The structure of the O-antigen polysaccharide (PS) from Escherichia coli O74 has been determined. Component analysis, together with (1)H and (13)C NMR spectroscopy as well as (1)H,(15)N-HSQC experiments were employed to elucidate the structure. Inter-residue correlations were determined by (1)H,(1)H-NOESY and (1)H,(13)C-heteronuclear multiple-bond correlation experiments. The PS is composed of tetrasaccharide repeating units with the following structure: Cross-peaks of low intensity from an α-linked N-acetylglucosamine residue were present in the NMR spectra, and spectral analysis indicates that they originate from the penultimate residue in the polysaccharide. Consequently, the biological repeating unit has a 3-substituted N-acetyl-d-glucosamine residue at its reducing end. The (1)H, (13)C and (15)N NMR chemical shifts of the α- and β-anomeric forms of d-Fucp3NAc are also reported. The repeating unit of the E. coli O74 O-antigen is identical to that of the capsular polysaccharide from E. coli K45.

Lipopolysaccharide, NMR, Escherichia coli, capsular polysaccharide, biological repeating unit

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6
3,4,2	Ac	175.5	23.2
3,4,3,3	Ac	175.1	22.8
3,4,3	bDFucp3N	104.6	68.8	55.6	70.5	72.6	16.3
3,4	aDGlcpN	100.1	53.1	81.3	68.9	71.4	69.1
3	bDGalpA	104.3	70.9	72.8	79.9	75.2	174.0
2	Ac	175.5	23.2
	bDGlcpN	102.4	55.6	84.0	69.7	76.4	61.6


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6
3,4,2	Ac	-	2.04
3,4,3,3	Ac	-	2.05
3,4,3	bDFucp3N	4.46	3.52	3.92	3.73	3.89	1.25
3,4	aDGlcpN	4.91	4.03	3.86	3.58	4.28	3.84-4.10
3	bDGalpA	4.49	3.59	3.79	4.29	4.09	-
2	Ac	-	2.07
	bDGlcpN	4.59	3.90	3.83	3.61	3.52	3.83-3.97


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6
3,4,2	Ac		23.2/2.04
3,4,3,3	Ac		22.8/2.05
3,4,3	bDFucp3N	104.6/4.46	68.8/3.52	55.6/3.92	70.5/3.73	72.6/3.89	16.3/1.25
3,4	aDGlcpN	100.1/4.91	53.1/4.03	81.3/3.86	68.9/3.58	71.4/4.28	69.1/3.84-4.10
3	bDGalpA	104.3/4.49	70.9/3.59	72.8/3.79	79.9/4.29	75.2/4.09	
2	Ac		23.2/2.07
	bDGlcpN	102.4/4.59	55.6/3.90	84.0/3.83	69.7/3.61	76.4/3.52	61.6/3.83-3.97

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

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

769.707 g/mol (C₃₀H₄₇R₂N₃O₂₀, R = next and previous repeats, not counted in mol weight)