The structure of a novel neutral lipid A from the lipopolysaccharide of Bradyrhizobium elkanii containing three mannose units in the backbone.
The chemical structure of the lipid A of the lipopolysaccharide (LPS) from Bradyrhizobium elkanii USDA 76 (a member of the group of slow-growing rhizobia) has been established. It differed considerably from lipids A of other Gram-negative bacteria, in that it completely lacks negatively charged groups (phosphate or uronic acid residues); the glucosamine (GlcpN) disaccharide backbone is replaced by one consisting of 2,3-dideoxy-2,3-diamino-D-glucopyranose (GlcpN3N) and it contains two long-chain fatty acids, which is unusual among rhizobia. The GlcpN3N disaccharide was further substituted by three D-mannopyranose (D-Manp) residues, together forming a pentasaccharide. To establish the structural details of this molecule, 1D and 2D NMR spectroscopy, chemical composition analyses and high-resolution mass spectrometry methods (electrospray ionisation Fourier-transform ion cyclotron resonance mass spectrometry (ESI FT-ICR MS) and tandem mass spectrometry (MS/MS)) were applied. By using 1D and 2D NMR spectroscopy experiments, it was confirmed that one D-Manp was linked to C-1 of the reducing GlcpN3N and an alpha-(1-->6)-linked D-Manp disaccharide was located at C-4' of the non-reducing GlcpN3N (alpha-linkage). Fatty acid analysis identified 12:0(3-OH) and 14:0(3-OH), which were amide-linked to GlcpN3N. Other lipid A constituents were long (omega-1)-hydroxylated fatty acids with 26-33 carbon atoms, as well as their oxo forms (28:0(27-oxo) and 30:0(29-oxo)). The 28:0(27-OH) was the most abundant acyl residue. As confirmed by high-resolution mass spectrometry techniques, these long-chain fatty acids created two acyloxyacyl residues with the 3-hydroxy fatty acids. Thus, lipid A from B. elkanii comprised six acyl residues. It was also shown that one of the acyloxyacyl residues could be further acylated by 3-hydroxybutyric acid (linked to the (omega-1)-hydroxy group).
Komaniecka
Iwona
I
Department of Genetics and Microbiology, Maria Curie-Sklodowska University Akademicka 19, 20-033 Lublin, Poland. ikoma@hektor.umcs.lublin.pl
Choma
Adam
A
Lindner
Buko
B
Holst
Otto
O
D020369:Bradyrhizobium / Q000378:metabolism*; D002240:Carbohydrate Sequence; D008050:Lipid A / Q000737:chemistry*; D008070:Lipopolysaccharides / Q000737:chemistry* / Q000302:isolation & purification; D009682:Magnetic Resonance Spectroscopy; D008358:Mannose / Q000737:chemistry*; D008969:Molecular Sequence Data; D021241:Spectrometry, Mass, Electrospray Ionization
D008050:Lipid A; D008070:Lipopolysaccharides; D008358:Mannose