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MetaCyc Pathway: CMP-legionaminate biosynthesis I
Inferred from experiment

Enzyme View:

Pathway diagram: CMP-legionaminate biosynthesis I

This view shows enzymes only for those organisms listed below, in the list of taxa known to possess the pathway. If an enzyme name is shown in bold, there is experimental evidence for this enzymatic activity.

Synonyms: CMP-legionaminic acid biosynthesis I, 5,7-diacetamido-3,5,7,9-tetradeoxy-D-glycero-D-galacto-nonulosonic acid biosynthesis I, CMP-N, N-diacetyllegionaminate biosynthesis I

Superclasses: BiosynthesisCarbohydrates BiosynthesisSugars BiosynthesisSugar Nucleotides BiosynthesisCMP-sugar BiosynthesisCMP-legionaminate biosynthesis

Some taxa known to possess this pathway include : Campylobacter jejuni, Campylobacter jejuni jejuni NCTC 11168 = ATCC 700819

Expected Taxonomic Range: Bacteria

General Background

5,7-diacetamido-3,5,7,9-tetradeoxy-D-glycero-D-galacto-nonulopyranosonate (legionaminic acid, 5,7-diacetamido-3,5,7,9-tetradeoxy-D-glycero-D-galacto-nonulosonic acid) is part of the diverse sialic acid family of α-keto sugars. Members of this family mediate or modulate many cellular interactions in prokaryotes and eukaryotes. Many pathogenic organisms can display, bind, or catabolize sialic acids, helping them to evade host immune responses and invade cells (in [Schoenhofen09]).

Bacteria incorporate sialic acid-like 5,7-diacetamido-3,5,7,9-tetradeoxy-nonulosonate derivatives into virulence-associated cell surface glycoconjugates which may include lipopolysaccharide (LPS), capsular polysaccharide, pili and flagella. These nonulosonate derivatives are unique to microorganisms and may show configurational differences as compared with sialic acid. 5,7-diacetamido-3,5,7,9-tetradeoxy-D-glycero-D-galacto-nonulopyranosonate is a nonulosonate derivative that shows the same D-glycero-D-galacto absolute configuration as sialic acid ( N-acetylneuraminate) (in [Schoenhofen09]). It was first identified in 1994 as a component of Legionella pneumophila serogroup 1 LPS [Knirel94]. Its correct stereochemistry was determined in 2001 [Tsvetkov01]. Legionella pneumophila serogroup 1 is the causative agent of Legionnaires' disease.

The gastrointestinal pathogen Campylobacter coli was shown to have novel 5,7-diacetamido-3,5,7,9-tetradeoxy-D-glycero-D-galacto-nonulopyranosonate derivatives O-linked to serine/threonine residues of flagellin, and genes critical to CMP-N,N'-diacetyllegionaminate biosynthesis were identified [McNally07]. In the gastrointestinal pathogen Campylobacter jejuni jejuni NCTC 11168 = ATCC 700819 the complete biosynthetic pathway for CMP-N,N'-diacetyllegionaminate has been elucidated and was shown to involve genes in the Campylobacter flagellin glycosylation locus [Schoenhofen09] (this pathway).

About This Pathway

The elucidation of this pathway was based on a combination of bioinformatic, comparative genomic, metabolomic, and functional analyses as discussed in [Schoenhofen09]. Eleven recombinant enzymes involved in the biosynthesis of CMP-N,N'-diacetyllegionaminate (CMP-legionaminic acid) the CMP-activated form of 5,7-diacetamido-3,5,7,9-tetradeoxy-D-glycero-D-galacto-nonulopyranosonate (legionaminic acid) in Campylobacter jejuni jejuni NCTC 11168 = ATCC 700819 were purified and characterized and a biosynthetic pathway of CMP-N,N'-diacetyllegionaminate was proposed. The work also provided a method for the large-scale production of 5,7-diacetamido-3,5,7,9-tetradeoxy-D-glycero-D-galacto-nonulopyranosonate [Schoenhofen09]. The pathway involves unique GDP-sugar intermediates, rather than the UDP-sugar intermediates found in other pathways (such as pathway CMP-pseudaminate biosynthesis), which may provide a means of independent pathway regulation (in [Schoenhofen09]).

In this pathway PtmF and PtmA were found to efficiently convert β-D-fructofuranose 6-phosphate to D-glucosamine 6-phosphate. The isomerase PtmF was stabilized by copurification with glutaminase PtmA, and together they constituted an unfused D-glucosamine-6-phosphate synthase. The next committed step is catalyzed by a phosphoglucosamine mutase PgmL in a reversible reaction. PtmE is a nucleotidyltransferase with an absolute specificity for GTP when D-glucosamine 1-phosphate is the substrate (this allowed large-scale purification of GDP-D-glucosamine). In "one pot" enzymatic reactions, PtmF, PtmA, PgmL and PtmE converted β-D-fructofuranose 6-phosphate to GDP-D-glucosamine. Metabolites were purified and identified by capillary electrophoresis/mass spectrometry analysis and NMR spectroscopy [Schoenhofen09].

