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Enzymatic synthesis of blood group A and B trisaccharide analogues
- Source :
- Carbohydrate Research. 324:161-169
- Publication Year :
- 2000
- Publisher :
- Elsevier BV, 2000.
-
Abstract
- Glycosyltransferases A and B utilize the donor substrates UDP-GalNAc and UDP-Gal, respectively, in the biosynthesis of the human blood group A and B trisaccharide antigens from the O(H)-acceptor substrates. These enzymes were cloned as synthetic genes and expressed in Escherichia coli, thereby generating large quantities of enzyme for donor specificity evaluations. The amino acid sequence of glycosyltransferase A only differs from glycosyltransferase B by four amino acids, and alteration of these four amino acid residues (Arg-176-->Gly, Gly-235-->Ser, Leu-266-->Met and Gly-268-->Ala) can change the donor substrate specificity from UDP-GalNAc to UDP-Gal. Crossovers in donor substrate specificity have been observed, i.e., the A transferase can utilize UDP-Gal and B transferase can utilize UDP-GalNAc donor substrates. We now report a unique donor specificity for each enzyme type. Only A transferase can utilize UDP-GlcNAc donor substrates synthesizing the blood group A trisaccharide analog alpha-D-Glcp-NAc-(1-->3)-[alpha-L-Fucp-(1-->2)]-beta-D-Galp-O-(CH2 )7CH3 (4). Recombinant blood group B was shown to use UDP-Glc donor substrates synthesizing blood group B trisaccharide analog alpha-D-Glcp-(1-->3)-[alpha-L-Fucp-(1-->2)]-beta-D-Galp-O-(CH2) 7CH3 (5). In addition, a true hybrid enzyme was constructed (Gly-235-->Ser, Leu-266-->Met) that could utilize both UDP-GlcNAc and UDP-Glc. Although the rate of transfer with UDP-GlcNAc by the A enzyme was 0.4% that of UDP-GalNAc and the rate of transfer with UDP-Glc by the B enzyme was 0.01% that of UDP-Gal, these cloned enzymes could be used for the enzymatic synthesis of blood group A and B trisaccharide analogs 4 and 5.
- Subjects :
- Oligosaccharides, Branched-Chain
Uridine Diphosphate Glucose
Stereochemistry
Molecular Sequence Data
Oligosaccharides
Biochemistry
ABO Blood-Group System
Substrate Specificity
Analytical Chemistry
chemistry.chemical_compound
Biosynthesis
Glycosyltransferase
Humans
Trisaccharide
Peptide sequence
chemistry.chemical_classification
Uridine Diphosphate N-Acetylglucosamine
biology
Chemistry
Organic Chemistry
General Medicine
Galactosyltransferases
Amino acid
carbohydrates (lipids)
Enzyme
Uridine diphosphate N-acetylglucosamine
Carbohydrate Sequence
biology.protein
Uridine diphosphate glucose
N-Acetylgalactosaminyltransferases
Trisaccharides
Subjects
Details
- ISSN :
- 00086215
- Volume :
- 324
- Database :
- OpenAIRE
- Journal :
- Carbohydrate Research
- Accession number :
- edsair.doi.dedup.....be909ae4f478618117117438e94e4a49