Derivatives of 5,7-diamino-3,5,7,9-tetradeoxynon-2-ulosonic acids are essential constituents of some bacterial polysaccharides and glycoproteins. In order to establish reliably the configuration of the natural sugars, nine stereoisomeric 5,7-diacetamido-3,5,7,9-tetradeoxynon-2-ulosonic acids were synthesized, including di-N-acetyl-legionaminic and -pseudaminic acids (the D-glycero-D-galacto and L-glycero-L-manno isomers, respectively) and their isomers at C-4, C-5, C-7, and C-8 having the L-glycero-D-galacto, D-glycero-D-talo, L-glycero-D-talo, D-glycero-L-altro, L-glycero-L-altro, D-glycero-L-manno, and L-glycero-L-gluco configurations. Synthesis was performed by condensation of 2,4-diacetamido-2,4,6-trideoxy-L-gulose, -D-mannose, -D-talose, and -L-allose with oxalacetic acid under basic conditions, the reaction of the last two precursors being accompanied by epimerisation at C-2. The 1H and 13C NMR data of the synthetic compounds are discussed. Acetylated methyl esters of the C-7 and C-8 isomeric nonulosonic acids were prepared and used for analysis of the side-chain conformation by NMR spectroscopy.

NMR, synthesis, LPS, Bacterial, identification, acid, NMR spectroscopy, pseudaminic acid, putative, spectroscopy, 5, 7-diamino-3, 7, 9-tetradeoxynon-2-ulosonic acid, lipopolysaccharide components, 7-diacetamido-3, 9-tetradeoxynon-2-ulosonic acids, 2, 4-diacetamido-2, 4, 6-trideoxyhexoses, legionaminic acid, 9-teradeoxynonulosonic acid, component, isomer

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

CC(=O)N[C@H]([C@@H]1OC(O)(C(=O)O)C[C@H](O)[C@H]1NC(C)=O)[C@@H](C)O

334.325 g/mol (C₁₃H₂₂N₂O₈)

Nonulosonic acids are found in the surface polysaccharides of many bacterial species and are often implicated in pathogenesis. Here, the structure of a novel 5,7-diacetamido-3,5,7,9-tetradeoxynon-2-ulosonic acid recovered from the capsular polysaccharide of a multiply antibiotic resistant Acinetobacter baumannii isolate was determined. The isolate carries a sugar synthesis module that differs by only a single gene from the module for the synthesis of 5,7-diacetamido-3,5,7,9-tetradeoxy-L-glycero-L-altro-non-2-ulosonic acid or 5,7-di-N-acetylacinetaminic acid, recently discovered in the capsule of another A. baumannii isolate. The new monosaccharide is the C8-epimer of acinetaminic acid (8eAci; 5,7-diacetamido-3,5,7,9-tetradeoxy-D-glycero-L-altro-non-2-ulosonic acid) and the C7-epimer of legionaminic acid. This monosaccharide had not previously been detected in a biological sample but had been synthesized chemically.

structure, capsular polysaccharide, Acinetobacter baumannii, nonulosonic acid, 5, 7, 7-diacetamido-3, legionaminic acid, capsule, surface polysaccharide, 7-Di-N-acetyl-8-epiacinetaminic acid, 9-tetradeoxy-D-glycero-L-altro-non-2-ulosonic acid

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6	C7	C8	C9
5	Ac
7	Ac
	aX8eAcip	?	?	40.5	68.3	55.0	76.1	53.5	66.5	20.3


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6	H7	H8	H9
5	Ac
7	Ac
	aX8eAcip	-	-	1.86-2.24	3.95	3.84	3.84	3.90	4.45	1.05


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6	C7/H7	C8/H8	C9/H9
5	Ac
7	Ac
	aX8eAcip			40.5/1.86-2.24	68.3/3.95	55.0/3.84	76.1/3.84	53.5/3.90	66.5/4.45	20.3/1.05

The spectrum also has 2 signals at unknown positions (not plotted).

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

CC(=O)N[C@H]([C@@H]1O[C@](O)(C(=O)O)C[C@H](O)[C@H]1NC(C)=O)[C@@H](C)O

334.325 g/mol (C₁₃H₂₂N₂O₈)

Nonulosonic acids are found in the surface polysaccharides of many bacterial species and are often implicated in pathogenesis. Here, the structure of a novel 5,7-diacetamido-3,5,7,9-tetradeoxynon-2-ulosonic acid recovered from the capsular polysaccharide of a multiply antibiotic resistant Acinetobacter baumannii isolate was determined. The isolate carries a sugar synthesis module that differs by only a single gene from the module for the synthesis of 5,7-diacetamido-3,5,7,9-tetradeoxy-L-glycero-L-altro-non-2-ulosonic acid or 5,7-di-N-acetylacinetaminic acid, recently discovered in the capsule of another A. baumannii isolate. The new monosaccharide is the C8-epimer of acinetaminic acid (8eAci; 5,7-diacetamido-3,5,7,9-tetradeoxy-D-glycero-L-altro-non-2-ulosonic acid) and the C7-epimer of legionaminic acid. This monosaccharide had not previously been detected in a biological sample but had been synthesized chemically.

structure, capsular polysaccharide, Acinetobacter baumannii, nonulosonic acid, 5, 7, 7-diacetamido-3, legionaminic acid, capsule, surface polysaccharide, 7-Di-N-acetyl-8-epiacinetaminic acid, 9-tetradeoxy-D-glycero-L-altro-non-2-ulosonic acid

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6	C7	C8	C9
5	Ac
7	Ac
	bX8eAcip	?	?	41.9	69.7	55.5	76.1	54.9	66.8	20.3


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6	H7	H8	H9
5	Ac
7	Ac
	bX8eAcip	-	-	1.64-2.70	3.70	3.78	3.40	3.93	4.42	1.04


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6	C7/H7	C8/H8	C9/H9
5	Ac
7	Ac
	bX8eAcip			41.9/1.64-2.70	69.7/3.70	55.5/3.78	76.1/3.40	54.9/3.93	66.8/4.42	20.3/1.04

The spectrum also has 2 signals at unknown positions (not plotted).