Although still uncertain, the next step was proposed to involve a GlmU catalyzed acetylation of GDP-D-glucosamine to GDP-N-acetyl-D-glucosamine. The enzyme is known to be a bifunctional glucosamine-1-phosphate N-acetyltransferase/UDP-N-acetylglucosamine pyrophosphorylase. The enzyme converted GDP-D-glucosamine to GDP-N-acetyl-D-glucosamine, but not to completion. The authors stated that other candidate enzymes for this reaction are being screened [Schoenhofen09].

The data and rationale used in elucidating the remaining biosynthetic route to 5,7-diacetamido-3,5,7,9-tetradeoxy-D-glycero-D-galacto-nonulopyranosonate (legionaminic acid) are detailed in [Schoenhofen09]. Large scale biosynthesis of this compound could be achieved using two separate "one pot" reactions converting GDP-N-acetyl-D-glucosamine to GDP-2,4-diacetamido-2,4,6-trideoxy-α-D-glucopyranose, and this product to 5,7-diacetamido-3,5,7,9-tetradeoxy-D-glycero-D-galacto-nonulopyranosonate. LegF could efficiently catalyze production of CMP-N,N'-diacetyllegionaminate (CMP-legionaminic acid) [Schoenhofen09].

Variants: CMP-legionaminate biosynthesis II

Created 11-Mar-2011 by Fulcher CA, SRI International


Angata02: Angata T, Varki A (2002). "Chemical diversity in the sialic acids and related alpha-keto acids: an evolutionary perspective." Chem Rev 102(2);439-69. PMID: 11841250

Cox02: Cox AD, Hood DW, Martin A, Makepeace KM, Deadman ME, Li J, Brisson JR, Moxon ER, Richards JC (2002). "Identification and structural characterization of a sialylated lacto-N-neotetraose structure in the lipopolysaccharide of Haemophilus influenzae." Eur J Biochem 269(16);4009-19. PMID: 12180977

Hood99: Hood DW, Makepeace K, Deadman ME, Rest RF, Thibault P, Martin A, Richards JC, Moxon ER (1999). "Sialic acid in the lipopolysaccharide of Haemophilus influenzae: strain distribution, influence on serum resistance and structural characterization." Mol Microbiol 33(4);679-92. PMID: 10447878

Knirel94: Knirel YA, Rietschel ET, Marre R, Zahringer U (1994). "The structure of the O-specific chain of Legionella pneumophila serogroup 1 lipopolysaccharide." Eur J Biochem 221(1);239-45. PMID: 8168511

McNally07: McNally DJ, Aubry AJ, Hui JP, Khieu NH, Whitfield D, Ewing CP, Guerry P, Brisson JR, Logan SM, Soo EC (2007). "Targeted metabolomics analysis of Campylobacter coli VC167 reveals legionaminic acid derivatives as novel flagellar glycans." J Biol Chem 282(19);14463-75. PMID: 17371878

Schoenhofen09: Schoenhofen IC, Vinogradov E, Whitfield DM, Brisson JR, Logan SM (2009). "The CMP-legionaminic acid pathway in Campylobacter: biosynthesis involving novel GDP-linked precursors." Glycobiology 19(7);715-25. PMID: 19282391

Severi05: Severi E, Randle G, Kivlin P, Whitfield K, Young R, Moxon R, Kelly D, Hood D, Thomas GH (2005). "Sialic acid transport in Haemophilus influenzae is essential for lipopolysaccharide sialylation and serum resistance and is dependent on a novel tripartite ATP-independent periplasmic transporter." Mol Microbiol 58(4);1173-85. PMID: 16262798

Severi08: Severi E, Muller A, Potts JR, Leech A, Williamson D, Wilson KS, Thomas GH (2008). "Sialic acid mutarotation is catalysed by the Escherichia coli beta -propeller protein YJHT." J Biol Chem 283(8):4841-9. PMID: 18063573

Tsvetkov01: Tsvetkov YE, Shashkov AS, Knirel YA, Zahringer U (2001). "Synthesis and identification in bacterial lipopolysaccharides of 5,7-diacetamido-3,5,7,9-tetradeoxy-D-glycero-D-galacto- and -D-glycero-D-talo-non-2-ulosonic acids." Carbohydr Res 331(3);233-7. PMID: 11383892

Other References Related to Enzymes, Genes, Subpathways, and Substrates of this Pathway

Badet87: Badet B, Vermoote P, Haumont PY, Lederer F, LeGoffic F (1987). "Glucosamine synthetase from Escherichia coli: purification, properties, and glutamine-utilizing site location." Biochemistry 1987;26(7);1940-8. PMID: 3297136

Badet88: Badet B, Vermoote P, Le Goffic F (1988). "Glucosamine synthetase from Escherichia coli: kinetic mechanism and inhibition by N3-fumaroyl-L-2,3-diaminopropionic derivatives." Biochemistry 1988;27(7);2282-7. PMID: 3132968

BadetDenisot92: Badet-Denisot MA, Badet B (1992). "Chemical modification of glucosamine-6-phosphate synthase by diethyl pyrocarbonate: evidence of histidine requirement for enzymatic activity." Arch Biochem Biophys 1992;292(2);475-8. PMID: 1731613

BadetDenisot95: Badet-Denisot MA, Leriche C, Massiere F, Badet B (1995). "Nitrogen transfer in E. coli glucosamine-6P synthase. Investigations using substrate and bisubstrate analogs." Bioorg. Med. Chem. Lett. 5(8);815-820.

