Taxonomic group: bacteria / Proteobacteria
(Phylum: Proteobacteria)
Host organism: Homo sapiens
Associated disease: meningitis [ICD11:
1D01 
];
infection due to Neisseria meningitidis [ICD11:
XN1DV ]
The structure was elucidated in this paperNCBI PubMed ID: 24849599Publication DOI: 10.1074/jbc.M114.575142Journal NLM ID: 2985121RPublisher: Baltimore, MD: American Society for Biochemistry and Molecular Biology
Correspondence: gerardy-schahn.rita
mh-hannover.de; francesco.berti
novartis.com
Institutions: Novartis Vaccines, Research, Via Fiorentina 1, 53100 Siena, Italy, Institute for Cellular Chemistry, Hannover Medical School, 30625 Hannover, Germany
The human pathogen Neisseria meningitidis (Nm) is a leading cause of bacterial meningitis and sepsis globally. A major virulence factor of Nm is the capsular polysaccharide (CPS), which in Nm serogroup A consists of N-acetyl-mannosamine-1-phosphate units linked together by phosphodiester linkages [→6)-α-D-ManNAc-(1→OPO3 (-)→]n. Acetylation in O-3 (to a minor extent in O-4) position results in immunologically active polymer. In the capsule gene cluster (cps) of Nm, region A contains the genetic information for CPSA biosynthesis. Thereby the open reading frames csaA, -B, and -C are thought to encode the UDP-N-acetyl-D-glucosamine-2-epimerase, poly-ManNAc-1-phosphate-transferase, and O-acetyltransferase, respectively. With the aim to use a minimal number of recombinant enzymes to produce immunologically active CPSA, we cloned the genes csaA, csaB, and csaC and functionally characterized the purified recombinant proteins. If recombinant CsaA and CsaB were combined in one reaction tube, priming CPSA-oligosaccharides were efficiently elongated with UDP-GlcNAc as the donor substrate, confirming that CsaA is the functional UDP-N-acetyl-D-glucosamine-2-epimerase and CsaB the functional poly-ManNAc-1-phosphate-transferase. Subsequently, CsaB was shown to transfer ManNAc-1P onto O-6 of the non-reducing end sugar of priming oligosaccharides, to prefer non-O-acetylated over O-acetylated primers, and to efficiently elongate the dimer of ManNAc-1-phosphate. The in vitro synthesized CPSA was purified, O-acetylated with recombinant CsaC, and proven to be identical to the natural CPSA by (1)H NMR, (31)P NMR, and immunoblotting. If all three enzymes and their substrates were combined in a one-pot reaction, nature identical CPSA was obtained. These data provide the basis for the development of novel vaccine production protocols.
polysaccharide, epimerase, Gram-negative bacteria, acetyltransferase, vaccine development, enzyme catalysis, nuclear magnetic resonance (NMR), Capsule Polymerase, Neisseria meningitidis Serogroup A, Recombinant Protein Expression
Structure type: monomer
Location inside paper: p.19396, Fig.1
Compound class: CPS
Contained glycoepitopes: IEDB_149549
Methods: 13C NMR, 1H NMR, NMR-2D, DNA cloning, SDS-PAGE, 31P NMR, genetic methods, radioactivity measurement, HPLC, immunoblotting, HPAEC-PAD
Enzymes that release or process the structure: CsaB (poly-ManNAc-1-phosphate-transferase), CsaC (O-acetyltransferase)
Comments, role: non-reducing end of CPS; part of the CPS (see RR:30228)
Related record ID(s): 30052, 30228, 30230, 30231, 30232, 30233
NCBI Taxonomy refs (TaxIDs): 65699
Show glycosyltransferases
There is only one chemically distinct structure:
Found 
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