A capsular polysaccharide was isolated from cells of opportunistic bacterial pathogen Acinetobacter baumannii strain MAR 55-66. According to carbohydrate analysis and 1H and 13C NMR spectroscopy, it is composed of heptasaccharide repeating units, which include five l-rhamnose (Rha) residues and one residue each of d-glucuronic acid (GlcA) and N-acetyl-d-glucosamine (GlcNAc). For structural analysis of this polysaccharide, the Smith degradation, depolymerization with a recombinant endo-glycosidase from a specific bacteriophage, and selective solvolysis with CF3CO2H were applied. The two last methods resulted in formation of the same linear heptasaccharide, which was subjected to the following chemical modifications: borohydride reduction, β-elimination with alkali, and partial acid hydrolysis. The resulting oligosaccharides were isolated by gel-permeation chromatography, and their structures were established by 1H and 13C NMR spectroscopy including 2D 1H-1H and 1H-13C NMR correlation experiments, as well as high-resolution electrospray ionization mass spectrometry. Based on the data obtained, the structure of the branched heptasaccharide repeating unit containing three Rha residues and one GlcNAc residue in the main chain along with two Rha residues and one GlcA residue in the side chain was established. The polysaccharide studied belongs to a group of structurally similar capsular polysaccharides of A. baumannii, which are built up of branched oligosaccharide repeating units that include four or five l-Rha residues and one residue each of d-GlcA and d-GlcNAc.

oligosaccharide, structure, capsular polysaccharide, Acinetobacter baumannii, depolymerization

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6
3,3,2,2,4,3	aLRhap	103.6	71.5	71.6	73.5	70.2	17.7-18.1
3,3,2,2,4	aLRhap	102.2	71.5	79.3	72.5	70.6	17.7-18.1
3,3,2,2	bDGlcpA	104.9	74.8	75.4	80.5	75.4	174.0
3,3,2	aLRhap	102.4	79.5	81.2	72.1	70.3	17.7-18.1
3,3	aLRhap	102.1	79.3	71.3	73.4	70.3	17.7-18.1
3	aLRhap	102.5	71.9	78.7	72.9	70.5	17.7-18.1
2	Ac	175.1-175.5	23.3-23.5
	bDGlcpN	103.4	56.8	82.8	69.6	77.1	62.1


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6
3,3,2,2,4,3	aLRhap	5.02	4.06	3.83	3.46	3.79	1.26-1.32
3,3,2,2,4	aLRhap	4.74	3.98	3.78	3.52	4.04	1.26-1.32
3,3,2,2	bDGlcpA	4.79	3.38	3.63	3.66	3.98	-
3,3,2	aLRhap	5.17	4.40	3.91	3.56	3.72	1.26-1.32
3,3	aLRhap	5.14	4.04	3.90	3.54	3.76	1.26-1.32
3	aLRhap	4.86	3.85	3.78	3.56	4.01	1.26-1.32
2	Ac	-	2.03-2.06
	bDGlcpN	4.73	3.87	3.64	3.57	3.49	3.78-3.95


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6
3,3,2,2,4,3	aLRhap	103.6/5.02	71.5/4.06	71.6/3.83	73.5/3.46	70.2/3.79	17.7-18.1/1.26-1.32
3,3,2,2,4	aLRhap	102.2/4.74	71.5/3.98	79.3/3.78	72.5/3.52	70.6/4.04	17.7-18.1/1.26-1.32
3,3,2,2	bDGlcpA	104.9/4.79	74.8/3.38	75.4/3.63	80.5/3.66	75.4/3.98	
3,3,2	aLRhap	102.4/5.17	79.5/4.40	81.2/3.91	72.1/3.56	70.3/3.72	17.7-18.1/1.26-1.32
3,3	aLRhap	102.1/5.14	79.3/4.04	71.3/3.90	73.4/3.54	70.3/3.76	17.7-18.1/1.26-1.32
3	aLRhap	102.5/4.86	71.9/3.85	78.7/3.78	72.9/3.56	70.5/4.01	17.7-18.1/1.26-1.32
2	Ac		23.3-23.5/2.03-2.06
	bDGlcpN	103.4/4.73	56.8/3.87	82.8/3.64	69.6/3.57	77.1/3.49	62.1/3.78-3.95

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

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

1110.028 g/mol (C₄₄H₇₁R₂NO₃₁, R = next and previous repeats, not counted in mol weight)