Shigella flexneri is an important human pathogen causing shigellosis. Strains of S. flexneri are serologically heterogeneous and, based on O-antigens, are currently classified into 14 types. Structures of the O-antigens (O-polysaccharides) of S. flexneri have been under study since 1960s but some gaps still remained. In this work, using one- and two-dimensional (1) H- and (13) C-NMR spectroscopy, the O-polysaccharides of several S. flexneri types were reinvestigated, and their structures were either confirmed (types 2b, 3b, 3c, 5b, X) or amended in respect to the O-acetylation pattern (types 3a, Y, 6, 6a). As a result, the O-acetylation sites were defined in all O-polysaccharides that had not been studied in detail earlier, and the long story of S. flexneri type strain O-antigen structure elucidation is thus completed. New and published data on the S. flexneri O-antigen structures are summarized and discussed in view of serological and genetic relationships of the O-antigens within the Shigella group and between S. flexneri and Escherichia coli.

Shigella flexneri, O-acetylation, serological classification, O-Polysaccharide structure, O-antigen diversity

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6
3,4,2,3	60%Ac	175.1	22.0
3,4,2,4	30%Ac	175.1	21.9
3,4,2	aLRhap	102.4	78.0	74.2	71.4	70.6	18.0
3,4	aLRhap	101.0	80.3	71.3	73.4	70.6	18.0
3	bDGalpA	105.7	71.4	75.1	77.6	76.0	174.2
2	Ac	175.9	23.8
	bDGalpN	104.2	53.0	81.2	68.9	74.5	62.4


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6
3,4,2,3	60%Ac	-	2.19
3,4,2,4	30%Ac	-	2.14
3,4,2	aLRhap	5.16	4.25	5.04	3.53	3.77	1.27
3,4	aLRhap	5.38	4.09	3.87	3.44	3.67	1.24
3	bDGalpA	4.50	3.63	4.23	4.34	3.62	-
2	Ac	-	2.06
	bDGalpN	4.51	4.00	3.90	4.24	3.82	3.77-3.77


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6
3,4,2,3	60%Ac		22.0/2.19
3,4,2,4	30%Ac		21.9/2.14
3,4,2	aLRhap	102.4/5.16	78.0/4.25	74.2/5.04	71.4/3.53	70.6/3.77	18.0/1.27
3,4	aLRhap	101.0/5.38	80.3/4.09	71.3/3.87	73.4/3.44	70.6/3.67	18.0/1.24
3	bDGalpA	105.7/4.50	71.4/3.63	75.1/4.23	77.6/4.34	76.0/3.62	
2	Ac		23.8/2.06
	bDGalpN	104.2/4.51	53.0/4.00	81.2/3.90	68.9/4.24	74.5/3.82	62.4/3.77-3.77

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

*O[C@H]1[C@H](O[C@H]2[C@H](O[C@H]3[C@@H](C(=O)O)O[C@@H](O[C@@H]4[C@@H](NC(C)=O)[C@H](*)O[C@H](CO)[C@@H]4O)[C@H](O)[C@H]3O)O[C@@H](C)[C@H](O)[C@H]2O)O[C@@H](C)[C@H](OC(C)=O)[C@H]1OC(C)=O

755.676 g/mol (C₃₀H₄₅R₂NO₂₁, R = next and previous repeats, not counted in mol weight)