Taxonomic group: bacteria / Proteobacteria
(Phylum: Proteobacteria)
Associated disease: infection due to Campylobacter jejuni [ICD11:
XN4Q5 
]
NCBI PubMed ID: 21542610Publication DOI: 10.1021/bi2003372Journal NLM ID: 0370623Publisher: American Chemical Society
Correspondence: imper
mit.edu
Institutions: Departments of Chemistry and Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA, Center for Molecular Biology and Neuroscience and Department of Molecular Biosciences, University of Oslo, 0216 Oslo, Norway
The O-linked protein glycosylation pathway in Neisseria gonorrhoeae is responsible for the synthesis of a complex oligosaccharide on undecaprenyl diphosphate and subsequent en bloc transfer of the glycan to serine residues of select periplasmic proteins. Protein glycosylation (pgl) genes have been annotated on the basis of bioinformatics and top-down mass spectrometry analysis of protein modifications in pgl-null strains [Aas, F. E., et al. (2007) Mol. Microbiol. 65, 607-624; Vik, A., et al. (2009) Proc. Natl. Acad. Sci. U.S.A. 106, 4447-4452], but relatively little biochemical analysis has been performed to date. In this report, we present the expression, purification, and functional characterization of seven Pgl enzymes. Specifically, the enzymes studied are responsible for synthesis of an uncommon uridine diphosphate (UDP)-sugar (PglD, PglC, and PglB-acetyltransferase domain), glycan assembly (PglB-phospho-glycosyltransferase domain, PglA, PglE, and PglH), and final oligosaccharide transfer (PglO). UDP-2,4-diacetamido-2,4,6-trideoxy-α-D-hexose (DATDH), which is the first sugar in glycan biosynthesis, was produced enzymatically, and the stereochemistry was assigned as uridine diphosphate N'-diacetylbacillosamine (UDP-diNAcBac) by nuclear magnetic resonance characterization. In addition, the substrate specificities of the phospho-glycosyltransferase, glycosyltransferases, and oligosaccharyltransferase (OTase) were analyzed in vitro, and in most cases, these enzymes exhibited strong preferences for the native substrates relative to closely related glycans. In particular, PglO, the O-linked OTase, and PglB(Cj), the N-linked OTase from Campylobacter jejuni, preferred the native N. gonorrhoeae and C. jejuni substrates, respectively. This study represents the first comprehensive biochemical characterization of this important O-linked glycosylation pathway and provides the basis for further investigations of these enzymes as antibacterial targets.
biosynthesis, Campylobacter jejuni, glycosyltransferases, Neisseria gonorrhoeae, bioinformatics, Uridine Diphosphate Sugars, O-linked glycosylation
Structure type: oligomer
Location inside paper: Fig.S12
Aglycon: undecaprenyl pyrophosphate
Trivial name: Und-PP-diNAcBac-(GalNAc)2
Contained glycoepitopes: IEDB_130648,IEDB_137473,IEDB_1391961,IEDB_141584,IEDB_885822
Methods: 1H NMR, glycosyltransferase assays, biochemical methods, radioactivity measurement, HPLC
Biological activity: data for glycosyltransferase substrate specificity
Related record ID(s): 29194, 29540, 29541, 29542, 29543
NCBI Taxonomy refs (TaxIDs): 197Reference(s) to other database(s): GTC:G46503VQ
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There is only one chemically distinct structure:
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