Morganella morganii is a commensal Gram-negative bacterium that has long been known to produce an antigen bearing phosphocholine groups. We determined the structure of this O-chain antigen and found that its repeating unit also contains a free amino group and a second phosphate: This alternating charge character places the M. morganii O-chain polysaccharide into a small family of zwitterionic polysaccharides (ZPSs) known to induce T-cell-dependent immune responses via presentation by class II major histocompatibility complex (MHCII) molecules. In vitro binding assays demonstrate that this O-chain interacts with MHCII in a manner that competes with binding of the prototypical ZPS antigen PSA from Bacteroides fragilis, despite its lack of a helical structure. Cellular studies also showed that the M. morganii polysaccharide induces activation of CD4(+) T-cells. Antibody binding experiments using acid hydrolyzed fragments representing the monomer and higher oligomers of the repeating unit showed that the phosphocholine group was the dominant element of the epitope with an overall affinity (K(D)) of about 5 ? 10(-5) M, a typical value for an IgM anti-carbohydrate antibody but much lower than the affinity for phosphocholine itself. These data show that the structure of the M. morganii polysaccharide contains a unique zwitterionic repeating unit which allows for immune recognition by T-cells, making it the first identified T-cell-dependent O-chain antigen.

polysaccharide structure, Morganella morganii, MHC interaction, phosphocholine antigen, zwitterionic polysaccharide

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6
0,3,3,2,0	xXCho	60.58	67.19	55.05
0,3,3,2	P
0,3,3,3	aDGalpN	91.80	51.18	67.64	69.37	72.47	61.69
0,3,3	bDGalp	103.57	74.34	74.34	70.32	75.67	62.38
0,3,2	Ac	175.4	23.35
0,3	bDGalpN	103.67	52.47	78.12	69.23	75.85	61.92
0	x?Gro	71.67	70.31	67.52
	P


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6
0,3,3,2,0	xXCho	4.34	3.66	3.24
0,3,3,2	P
0,3,3,3	aDGalpN	5.55	3.56	4.35	4.04	4.34	3.77
0,3,3	bDGalp	4.75	4.24	4.04	4.77	3.76	3.77
0,3,2	Ac	-	2.07
0,3	bDGalpN	4.46	4.01	4.05	4.15	3.73	3.79-3.83
0	x?Gro	3.99	4.01	3.67
	P


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6
0,3,3,2,0	xXCho	60.58/4.34	67.19/3.66	55.05/3.24
0,3,3,2	P
0,3,3,3	aDGalpN	91.80/5.55	51.18/3.56	67.64/4.35	69.37/4.04	72.47/4.34	61.69/3.77
0,3,3	bDGalp	103.57/4.75	74.34/4.24	74.34/4.04	70.32/4.77	75.67/3.76	62.38/3.77
0,3,2	Ac		23.35/2.07
0,3	bDGalpN	103.67/4.46	52.47/4.01	78.12/4.05	69.23/4.15	75.85/3.73	61.92/3.79-3.83
0	x?Gro	71.67/3.99	70.31/4.01	67.52/3.67
	P

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

*O[C@@H]1[C@H](O[C@H]2O[C@H](CO)[C@H](O)[C@H](O)[C@H]2N)[C@@H](OP(=O)(O)OCC[N+](C)(C)C)[C@H](O[C@H]2[C@@H](O)[C@@H](CO)O[C@@H](OCC(O)COP(*)(=O)O)[C@@H]2NC(C)=O)O[C@@H]1CO

846.687 g/mol (C₂₈H₅₄R₂N₃O₂₂P₂⁺, R = next and previous repeats, not counted in mol weight)