Bearne00: Bearne SL, Blouin C (2000). "Inhibition of Escherichia coli glucosamine-6-phosphate synthase by reactive intermediate analogues. The role of the 2-amino function in catalysis." J Biol Chem 275(1);135-40. PMID: 10617596

Bearne95: Bearne SL, Wolfenden R (1995). "Glutamate gamma-semialdehyde as a natural transition state analogue inhibitor of Escherichia coli glucosamine-6-phosphate synthase." Biochemistry 34(36);11515-20. PMID: 7547881

Bearne96: Bearne SL (1996). "Active site-directed inactivation of Escherichia coli glucosamine-6-phosphate synthase. Determination of the fructose 6-phosphate binding constant using a carbohydrate-based inactivator." J Biol Chem 271(6);3052-7. PMID: 8621700

BRENDA14: BRENDA team (2014). Imported from BRENDA version existing on Aug 2014.

Broschat02: Broschat KO, Gorka C, Page JD, Martin-Berger CL, Davies MS, Huang Hc HC, Gulve EA, Salsgiver WJ, Kasten TP (2002). "Kinetic characterization of human glutamine-fructose-6-phosphate amidotransferase I: potent feedback inhibition by glucosamine 6-phosphate." J Biol Chem 277(17);14764-70. PMID: 11842094

Chmara84: Chmara H, Zahner H (1984). "The inactivation of glucosamine synthetase from bacteria by anticapsin, the C-terminal epoxyamino acid of the antibiotic tetaine." Biochim Biophys Acta 787(1);45-52. PMID: 6426523

Chmara85: Chmara H (1985). "Inhibition of glucosamine synthase by bacilysin and anticapsin." J Gen Microbiol 131(2);265-71. PMID: 3884731

Deng06: Deng MD, Grund AD, Wassink SL, Peng SS, Nielsen KL, Huckins BD, Burlingame RP (2006). "Directed evolution and characterization of Escherichia coli glucosamine synthase." Biochimie 88(5);419-29. PMID: 16871653

DutkaMalen88: Dutka-Malen S, Mazodier P, Badet B (1988). "Molecular cloning and overexpression of the glucosamine synthetase gene from Escherichia coli." Biochimie 70(2);287-90. PMID: 3134953

Edebrink96: Edebrink P, Jansson PE, Bogwald J, Hoffman J (1996). "Structural studies of the Vibrio salmonicida lipopolysaccharide." Carbohydr Res 287(2);225-45. PMID: 8766209

Floquet07: Floquet N, Mouilleron S, Daher R, Maigret B, Badet B, Badet-Denisot MA (2007). "Ammonia channeling in bacterial glucosamine-6-phosphate synthase (Glms): molecular dynamics simulations and kinetic studies of protein mutants." FEBS Lett 581(16);2981-7. PMID: 17559838

Ghosh60: Ghosh S, Blumenthal HJ, Davidson E, Roseman S (1960). "Glucosamine metabolism. V. Enzymatic synthesis of glucosamine 6-phosphate." J Biol Chem 235;1265-73. PMID: 13827775

Ghosh60a: Ghosh S, Blumenthal HJ, Davidson E, Roseman S (1960). "Glucosamine Metabolism V. Enzymatic synthesis of glucosamine 6-phosphate." J Biol Chem 1960;235(5):1265-1273.

Glaze08: Glaze PA, Watson DC, Young NM, Tanner ME (2008). "Biosynthesis of CMP-N,N'-diacetyllegionaminic acid from UDP-N,N'-diacetylbacillosamine in Legionella pneumophila." Biochemistry 47(10);3272-82. PMID: 18275154

GolinelliPimpan91: Golinelli-Pimpaneau B, Badet B (1991). "Possible involvement of Lys603 from Escherichia coli glucosamine-6-phosphate synthase in the binding of its substrate fructose 6-phosphate." Eur J Biochem 1991;201(1);175-82. PMID: 1915361

Graack01: Graack HR, Cinque U, Kress H (2001). "Functional regulation of glutamine:fructose-6-phosphate aminotransferase 1 (GFAT1) of Drosophila melanogaster in a UDP-N-acetylglucosamine and cAMP-dependent manner." Biochem J 360(Pt 2);401-12. PMID: 11716769

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Please cite the following article in publications resulting from the use of MetaCyc: Caspi et al, Nucleic Acids Research 42:D459-D471 2014
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