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

CC(=O)N[C@H]([C@@H]1O[C@@](O)(C(=O)O)C[C@H](O)[C@H]1NC(C)=O)[C@@H](C)O

334.325 g/mol (C₁₃H₂₂N₂O₈)

Structures of capsular polysaccharides of Acinetobacter baumannii isolates carrying KL13 and KL73 gene clusters were established. The closely related KL73 and KL13 gene clusters differ only by one gene in the module responsible for synthesis of the non-2-ulosonic acids. The K13 and K73 polysaccharides differ only in a single side-chain sugar, which is either 5,7-diacetamido-3,5,7,9-tetradeoxy-l-glycero-l-altro- or -d-glycero-l-altro-non-2-ulosonic acid [di-N-acetylated forms of acinetaminic acid (Aci5Ac7Ac) or 8-epiacinetaminic acid (8eAci5Ac7Ac), respectively]. The KL13 also is closely related to the KL12 gene cluster, which contains a different wzy gene encoding the K unit polymerase. Accordingly, the otherwise near identical K units are linked differently via an α-d-FucpNAc-(1→4)-d-Galp linkage in K13 and K73 or an α-d-FucpNAc-(1→3)-d-GalpNAc linkage in K12. This finding confirms the predicted substrate of the ItrB3 initiating transferase as d-FucpNAc. Glycosyltransferases predicted to catalyse the linkage of d-Galp or d-GalpNAc to l-FucpNAc in the growing K13 and K73 or K12 units, respectively, differ by only two amino acids.

capsular polysaccharide, Acinetobacter baumannii, capsular polysaccharide structure, 5, 7-Di-N-acetyl-8-epiacinetaminic acid, Capsule biosynthesis gene cluster, Non-2-ulosonic acid 5, 7-Di-N-acetylacinetaminic acid

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6	C7	C8	C9
3,3,6,5	Ac	174.6-175.8	23.3-23.6
3,3,6,7	Ac	174.6-175.8	23.3-23.6
3,3,6	aX8eAcip	?	?	40.6	67.8	54.2	76.7	53.5	66.3	20.2
3,3	aDGalp	102.2	69.5	69.8	78.5	71.1	62.1
3,2	Ac	174.6-175.8	23.3-23.6
3	aLFucpN	100.0	49.5	77.5	72.2	68.3	16.7
2	Ac	174.6-175.8	23.3-23.6
	aDFucpN	99.1	49.8	74.6	72.3	68.2	16.4


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6	H7	H8	H9
3,3,6,5	Ac	-	2.01-2.10
3,3,6,7	Ac	-	2.01-2.10
3,3,6	aX8eAcip	-	-	1.70-2.41	3.88	3.84	3.45	3.92	4.48	1.07
3,3	aDGalp	5.10	3.83	3.98	3.99	4.14	3.39-3.54
3,2	Ac	-	2.01-2.10
3	aLFucpN	5.02	4.32	4.04	3.99	4.16	1.26
2	Ac	-	2.01-2.10
	aDFucpN	4.83	4.37	3.98	3.83	4.60	1.17


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6	C7/H7	C8/H8	C9/H9
3,3,6,5	Ac		23.3-23.6/2.01-2.10
3,3,6,7	Ac		23.3-23.6/2.01-2.10
3,3,6	aX8eAcip			40.6/1.70-2.41	67.8/3.88	54.2/3.84	76.7/3.45	53.5/3.92	66.3/4.48	20.2/1.07
3,3	aDGalp	102.2/5.10	69.5/3.83	69.8/3.98	78.5/3.99	71.1/4.14	62.1/3.39-3.54
3,2	Ac		23.3-23.6/2.01-2.10
3	aLFucpN	100.0/5.02	49.5/4.32	77.5/4.04	72.2/3.99	68.3/4.16	16.7/1.26
2	Ac		23.3-23.6/2.01-2.10
	aDFucpN	99.1/4.83	49.8/4.37	74.6/3.98	72.3/3.83	68.2/4.60	16.4/1.17

The spectrum also has 2 signals at unknown positions (not plotted).

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

[*]O[C@H]1[C@@H](CO[C@@]2(C(=O)O)C[C@H](O)[C@@H](NC(C)=O)[C@H]([C@@H](NC(C)=O)[C@@H](C)O)O2)O[C@H](O[C@H]2[C@H](NC(C)=O)[C@H](O[C@@H]3[C@@H](NC(C)=O)[C@@H]([*])O[C@H](C)[C@@H]3O)O[C@@H](C)[C@H]2O)[C@H](O)[C@H]1O

852.841 g/mol (C₃₅H₅₆R₂N₄O₂₀, R = next and previous repeats, not counted in mol weight)