 or —CH2CH3), substituted alkyl, aryl, and substituted aryl (such as but not limited to a p-nitrophenyl group).
A synthetic chondroitin sulfate oligosaccharide, wherein the synthetic chondroitin sulfate oligosaccharides comprise a structure selected from the group consisting of:
- wherein n=4 (CS-A 11-mer), n=5 (CS-A 13-mer), n=6 (CS-A 15-mer), n=7 (CS-A 17-mer), or n=8 (CS-A 19-mer);
- wherein n=4 (CS-C 11-mer), n=5 (CS-C 13-mer), n=6 (CS-C 15-mer), n=7 (CS-C 17-mer), or n=8 (CS-C 19-mer);
- and
- wherein n=1 (CS-E 5-mer), n=2 (CS-E 7-mer), n=3 (CS-E 9-mer), n=4 (CS-E 11-mer), n=5 (CS-E 13-mer), n=6 (CS-E 15-mer), n=7 (CS-E 17-mer) or n=8 (CS-E 19-mer),
- wherein R is selected from the group consisting of —H, alkyl (such as but not limited to —CH3 or —CH2CH3), substituted alkyl, aryl, and substituted aryl (such as but not limited to a p-nitrophenyl group).
Dependencies are inferred from the claim text.