Taxonomic group: bacteria / Proteobacteria
(Phylum: Proteobacteria)
Host organism: Homo sapiens
Associated disease: bacillary dysentery [ICD11:
1A02 
, ICD11:
XN7HG ];
bacillary dysentery (shigellosis) [ICD11:
1A02 , ICD11:
SA56 , ICD11:
XN7HG ];
infection due to Shigella boydii [ICD11:
XN8RN ]
NCBI PubMed ID: 21057010Journal NLM ID: 2985120RPublisher: American Society for Microbiology
Correspondence: brockhau
queensu.ca; wanglei
nankai.edu.cn
Institutions: TEDA School of Biological Sciences and Biotechnology, Nankai University, Hongda Street, TEDA, Tianjin 300457, P.R.China, Tianjin Key Laboratory of Microbial Functional Genomics, P. R. China, Department of Medicine and Department of Biochemistry, Queen's University, Kingston, ON, Canada, and Key Laboratory of Molecular Microbiology and Technology, Ministry of Education, P. R. China, Department of Chemistry, Queen's University, Kingston, ON, Canada.
The O-antigen is the outer part of the lipopolysaccharide (LPS) in the outer membrane of Gram-negative bacteria and contains many repeats of an oligosaccharide unit. It contributes to antigenic variability and is essential to the full function and virulence of bacteria. Shigella is a Gram-negative human pathogen that causes diarrhea in humans. The O-antigen of Shigella boydii type 14 consists of repeating oligosaccharide units with the structure [→6 DGalp α1→4 DGlcpA β1→6 DGalp β1→4 DGalp β1→4 DGlcpNAc β1→]n. The wfeD gene in the O-antigen gene cluster of Shigella boydii type 14 was proposed to encode a galactosyltransferase (GalT) involved in O-antigen synthesis. We confirmed here that the wfeD gene product is a β4GalT that synthesizes the Galβ1-4GlcNAcα-R linkage. WfeD was expressed in E.coli and the activity characterized using UDP-[(3)H]Gal as the donor substrate and the synthetic acceptor substrate GlcNAcα-pyrophosphate-(CH2)11-O-phenyl. Enzyme product was analyzed by LC-MS, HPLC, NMR and galactosidase digestion. The enzyme was shown to be specific for the UDP-Gal donor substrate and required pyrophosphate in the acceptor substrate. Divalent metal ions such as Mn(2+), Ni(2+), and surprisingly also Pb(2+), enhanced the enzyme activity. Mutational analysis showed that the Glu101 residue within a DxD motif is essential for activity, possibly by forming the catalytic nucleophile. The Lys211 residue was also shown to be required for activity and may be involved in binding the negatively charged acceptor substrate. Our study revealed that the β4-GalT WfeD is a novel enzyme that has virtually no sequence similarity to mammalian β4GalT although it catalyzes a similar reaction.
structure, virulence, O-antigen, gene cluster, specific, galactosyltransferase, Shigella boydii
Structure type: polymer chemical repeating unit
Location inside paper: abstract, p.449
Compound class: O-polysaccharide, O-antigen
Contained glycoepitopes: IEDB_115136,IEDB_130646,IEDB_135813,IEDB_136044,IEDB_136906,IEDB_137340,IEDB_137472,IEDB_140108,IEDB_140122,IEDB_140630,IEDB_141794,IEDB_141807,IEDB_151528,IEDB_151531,IEDB_158533,IEDB_190606,IEDB_221845,IEDB_423153,SB_165,SB_166,SB_187,SB_195,SB_30,SB_7,SB_88
Methods: 13C NMR, 1H NMR, SDS-PAGE, ESI-MS, Western blotting, MALDI-TOF MS, genetic methods, biochemical methods, HPLC, LC-MS, mutation analysis
Biological activity: galactosyltransferase activity data
Biosynthesis and genetic data: genetic data
Synthetic data: chemical and chemoenzymatic
Related record ID(s): 27012
NCBI Taxonomy refs (TaxIDs): 621Reference(s) to other database(s): GTC:G17781ZM, GlycomeDB:
3526
Show glycosyltransferases
There is only one chemically distinct structure:
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