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Structure type: monomer
Trivial name: UDP-2,4-diacetamido-2,4,6-trideoxy-α-D-glucopyranose, UDP-2,4-diacetamido-Bacillose, UDP-N,N-diacetylbacillosamine, UPD-2,4-diacetamido-2,4,6-trideoxy-α-D-glucopyranose, UDP-N,N'-diacetyl-bacillosamine, UDP-Bac2Ac4Ac, UDP-diNAcBac
Compound class: nucleoside diphosphate sugar

• Article ID: 3404
  Guerry P, Ewing CP, Schoenhofen IC, Logan SM "Protein glycosylation in Campylobacter jejuni: partial suppression of pglF by mutation of pseC" - Journal of Bacteriology 189(18) (2007) 6731-6733
  

  Campylobacter jejuni has systems for N- and O-linked protein glycosylation. Although biochemical evidence demonstrated that a pseC mutant in the O-linked pathway accumulated the product of pglF in the N-linked pathway, analyses of transformation frequencies and glycosylation statuses of N-glycosylated proteins indicated a partial suppression of pglF by pseC

  biosynthesis, Bacterial Proteins, Campylobacter jejuni, glycosylation

  NCBI PubMed ID: 17631632
  Journal NLM ID: 2985120R
  Publisher: American Society for Microbiology
  Correspondence: guerryp@nmrc.navy.mil
  Institutions: Enteric Diseases Department, Naval Medical Research Center, 503 Robert Grant Ave., Silver Spring, MD 20910, USA
  Methods: PCR, serological methods, genetic methods
  
   
  
  Campylobacter jejuni
  CSDB ID 21967 (all data & tools)
• Article ID: 3475
  Glaze PA, Watson DC, Young NM, Tanner ME "Biosynthesis of CMP-N,N'-diacetyllegionaminic acid from UDP-N,N'-diacetylbacillosamine in Legionella pneumophila" - Biochemistry 47(10) (2008) 3272-3282
  

  Legionaminic acid is a nine-carbon α-keto acid that is similar in structure to other members of the sialic acid family that includes neuraminic acid and pseudaminic acid. It is found as a component of the lipopolysaccharide in several bacterial species and is perhaps best known for its presence in the O-antigen of the causative agent of Legionnaires' disease, Legionella pneumophila. In this work, the enzymes responsible for the biosynthesis and activation of N, N'-diacetyllegionaminic acid are identified for the first time. A cluster of three L. pneumophila genes bearing homology to known sialic acid biosynthetic genes (neuA,B,C) were cloned and overexpressed in Escherichia coli. The NeuC homologue was found to be a hydrolyzing UDP- N, N'-diacetylbacillosamine 2-epimerase that converts UDP- N, N'-diacetylbacillosamine into 2,4-diacetamido-2,4,6-trideoxymannose and UDP. Stereochemical and isotopic labeling studies showed that the enzyme utilizes a mechanism involving an initial anti elimination of UDP to form a glycal intermediate and a subsequent syn addition of water to generate product. This is similar to the hydrolyzing UDP- N-acetylglucosamine 2-epimerase (NeuC) of sialic acid biosynthesis, but the L. pneumophila enzyme would not accept UDP-GlcNAc as an alternate substrate. The NeuB homologue was found to be a N, N'-diacetyllegionaminic acid synthase that condenses 2,4-diacetamido-2,4,6-trideoxymannose with phosphoenolpyruvate (PEP), although the in vitro activity of the recombinant enzyme (isolated as a MalE fusion protein) was very low. The synthase activity was dependent on the presence of a divalent metal ion, and the reaction proceeded via a C-O bond cleavage process, similar to the reactions catalyzed by the sialic acid and pseudaminic acid synthases. Finally, the NeuA homologue was shown to possess the CMP- N, N'-diacetyllegionaminic acid synthetase activity that generates the activated form of legionaminic acid used in lipopolysaccharide biosynthesis. Together, the three enzymes constitute a pathway that converts a UDP-linked bacillosamine derivative into a CMP-linked legionaminic acid derivative

  O-antigen, lipopolysaccharide biosynthesis, pseudaminic acid, neuraminic acid, Legionella pneumophila, legionaminic acid, Legionella, bacillosamine

