Bacterial extracellular membrane vesicles (EMVs) play an active role in many physiological and pathogenic processes. Here, we report the identification and the detailed structural characterization of the capsular polysaccharide from both cells and EMVs from Shewanella vesiculosa by NMR and chemical analysis. The polysaccharide consists of a pentasaccharide repeating unit containing neutral monosaccharides together with amino sugars, of which one has never been isolated from a natural source. The adhesion ability of the polymer both on synthetic surfaces, such as polystyrene nanoparticles and on vesicles with a bilayer mimicking the bacterial membrane in the presence and absence of lipopolysaccharide was investigated. In both cases, a 'CPS-corona' that could be the first stage of biofilm formation was observed. The polymer also activates Caspases on colon cancer cells, making S. vesiculosa EMVs as natural nanocarriers for drug delivery.

NMR, capsule, Biofilm, nanoparticles, SANS, Shewanosamine

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6
2	Ac	174.6	22.0
3	Ac	174.4	21.8
4	Ac	175.4	21.8
	aDSugp	90.4	47.6	47.6	51.3	64.8	15.7


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6
2	Ac	-	1.91
3	Ac	-	1.83
4	Ac	-	2.02
	aDSugp	5.14	3.99	4.25	4.18	4.37	1.03


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6
2	Ac		22.0/1.91
3	Ac		21.8/1.83
4	Ac		21.8/2.02
	aDSugp	90.4/5.14	47.6/3.99	47.6/4.25	51.3/4.18	64.8/4.37	15.7/1.03

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

CC(=O)N[C@@H]1[C@@H](NC(C)=O)C(O)O[C@H](C)[C@@H]1NC(C)=O

287.316 g/mol (C₁₂H₂₁N₃O₅)

Bacterial extracellular membrane vesicles (EMVs) play an active role in many physiological and pathogenic processes. Here, we report the identification and the detailed structural characterization of the capsular polysaccharide from both cells and EMVs from Shewanella vesiculosa by NMR and chemical analysis. The polysaccharide consists of a pentasaccharide repeating unit containing neutral monosaccharides together with amino sugars, of which one has never been isolated from a natural source. The adhesion ability of the polymer both on synthetic surfaces, such as polystyrene nanoparticles and on vesicles with a bilayer mimicking the bacterial membrane in the presence and absence of lipopolysaccharide was investigated. In both cases, a 'CPS-corona' that could be the first stage of biofilm formation was observed. The polymer also activates Caspases on colon cancer cells, making S. vesiculosa EMVs as natural nanocarriers for drug delivery.

NMR, capsule, Biofilm, nanoparticles, SANS, Shewanosamine

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6
2	Ac	174.8	22.0
3	Ac	174.3	21.8
4	Ac	175.3	22.0
	bDSugp	96.0	51.0	51.8	51.1	70.8	15.7


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6
2	Ac	-	1.91
3	Ac	-	1.83
4	Ac	-	2.03
	bDSugp	4.67	3.70	4.07	4.13	3.93	1.08


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6
2	Ac		22.0/1.91
3	Ac		21.8/1.83
4	Ac		22.0/2.03
	bDSugp	96.0/4.67	51.0/3.70	51.8/4.07	51.1/4.13	70.8/3.93	15.7/1.08

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

CC(=O)N[C@@H]1[C@@H](NC(C)=O)C(O)O[C@H](C)[C@@H]1NC(C)=O

287.316 g/mol (C₁₂H₂₁N₃O₅)

Bacterial extracellular membrane vesicles (EMVs) play an active role in many physiological and pathogenic processes. Here, we report the identification and the detailed structural characterization of the capsular polysaccharide from both cells and EMVs from Shewanella vesiculosa by NMR and chemical analysis. The polysaccharide consists of a pentasaccharide repeating unit containing neutral monosaccharides together with amino sugars, of which one has never been isolated from a natural source. The adhesion ability of the polymer both on synthetic surfaces, such as polystyrene nanoparticles and on vesicles with a bilayer mimicking the bacterial membrane in the presence and absence of lipopolysaccharide was investigated. In both cases, a 'CPS-corona' that could be the first stage of biofilm formation was observed. The polymer also activates Caspases on colon cancer cells, making S. vesiculosa EMVs as natural nanocarriers for drug delivery.

