Taxonomic group: bacteria / Proteobacteria
(Phylum: Proteobacteria)
Associated disease: infection due to Neisseria meningitidis [ICD11:
XN1DV 
];
infection due to Escherichia coli [ICD11:
XN6P4 ]
NCBI PubMed ID: 10521543Publication DOI: 10.1093/glycob/9.10.1061Journal NLM ID: 9104124Publisher: IRL Press at Oxford University Press
Institutions: Department of Chemistry, Swedish University of Agricultural Sciences, Uppsala, Sweden, Department of Medical Chemistry, Vrije Universiteit, Van der Boechorstraat 7, 1081 BT Amsterdam, The Netherlands
We have expressed the Neisseria meningitidis lgtA gene at a high level in Escherichia coli. The encoded β-N-acetylglucosaminyltransferase, referred to as LgtA, which in the bacterium is involved in the synthesis of the lacto-N-neo-tetraose structural element of the bacterial lipooligosaccharide, was obtained in an enzymatically highly active form. This glycosyltransferase appeared to be unusual in that it displays a broad acceptor specificity toward both α- and β-galactosides, whether structurally related to N- or O-protein-, or lipid-linked oligosaccharides. Product analysis by one- and two-dimensional 400 MHz 1H- and 13C NMR spectroscopy reveals that LgtA catalyzes the introduction of GlcNAc from UDP-GlcNAc in a β1→3-linkage to accepting Gal residues. The enzyme can thus be characterized as a UDP-GlcNAc:Gal α/β-R β 3-N-acetylglucosaminyltransferase. Although lactose is a highly preferred acceptor substrate the recombinant enzyme also acts efficiently on monomeric and dimeric N-acetyllactosamine revealing its potential value in the synthesis of polylactosaminoglycan structures in enzyme assisted procedures. Furthermore, LgtA shows a high donor promiscuity toward UDP-GalNAc, but not toward other UDP-sugars, and can catalyze the introduction of GalNAc in β1→3-linkage to α- or β-Gal in the acceptor structures at moderate rates. LgtA therefore shows promise to be a useful catalyst in the preparative synthesis of both GlcNAc β1→3 Gal and GalNAc β1→3 Gal linkages.
oligosaccharide, enzyme-assisted-synthesis, recombinant glycosyltransferase, glycosidic linkage, polylactosaminoglycan, recombinant glycosyltrasferase
Structure type: oligomer
Location inside paper: Fig. 4, structure A, Table VII
Aglycon: p-nitrophenyl
Contained glycoepitopes: IEDB_135813,IEDB_136044,IEDB_137340,IEDB_137472,IEDB_1391966,IEDB_141794,IEDB_141807,IEDB_142351,IEDB_142487,IEDB_142488,IEDB_146664,IEDB_151531,IEDB_190606,IEDB_983931,SB_145,SB_165,SB_166,SB_173,SB_187,SB_192,SB_195,SB_6,SB_7,SB_88
Methods: 13C NMR, 1H NMR, NMR-2D, SDS-PAGE, enzyme-assisted synthesis, DNA techniques, glycosyltransferase assays, kinetics assays
Enzymes that release or process the structure: LgtA, N-acetylglucosaminyltransferase
Biosynthesis and genetic data: biosynthesis data
Synthetic data: enzymatic
Comments, role: product obtained in large scale incubation using LgdtA
Related record ID(s): 942, 1627, 1715, 1717, 1719, 1720, 1721, 1722, 1723, 1724, 1725, 1726, 1728, 1729, 1730, 1731, 1732, 1734, 1735, 1736, 1737, 1738, 1739, 1740, 1741, 1742, 1743, 1744, 1745, 1746, 1747, 1748, 1749, 1750, 1751, 1752, 1753, 1754, 1755, 1756, 1757, 1758, 1759, 1760, 1761, 1763, 1764, 8942, 9073, 9074, 9075, 9078, 9079, 9080
NCBI Taxonomy refs (TaxIDs): 487,
562Reference(s) to other database(s): GTC:G06239NA, GlycomeDB:
599
Show glycosyltransferases
NMR conditions: in D2O at 303 K
[as TSV]
13C NMR data:
Linkage Residue C1 C2 C3 C4 C5 C6
4,3,2 Ac ? 23.1
4,3 bDGlcpN 103.4 ? ? ? ? ?
4 bDGalp 103.6 70.4 82.6 69.1 ? ?
bDGlcp 99.8 72.9 78.4 75.6 ? ?
1H NMR data:
Linkage Residue H1 H2 H3 H4 H5 H6
4,3,2 Ac - 2.048
4,3 bDGlcpN 4.698 ? ? ? ? ?
4 bDGalp 4.480 3.631 3.738 4.162 ? ?
bDGlcp 5.310 3.684 3.791 3.856 ? ?
1H/13C HSQC data:
Linkage Residue C1/H1 C2/H2 C3/H3 C4/H4 C5/H5 C6/H6
4,3,2 Ac 23.1/2.048
4,3 bDGlcpN 103.4/4.698 ?/? ?/? ?/? ?/? ?/?
4 bDGalp 103.6/4.480 70.4/3.631 82.6/3.738 69.1/4.162 ?/? ?/?
bDGlcp 99.8/5.310 72.9/3.684 78.4/3.791 75.6/3.856 ?/? ?/?
1H NMR data:
| Linkage | Residue | H1 | H2 | H3 | H4 | H5 | H6 |
|---|
| 4,3,2 | Ac |
| 2.048 | |
| 4,3 | bDGlcpN | 4.698 | ? | ? | ? | ? | ? |
| 4 | bDGalp | 4.480 | 3.631 | 3.738 | 4.162 | ? | ? |
| | bDGlcp | 5.310 | 3.684 | 3.791 | 3.856 | ? | ? |
|
13C NMR data:
| Linkage | Residue | C1 | C2 | C3 | C4 | C5 | C6 |
|---|
| 4,3,2 | Ac | ? | 23.1 | |
| 4,3 | bDGlcpN | 103.4 | ? | ? | ? | ? | ? |
| 4 | bDGalp | 103.6 | 70.4 | 82.6 | 69.1 | ? | ? |
| | bDGlcp | 99.8 | 72.9 | 78.4 | 75.6 | ? | ? |
|
![](nmr_drawer.py?width=750&height=130&nucleus=C&rotate_labels=false&spectrum=[{"s":23.1,"l":"2","c":[0,0,0]},{"s":103.4,"l":"1","c":[0,0,225]},{"s":103.6,"l":"1","c":[230,0,30]},{"s":70.4,"l":"2","c":[230,0,30]},{"s":82.6,"l":"3","c":[230,0,30]},{"s":69.1,"l":"4","c":[230,0,30]},{"s":99.8,"l":"1","c":[0,165,0]},{"s":72.9,"l":"2","c":[0,165,0]},{"s":78.4,"l":"3","c":[0,165,0]},{"s":75.6,"l":"4","c":[0,165,0]}])
The spectrum also has 10 signals at unknown positions (not plotted). |
There is only one chemically distinct structure:
Found 
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