  NCBI PubMed ID: 18275154
  Journal NLM ID: 0370623
  Publisher: American Chemical Society
  Correspondence: mtanner@chem.ubc.ca
  Institutions: Department of Chemistry, University of British Columbia, Vancouver, British Columbia, Canada V6T 1Z1
  Methods: 13C NMR, 1H NMR, NMR-2D, SDS-PAGE, 31P NMR, genetic methods, biochemical methods
  
   
  
  Legionella pneumophila
  CSDB ID 22963 (all data & tools)
• Article ID: 3516
  McNally DJ, Schoenhofen IC, Houliston RS, Khieu NH, Whitfield DM, Logan SM, Jarrell HC, Brisson JR "CMP-Pseudaminic Acid is a Natural Potent Inhibitor of PseB, the First Enzyme of the Pseudaminic Acid Pathway in Campylobacter jejuni and Helicobacter pylori" - ChemMedChem 3(1) (2008) 55-59
  

  No abstract available

  Research, acid, Campylobacter, Campylobacter jejuni, NMR spectroscopy, pseudaminic acid, biological, Helicobacter pylori, enzyme, natural, pathway, Helicobacter, inhibitors, STD, STD NMR spectroscopy, potent

  NCBI PubMed ID: 17893902
  Journal NLM ID: 101259013
  Publisher: Wiley-VCH
  Correspondence: david.mcnally@nrc-cnrc.gc.ca
  Institutions: National Research Council of Canada-Institute for Biological Sciences, Ottawa ON, K1A 0R6, Canada, Fax: (+1) 613-952-9092
  Methods: 13C NMR, 1H NMR, NMR-2D, sugar analysis, NMR-1D, genetic methods, biochemical methods, STD NMR
  
   
  
  Campylobacter jejuni
  CSDB ID 23137 (all data & tools)
• Article ID: 3649
  Olivier NB, Chen MM, Behr JR, Imperiali B "In vitro biosynthesis of UDP-N,N'-diacetylbacillosamine by enzymes of the Campylobacter jejuni general protein glycosylation system" - Biochemistry 45(45) (2006) 13659-13669
  

  In Campylobacter jejuni 2,4-diacetamido-2,4,6-trideoxy-α-D-glucopyranose, termed N,N'-diacetylbacillosamine (Bac2,4diNAc), is the first carbohydrate in the glycoprotein N-linked heptasaccharide. With uridine diphosphate-N-acetylglucosamine (UDP-GlcNAc) as a starting point, two enzymes of the general protein glycosylation (Pgl) pathway in C. jejuni (PglF and PglE) have recently been shown to modify this sugar nucleotide to form UDP-2-acetamido-4-amino-2,4,6-trideoxy-α-D-glycopyranose (UDP-4-amino-sugar) [Schoenhofen, I. C., et al. (2006) J. Biol. Chem. 281, 723-732]. PglD has been proposed to catalyze the final step in N,N'-diacetylbacillosamine synthesis by N-acetylation of the UDP-4-amino-sugar at the C4 position. We have cloned, overexpressed, and purified PglD from the pgl locus of C. jejuni NCTC 11168 and identified it as the acetyltransferase that modifies the UDP-4-amino-sugar to form UDP-N,N'-diacetylbacillosamine, utilizing acetyl-coenzyme A as the acetyl group donor. The UDP-N,N'-diacetylbacillosamine product was purified from the reaction by reverse phase C18 HPLC and the structure determined by NMR analysis. Additionally, the full-length PglF was overexpressed and purified in the presence of detergent as a GST fusion protein, allowing for derivation of kinetic parameters. We found that the UDP-4-amino-sugar was readily synthesized from UDP-GlcNAc in a coupled reaction using PglF and PglE. We also demonstrate the in vitro biosynthesis of the complete heptasaccharide lipid-linked donor by coupling the action of eight enzymes (PglF, PglE, PglD, PglC, PglA, PglJ, PglH, and PglI) in the Pgl pathway in a single reaction vessel.

