Mild acid hydrolysis of the lipopolysaccharide of Escherichia coli O54 afforded an O-polysaccharide, which was studied by sugar analysis, solvolysis with anhydrous trifluoroacetic acid, and 1H and 13C NMR spectroscopy. Solvolysis cleaved predominantly the linkage of β-d-Ribf and, to a lesser extent, that of β-d-GlcpNAc, whereas the other linkages, including the linkage of α-l-Rhap, were stable under selected conditions (40 °C, 5 h). The following structure of the O-polysaccharide was established: →4)-α-d-GalpA-(1 → 2)-α-l-Rhap-(1 → 2)-β-d-Ribf-(1 → 4)-β-d-Galp-(1 → 3)-β-d-GlcpNAc-(1→ The O-antigen gene cluster of E. coli O54 was analyzed and found to be consistent in general with the O-polysaccharide structure established but there were two exceptions: i) in the cluster, there were genes for phosphoserine phosphatase and serine transferase, which have no apparent role in the O-polysaccharide synthesis, and ii) no ribofuranosyltransferase gene was present in the cluster. Both uncommon features are shared by some other enteric bacteria.

O-antigen, Escherichia coli, O-polysaccharide, bacterial polysaccharide structure, O-antigen gene cluster

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6
2,2,4,3	bDGalp	104.7	72.0	73.7	69.8	76.5	62.3
2,2,4,2	Ac	176.1-176.2	23.7
2,2,4	bDGlcpN	103.2	56.2	83.5	70.1	76.4	62.5
2,2	aDGalpA	98.8	69.6	70.9	80.7	73.0	176.6
2	aLRhap	96.9	77.2	70.7	72.9	70.7	17.7
	bDRibp	93.7	77.2	67.8	68.1	64.1


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6
2,2,4,3	bDGalp	4.45	3.53	3.65	3.92	3.71	3.76-3.88
2,2,4,2	Ac	-	2.03-2.05
2,2,4	bDGlcpN	4.68	3.85	3.79	3.50	3.39	3.71-3.86
2,2	aDGalpA	5.07	3.73	4.06	4.38	4.65	-
2	aLRhap	5.08	4.06	3.91	3.50	3.92	1.29
	bDRibp	4.98	3.58	4.29	3.86	3.71-3.84


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6
2,2,4,3	bDGalp	104.7/4.45	72.0/3.53	73.7/3.65	69.8/3.92	76.5/3.71	62.3/3.76-3.88
2,2,4,2	Ac		23.7/2.03-2.05
2,2,4	bDGlcpN	103.2/4.68	56.2/3.85	83.5/3.79	70.1/3.50	76.4/3.39	62.5/3.71-3.86
2,2	aDGalpA	98.8/5.07	69.6/3.73	70.9/4.06	80.7/4.38	73.0/4.65	
2	aLRhap	96.9/5.08	77.2/4.06	70.7/3.91	72.9/3.50	70.7/3.92	17.7/1.29
	bDRibp	93.7/4.98	77.2/3.58	67.8/4.29	68.1/3.86	64.1/3.71-3.84

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

CC(=O)N[C@H]1[C@H](O[C@H]2[C@@H](C(=O)O)O[C@H](O[C@H]3[C@H](O[C@H]4[C@H](O)OC[C@@H](O)[C@H]4O)O[C@@H](C)[C@H](O)[C@H]3O)[C@H](O)[C@H]2O)O[C@H](CO)[C@@H](O)[C@@H]1O[C@@H]1O[C@H](CO)[C@H](O)[C@H](O)[C@H]1O

837.731 g/mol (C₃₁H₅₁NO₂₅)