O-antigen representing the O-polysaccharide chain of the lipopolysaccharide is the most variable constituent on the cell surface of Gram-negative bacteria and a player in their pathogenicity. The O-polysaccharide of Escherichia coli O109 was studied by sugar analysis and nuclear magnetic resonance spectroscopy and found to contain a rarely occurring monosaccharide, 2,3-diacetamido-2,3,6-trideoxy-l-mannose (l-RhaNAc3NAc). The following structure of the tetrasaccharide repeating unit of the O-polysaccharide was established, which is closely related to that of Proteus penneri O66: Ac--4-β-L-RhapNAc3NAc→4)-α-D-Glcp-(1→3)-α-L-6dTalp-(1→3)-β-D-GlcpNAc-(1→. The O-antigen gene cluster of E. coli O109 was sequenced and all 14 genes found were assigned functions based on their similarity to genes from the available databases. Putative genes for synthesis of l-RhaN3N were found in E. coli O109 and their homologues in E. coli O119, whose O-antigen has been reported earlier to contain 2-acetamido-2,3,6-trideoxy-3-formamido-d-mannose (d-RhaNAc3NFo). Analysis by GLC of the (S)-2-octyl glycosides confirmed that the absolute configuration of RhaN3N in E. coli O119 should be revised from D TO L.

O-antigen, Escherichia coli, 2, 3, 3-diamino-2, O-antigen gene cluster, 6-trideoxy-L-mannose

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6
3,3,3,2	Ac	176.7	23.5
3,3,3,3	Ac	175.5	23.2
3,3,3,4	Ac	174.8	21.6
3,3,3	bLRhapN3N	100.7	53.0	53.4	72.9	73.1	18.2
3,3	aDGlcp	96.8	71.0	79.9	76.5	72.9	61.3
3,2	Ac	174.4	21.9
3	aL6dTalp	101.0	68.2	70.9	71.4	68.7	16.8
2	Ac	173.0	23.7
	bDGlcpN	101.4	57.5	83.4	70.3	77.8	62.2


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6
3,3,3,2	Ac	-	2.08
3,3,3,3	Ac	-	1.91
3,3,3,4	Ac	-	2.09
3,3,3	bLRhapN3N	5.21	4.65	4.26	4.70	3.75	1.23
3,3	aDGlcp	5.07	3.55	4.01	3.82	3.71	3.65-3.80
3,2	Ac	-	2.13
3	aL6dTalp	4.96	5.12	4.15	3.86	4.26	1.22
2	Ac	-	2.03
	bDGlcpN	4.51	3.83	3.61	3.54	3.40	3.81-3.88


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6
3,3,3,2	Ac		23.5/2.08
3,3,3,3	Ac		23.2/1.91
3,3,3,4	Ac		21.6/2.09
3,3,3	bLRhapN3N	100.7/5.21	53.0/4.65	53.4/4.26	72.9/4.70	73.1/3.75	18.2/1.23
3,3	aDGlcp	96.8/5.07	71.0/3.55	79.9/4.01	76.5/3.82	72.9/3.71	61.3/3.65-3.80
3,2	Ac		21.9/2.13
3	aL6dTalp	101.0/4.96	68.2/5.12	70.9/4.15	71.4/3.86	68.7/4.26	16.8/1.22
2	Ac		23.7/2.03
	bDGlcpN	101.4/4.51	57.5/3.83	83.4/3.61	70.3/3.54	77.8/3.40	62.2/3.81-3.88

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

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

823.799 g/mol (C₃₄H₅₃R₂N₃O₂₀, R = next and previous repeats, not counted in mol weight)