  biosynthesis, Campylobacter jejuni, glycosylation, acetyltransferase

  NCBI PubMed ID: 17087520
  Journal NLM ID: 0370623
  Publisher: American Chemical Society
  Correspondence: imper@mit.edu
  Institutions: Department of Chemistry, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, Massachusetts 02139, USA
  Methods: 1H NMR, 31P NMR, ESI-MS, MALDI-MS, genetic methods, biochemical methods, HPLC, capillary electrophoresis (CE)
  
   
  
  Campylobacter jejuni 11168
  CSDB ID 23936 (all data & tools)
• Article ID: 4134
  Watson DC, Leclerc S, Wakarchuk WW, Young NM "Enzymatic synthesis and properties of glycoconjugates with legionaminic acid as a replacement for neuraminic acid" - Glycobiology 21(1) (2011) 99-108
  

  In addition to sialic acid, bacteria produce several other nonulosonic acids, including legionaminic acid (Leg). This has exactly the same stereochemistry as sialic acid, with the added features of 9-deoxy and 7-amino groups. In order to explore the biological effects of replacing sialic acid residues (Neu5Ac) in glycoconjugates with Leg in its diacetylated form, diacetyllegionaminic acid (Leg5Ac7Ac), we tested CMP-Leg5Ac7Ac as a donor substrate with a selection of bacterial and mammalian sialyltransferases. The CMP-Leg5Ac7Ac was synthesized in vitro by means of cloned enzymes from the bacillosamine portion of the Campylobacter jejuni N-glycan pathway and from the Leg pathway of Legionella pneumophila. Using fluorescent derivatives of lactose, Galβ1,4GlcNAcβ and T-antigen (Galβ1,3GalNAcα) as acceptors, we tested eight different sialyltransferases and found that the Pasteurella multocida PM0188h and porcine ST3Gal1 sialyltransferases were significantly active with CMP-Leg5Ac7Ac, showing approximately 60% activity when compared with CMP-Neu5Ac. The Photobacterium α2,6 sialyltransferase was weakly active, with approximately 6% relative activity. The Leg5Ac7Ac-α-2,3-lactose product was then tested as a substrate with six sialidases of viral, bacterial and mammalian origin. All showed much lower activities than with the corresponding sialic acid substrate, with the influenza virus N1 being the most active and human NEU2 being the least active. These results show the feasibility of producing glycoconjugates with Leg5Ac7Ac residues as the terminal sugars, which should display novel biological properties.

  sialyltransferase, sialic acid, legionaminic acid, glycoconjugate, neuraminidase

  NCBI PubMed ID: 20978010
  Publication DOI: 10.1093/glycob/cwq135
  Journal NLM ID: 9104124
  Publisher: IRL Press at Oxford University Press
  Correspondence: martin.young@nrc-cnrc.gc.ca
  Institutions: Institute for Biological Sciences, National Research Council of Canada, 100 Sussex Drive, Ottawa, Ontario, Canada
  Methods: DNA cloning, genetic methods, biochemical methods, CE-MS, CE
  
   
  
  Campylobacter jejuni, Legionella pneumophila
  CSDB ID 27008 (all data & tools)
• Article ID: 4531
  Hartley MD, Morrison MJ, Aas FE, Borud B, Koomey M, Imperiali B "Biochemical characterization of the O-linked glycosylation pathway in Neisseria gonorrhoeae responsible for biosynthesis of protein glycans containing N,N'-diacetylbacillosamine" - Biochemistry 50(22) (2011) 4936-4948
  

  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

  NCBI PubMed ID: 21542610
  Publication DOI: 10.1021/bi2003372
  Journal NLM ID: 0370623
  Publisher: 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
  Methods: 1H NMR, glycosyltransferase assays, biochemical methods, radioactivity measurement, HPLC
  
   
  
  Neisseria gonorrhoeae
  CSDB ID 29540 (all data & tools)
• Article ID: 4581
  Morrison MJ, Imperiali B "Biosynthesis of UDP-N,N'-diacetylbacillosamine in Acinetobacter baumannii: Biochemical characterization and correlation to existing pathways" - Archives of Biochemistry and Biophysics 536(1) (2013) 72-80
  

