Exopolysaccharides contribute significantly to attachment and biofilm formation in the opportunisitc pathogen Pseudomonas aeruginosa. The Psl polysaccharide, which is synthesized by the polysaccharide synthesis locus (psl), is required for biofilm formation in non-mucoid strains that do not rely on alginate as the principal biofilm polysaccharide. In-frame deletion and complementation studies of individual psl genes revealed that 11 psl genes, pslACDEFGHIJKL, are required for Psl production and surface attachment. We also present the first structural analysis of the psl-dependent polysaccharide, which consists of a repeating pentasaccharide containing d-mannose, d-glucose and l-rhamnose: [See text]. In addition, we identified the sugar nucleotide precursors involved in Psl generation and demonstrated the requirement for GDP-d-mannose, UDP-d-glucose and dTDP-l-rhamnose in Psl production and surface attachment. Finally, genetic analyses revealed that wbpW restored Psl production in a pslB mutant and pslB promoted A-band LPS synthesis in a wbpW mutant, indicating functional redundancy and overlapping roles for these two enzymes. The structural and genetic data presented here provide a basis for further investigation of the Psl proteins and potential roles for Psl in the biology and pathogenesis of P. aeruginosa

biosynthesis, gene, Pseudomonas aeruginosa, exopolysaccharides, Enzymes

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6
3,3,3,2	aDManp	101.5	71.3	71.5	67.8	73.5	61.5
3,3,3	bDManp	97.8	74.4	82.4	66.8	77.7	61.5
3,3	bDManp	102.4	68.9	80.3	66.3	77.4	61.5
3	aLRhap	102.1	71.5	80.6	72.3	70.0	17.7
	bDGlcp	101.6	74.9	83.8	69.4	77.2	62.1


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6
3,3,3,2	aDManp	5.38	4.06	3.97	3.70	4.20	3.79-3.89
3,3,3	bDManp	4.87	4.39	4.00	3.75	3.44	3.79-3.89
3,3	bDManp	4.88	4.31	3.90	3.66	3.40	3.79-3.89
3	aLRhap	5.14	4.30	3.94	3.59	4.08	1.27
	bDGlcp	4.65	3.47	3.63	3.48	3.48	3.73-3.93


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6
3,3,3,2	aDManp	101.5/5.38	71.3/4.06	71.5/3.97	67.8/3.70	73.5/4.20	61.5/3.79-3.89
3,3,3	bDManp	97.8/4.87	74.4/4.39	82.4/4.00	66.8/3.75	77.7/3.44	61.5/3.79-3.89
3,3	bDManp	102.4/4.88	68.9/4.31	80.3/3.90	66.3/3.66	77.4/3.40	61.5/3.79-3.89
3	aLRhap	102.1/5.14	71.5/4.30	80.6/3.94	72.3/3.59	70.0/4.08	17.7/1.27
	bDGlcp	101.6/4.65	74.9/3.47	83.8/3.63	69.4/3.48	77.2/3.48	62.1/3.73-3.93

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

794.706 g/mol (C₃₀H₅₀R₂O₂₄, R = next and previous repeats, not counted in mol weight)