NMR, capsule, Biofilm, nanoparticles, SANS, Shewanosamine

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6
3,6,2	Ac	174.3	?
3,6	bDGlcpN	100.6	55.3	73.3	77.4	74.0	60.6
3,4,4,2	Ac	173.9	?
3,4,4,3	Am	166.0	?
3,4,4,4	Ac	175.6	?
3,4,4	aDSugp	95.4	46.7	50.6	49.7	65.0	15.6
3,4,2	Ac	173.9-174.1	?
3,4,3	Ac	173.9-174.1	?
3,4	aDGlcpN3NA	99.1	52.6	52.5	72.6	73.8	174.4
3	aDGlcp	93.9	70.8	72.6	79.1	68.8	66.8
2	Ac	173.4	?
	bLRhap	99.0	68.1	74.3	70.8	72.3	17.0


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6
3,6,2	Ac	-	2.03
3,6	bDGlcpN	4.39	3.75	3.62	3.71	3.44	3.79-3.91
3,4,4,2	Ac	-	1.99
3,4,4,3	Am	?	2.14
3,4,4,4	Ac	-	2.09
3,4,4	aDSugp	5.25	4.16	4.06	4.36	4.27	1.12
3,4,2	Ac	-	1.91-1.97
3,4,3	Ac	-	1.91-1.97
3,4	aDGlcpN3NA	4.97	4.07	4.25	3.95	4.22	-
3	aDGlcp	5.03	3.53	4.00	3.60	4.13	3.77-4.12
2	Ac	-	2.18
	bLRhap	5.08	5.69	3.88	3.52	3.53	1.38


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6
3,6,2	Ac		?/2.03
3,6	bDGlcpN	100.6/4.39	55.3/3.75	73.3/3.62	77.4/3.71	74.0/3.44	60.6/3.79-3.91
3,4,4,2	Ac		?/1.99
3,4,4,3	Am	166.0/?	?/2.14
3,4,4,4	Ac		?/2.09
3,4,4	aDSugp	95.4/5.25	46.7/4.16	50.6/4.06	49.7/4.36	65.0/4.27	15.6/1.12
3,4,2	Ac		?/1.91-1.97
3,4,3	Ac		?/1.91-1.97
3,4	aDGlcpN3NA	99.1/4.97	52.6/4.07	52.5/4.25	72.6/3.95	73.8/4.22	
3	aDGlcp	93.9/5.03	70.8/3.53	72.6/4.00	79.1/3.60	68.8/4.13	66.8/3.77-4.12
2	Ac		?/2.18
	bLRhap	99.0/5.08	68.1/5.69	74.3/3.88	70.8/3.52	72.3/3.53	17.0/1.38

The spectrum also has 7 signals at unknown positions (not plotted).

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

[*]O[C@@H]1[C@@H](CO)O[C@@H](OC[C@H]2O[C@H](O[C@@H]3[C@@H](O)[C@H](C)O[C@H]([*])[C@@H]3OC(C)=O)[C@H](O)[C@@H](O)[C@@H]2O[C@H]2O[C@H](C(=O)O)[C@@H](OC3O[C@H](C)[C@H](NC(C)=O)[C@H](NC(C)=N)[C@H]3NC(C)=O)[C@H](NC(C)=O)[C@H]2NC(C)=O)[C@H](NC(C)=O)[C@H]1O

1080.061 g/mol (C₄₄H₆₉R₂N₇O₂₄, R = next and previous repeats, not counted in mol weight)