  The Gram-negative, opportunistic pathogen Acinetobacter baumannii has recently captured headlines due to its ability to circumvent current antibiotic therapies. Herein we show that the multi-drug resistant (MDR) AYE strain of A. baumannii contains a gene locus that encodes three enzymes responsible for the biosynthesis of the highly-modified bacterial nucleotide sugar, UDP-N,N'-diacetylbacillosamine (UDP-diNAcBac). Previously, this UDP-sugar has been implicated in the pgl pathway of Campylobacter jejuni. Here we report the overexpression, purification, and biochemical characterization of the A. baumannii enzymes WeeK, WeeJ, and WeeI that are responsible for the production of UDP-diNAcBac. We also demonstrate the function of the phosphoglycosyltransferase (WeeH), which transfers the diNAcBac moiety to undecaprenyl-phosphate. UDP-diNAcBac biosynthesis in A. baumannii is also directly compared to the homologous pathways in the pathogens C. jejuni and Neisseria gonorrhoeae. This work demonstrates for the first time the ability of A. baumannii to generate the highly-modified, UDP-diNAcBac nucleotide sugar found previously in other bacteria adding to the growing list of pathogens that assemble glycoconjugates including bacillosamine. Additionally, characterization of these pathway enzymes highlights the opportunity for investigating the significance of highly-modified sugars in bacterial pathogenesis.

  biosynthesis, Acinetobacter baumannii, bacillosamine, acetyltransferases, aminotransferase, bacterial O-linked glycosylation, UDP-diNAcBac

  NCBI PubMed ID: 23747578
  Publication DOI: 10.1016/j.abb.2013.05.011
  Journal NLM ID: 0372430
  Correspondence: B. Imperiali
  Institutions: Department of Chemistry, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, MA 02139, United States, Department of Biology, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, MA 02139, United States
  Methods: SDS-PAGE, DNA techniques, glycosyltransferase assays, kinetics assays, genetic methods, biochemical methods, CE
  
   
  
  Acinetobacter baumannii AYE
  CSDB ID 29288 (all data & tools)
• Article ID: 4582
  Mostafavi AZ, Troutman JM "Biosynthetic assembly of the Bacteroides fragilis capsular polysaccharide A precursor bactoprenyl diphosphate-linked acetamido-4-amino-6-deoxygalactopyranose" - Biochemistry 52(11) (2013) 1939-1949
  

  The sugar capsule capsular polysaccharide A (CPSA), which coats the surface of the mammalian symbiont Bacteroides fragilis, is a key mediator of mammalian immune system development. In addition, this sugar polymer has shown therapeutic potential in animal models of multiple sclerosis and other autoimmune disorders. The structure of the CPSA polymer includes a rare stereoconfiguration sugar acetamido-4-amino-6-deoxygalactopyranose (AADGal) that we propose is the first sugar linked to a bactoprenyl diphosphate scaffold in the production of CPSA. In this report, we have utilized a heterologous system to reconstitute bactoprenyl diphosphate-linked AADGal production. Construction of this system included a previously reported Campylobacter jejuni dehydratase, PglF, coupled to a B. fragilis-encoded aminotransferase (WcfR) and initiating hexose-1-phosphate transferase (WcfS). The function of the aminotransferase was confirmed by capillary electrophoresis and a novel high-performance liquid chromatography (HPLC) method. Production of the rare uridine diphosphate (UDP)-AADGal was confirmed through a series of one- and two-dimensional nuclear magnetic resonance experiments and high-resolution mass spectrometry. A spectroscopically unique analogue of bactoprenyl phosphate was utilized to characterize the transfer reaction catalyzed by WcfS and allowed HPLC-based isolation of the isoprenoid-linked sugar product. Importantly, the entire heterologous system was utilized in a single-pot reaction to biosynthesize the bactoprenyl-linked sugar. This work provides the first critical step in the in vitro reconstitution of CPSA biosynthesis.

