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Taxonomic group: bacteria / Proteobacteria (Phylum: Proteobacteria)
Associated disease: nosocomial infections [ICD11: XB25

]; urinary tract infections (UTI) [ICD11: GC08

]; bacteremia [ICD11: MA15.0

]; septicemia [ICD11: MA15.Y

]; infection due to Klebsiella pneumoniae [ICD11: XN741

]

NCBI PubMed ID: 31138653
Publication DOI: 10.1074/jbc.RA119.008969
Journal NLM ID: 2985121R
Publisher: Baltimore, MD: American Society for Biochemistry and Molecular Biology
Correspondence: Chris Whitfield <cwhitfie

uoguelph.ca>
Institutions: Alberta Glycomics Centre and Department of Chemistry, University of Alberta, Edmonton, AB, Canada, Department of Molecular and Cellular Biology, University of Guelph, Ontario, Canada

A limited range of different structures is observed in O-antigenic polysaccharides (OPSs) from Klebsiella pneumoniae lipopolysaccharides. Among these, several are based on modifications of a conserved core element of serotype O2a OPS, which has a disaccharide repeat structure [→3)-α-Galp-(1→3)-β-Galf-(1→]. Here, we describe the enzymatic pathways for a highly unusual modification strategy involving the attachment of a second glycan repeat unit structure to the non-reducing terminus of O2a. This occurs by the addition of the O1 [→3)-α-Galp-(1→3)-β-Galp-(1→] or O2c [→3)-β-GlcpNAc-(1→5)-β-Galf-(1→] antigens. The organization of the enzyme activities performing these modifications differs, with the enzyme WbbY possessing two glycosyltransferase catalytic sites solely responsible for O1-antigen polymerization and forming a complex with the O2a glycosyltransferase, WbbM. In contrast, O2c polymerization requires glycosyltransferases WbmV and WbmW, which interact with one another, but apparently not with WbbM. Using defined synthetic acceptors and site-directed mutants to assign the activities of the WbbY catalytic sites, we found that the C-terminal WbbY domain is a UDP-Galp-dependent GT-A galactosyltransferase adding β-(1→3)-linked D-Galp, whereas the WbbY N terminus includes a GT-B enzyme adding α-(1→3)-linked D-Galp These activities build the O1 antigen on a terminal Galp in the O2a domain. Using similar approaches, we identified WbmV as the UDP-GlcNAc-dependent transferase and noted that WbmW represents a UDP-Galf-dependent enzyme and that both are GT-A members. WbmVW polymerizes the O2c antigen on a terminal Galf Our results provide mechanistic and conceptual insights into an important strategy for polysacharide antigen diversification in bacteria.

polysaccharide, O antigen, Gram-negative bacteria, glycosyltransferase, Klebsiella pneumoniae, serotyping, lipopolysaccharide (LPS), antigenic diversity, cell surface, enzyme complex

Structure type: oligomer
Location inside paper: fig.1, O1/O2 OPS
Compound class: O-polysaccharide
Contained glycoepitopes: IEDB_115013,IEDB_130645,IEDB_136044,IEDB_136095,IEDB_136906,IEDB_137472,IEDB_141794,IEDB_144987,IEDB_149558,IEDB_151528,IEDB_190606,IEDB_918314,SB_165,SB_166,SB_187,SB_195,SB_31,SB_7,SB_87,SB_88

Methods: 13C NMR, 1H NMR, gel filtration, NMR-2D, PCR, DNA sequencing, SDS-PAGE, TLC, chemical synthesis, HPLC, UV, immunoblotting, LC-MS, bioinformatic analysis, CC, LC-ESI-MS, enzymatic synthesis, fluorescent labeling
Biosynthesis and genetic data: gmlABC

Related record ID(s): 792, 793, 794, 796, 797, 798, 1044
NCBI Taxonomy refs (TaxIDs): 573
Reference(s) to other database(s): GTC:G08095DU
Show glycosyltransferases

There is only one chemically distinct structure:

OC[C@@H](O)[C@@H]1O[C@@H](O[C@H]2[C@@H](O[C@H]3O[C@H](CO)[C@H](O)[C@H](O)[C@H]3O)[C@@H](CO)O[C@H](O[C@H]3[C@H]([C@H](O)CO)O[C@@H](O[C@H]4[C@@H](O[C@H]5O[C@H](CO)[C@H](O)[C@H](O)[C@H]5O)[C@@H](CO)O[C@H](O[C@H]5[C@H]([C@H](O)CO)O[C@@H](O[C@H]6[C@@H](O[C@H]7O[C@H](CO)[C@H](O)[C@H](O)[C@H]7O)[C@@H](CO)O[C@H](O[C@H]7[C@H]([C@H](O)CO)O[C@@H](O[C@H]8[C@@H](O[C@H]9O[C@H](CO)[C@H](O)[C@H](O)[C@H]9O)[C@@H](CO)O[C@H](O)[C@@H]8O)[C@@H]7O)[C@@H]6O)[C@@H]5O)[C@@H]4O)[C@@H]3O)[C@@H]2O)[C@H](O)[C@H]1O[C@H]1O[C@H](CO)[C@H](O)[C@H](O[C@@H]2O[C@H](CO)[C@H](O)[C@H](O[C@H]3O[C@H](CO)[C@H](O)[C@H](O[C@@H]4O[C@H](CO)[C@H](O)[C@H](O[C@H]5O[C@H](CO)[C@H](O)[C@H](O[C@@H]6O[C@H](CO)[C@H](O)[C@H](O)[C@H]6O)[C@H]5O)[C@H]4O)[C@H]3O)[C@H]2O)[C@H]1O

SVG MW SMILES 3D mol Ionize

Warning = sub-repeat ending at aDGalp: unknown unit count was assumed to be 3
Warning = sub-repeat ending at aDGalp: unknown unit count was assumed to be 3