  Bacteroides fragilis, aminotransferase, caspsular polysaccharide A, biosynthesis pathway, WcfR, WcfS, acetamido-4-amino-6-deoxy-galactopyranose

  NCBI PubMed ID: 23458065
  Publication DOI: 10.1021/bi400126w
  Journal NLM ID: 0370623
  Publisher: American Chemical Society
  Correspondence: jerry.troutman@uncc.edu
  Institutions: Department of Chemistry, University of North Carolina at Charlotte , 9201 University City Boulevard, Charlotte, North Carolina 28223-0001, United States
  Methods: 1H NMR, NMR-2D, PCR, SDS-PAGE, TLC, ESI-MS, NMR-1D, genetic methods, biochemical methods, radioactivity measurement, HPLC, CE
  
   
  
  Campylobacter jejuni
  CSDB ID 29657 (all data & tools)

Show legend Show as text

Structure type: polymer chemical repeating unit
Compound class: CPS
Contained glycoepitopes: IEDB_137340,IEDB_141807,IEDB_151531

• Article ID: 6309
  Rudenko N, Karatovskaya A, Zamyatina A, Shepelyakovskaya A, Semushina S, Brovko F, Shpirt A, Torgov V, Kolotyrkina N, Zinin A, Kasimova A, Perepelov A, Shneider M, Knirel Y "Immune Response to Conjugates of Fragments of the Type K9 Capsular Polysaccharide of Acinetobacter baumannii with Carrier Proteins" - Microbiology Spectrum 10(5) (2022) e0167422
  

  The clonal bacterial species Acinetobacter baumannii is an emerging multidrug-resistant pathogen which causes high-lethality infections. Cells of A. baumannii are surrounded by the type-specific capsular polysaccharide (CPS), which provides resistance to the protective mechanisms of the host and is considered a target for immunization. The conjugates of three inert carrier proteins and A. baumannii type K9 CPS fragments, which contained various numbers of oligosaccharide repeats (K-units), were synthesized by periodate oxidation and squaric acid chemistry. The conjugates were applied to immunize mice, and chemical synthesis by squaric acid was shown to significantly improve the immunogenic properties of glycoconjugate. In BALB/c mice, IgG antibodies were predominant among type K9 CPS reactive antibodies, and their total content was several times higher than that of IgM. Immune sera were characterized by their opsonization ability during practically the entire lives of the experimental mice. The sera were cross-reactive, but the highest specificity was observed against the antigen (type K9 CPS) used for immunization. The immunization of BALB/c and ICR-1 mice with a glycoconjugate without adjuvants led to varying degrees of stimulation of IL-10, IL-17A, and TNF-alpha production, but not IL-4 production in the ICR-1 mice. This is in contrast to the BALB/c mice, in which gamma-IFN production was also activated. The protective effectiveness of the glycoconjugates obtained by squaric acid chemistry was demonstrated by experiments that involved challenging immunized and nonimmunized animals with a lethal dose of A. baumannii K9. IMPORTANCE Immunization by glycoconjugates with A. baumannii type K9 CPS fragments induced a high level of antibodies (predominantly IgG) in sera, which reacted specifically with the CPS of A. baumannii type K9, as well as a long immunological memory. The sera of immunized animals efficiently opsonized A. baumannii type K9. Immunization resulted in the balanced production of pro/anti-inflammatory lymphokines and protective antibodies to ensure the survival of the mice infected with A. baumannii. The level of specific antibodies was sufficient to provide protective immunity against the challenge by A. baumannii, making this approach applicable in the development of vaccine preparations.

  carbohydrates, carbohydrate, Acinetobacter baumannii, capsular polysaccharide, glycoconjugate, immunochemistry, interleukins, opsonisation assay

  NCBI PubMed ID: 35980044
  Publication DOI: 10.1128/spectrum.01674-22
  Journal NLM ID: 101634614
  Publisher: Washington, DC: ASM Press
  Correspondence: N.Rudenko
  Institutions: Laboratory of Immunochemistry, Pushchino Branch, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Pushchino, Moscow Region, Russia, Laboratory of Carbohydrates and Biocides, N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences, Moscow, Russia, Laboratory of Molecular Bioengineering, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russia
  Methods: periodate oxidation, EIA, chemical synthesis, MALDI-TOF MS, statistical analysis, immunization, HR-ESI-MS, opsonization assay
  
   
  
  Acinetobacter baumannii AYE
  CSDB ID 9189 (all data & tools)