Studies of lipid A fractions from the lipopolysaccharides of <i>Pseudomonas aeruginosa</i> and <i>Pseudomonas alcaligenes</i>

Drewry, David T.; Lomax, James A.; Gray, G. W.; Wilkinson, Stephen G.

doi:10.1042/bj1330563

Cited by 35 publications

(24 citation statements)

References 32 publications

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“…The purified LF (20 mg) was treated with alkaline phosphatase by the method of Gray et al [22]. After alkaline phosphatase treatment, the remnant of LF was tested by TLC using Dittmer staining and TLC-Ctxb overlay.…”

Section: Structural Analysis Of Lfmentioning

confidence: 99%

“…The molecular weight of the purified LF was determined by MALDI-TOF mass spectrometry. Phosphate was released from the purified LF by alkaline phosphatase treatment as described in the method of Gray et al [22]. After chloroform partitioning, the phosphate content of the aqueous layer was quantitated by the method of Touster et al [23].…”

Section: Structural Determination Of Lfmentioning

confidence: 99%

“…The purified LF (20 mg) was treated with alkaline phosphatase (from E. coli, Sigma, St. Louis, MO, USA) using the method of Gray et al [22]. Briefly, 1 ml of alkaline phosphatase solution containing 50 units of the enzyme in 0.05M (NH 4 ) 2 CO 3 , pH 9.6, was incubated with the purified LF (20 mg), covered with 0.1 ml of toluene to retard bacterial growth, and incubated for 48 h at 37°.…”

Section: Alkaline Phosphatase Treatmentmentioning

confidence: 99%

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Chemical validation of molecular mimicry: interaction of cholera toxin with Campylobacter lipooligosaccharides

et al. 2007

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It is generally believed that molecular mimicry between bacterial lipooligosaccharide (LOS) and nerve glycolipids may play an important pathogenic role in immune-mediated peripheral neuropathy. One of the putative infectious agents is Campylobacter jejuni (C. jejuni). To elucidate the structural basis for the molecular mimicry, we investigated the structure of the lipooligosaccharide (LOS) fraction of C. jejuni, strain HS19, and found that it includes at least two components, characterized as fast-and slow-moving bands (LF and LS) by thin-layer chromatography as revealed by cholera toxin B subunit (Ctxb) overlay. Structural analysis of the oligosaccharide portion of LS established that it had the following structure: Gal-GalNAc-(NeuAc)Gal-Hep-(Glc;PO 3 H)Hep-Kdo. The GM1-like epitope was validated by a terminal tetrasaccharide unit within this structure. On the other hand, analysis of LF revealed an entirely different structure: 1, 4′-bisphosphoryl glucosamine disaccharide N, N'-acylated by 3-(2-hydroxytetracosanoyloxy)octadecanoic acid at 2-and 2′-positions, which is consistent with that of lipid A. No GM1-like epitope was observed in LF. Both LS and LF interacted with Ctxb as demonstrated by TLC-overlay and sucrose density gradient centrifugation. Surprisingly, LF does not have the basic GM1 structure for interacting with Ctxb. Instead, the affinity of LF to Ctxb required that one or both of the phosphate groups be present in the glucosamine disaccharide residue because after alkaline phosphatase treatment the dephosphorylated LF was unable to bind to Ctxb. We conclude that LS is likely the component contributing to GM1-mimicry in autoimmune peripheral neuropathy and that the role of LF is not clear but may be associated with the initial activation of autoreactive T cells.

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Section: Structural Analysis Of Lfmentioning

confidence: 99%

Section: Structural Determination Of Lfmentioning

confidence: 99%

Section: Alkaline Phosphatase Treatmentmentioning

confidence: 99%

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Chemical validation of molecular mimicry: interaction of cholera toxin with Campylobacter lipooligosaccharides

et al. 2007

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“…aeruginosa strains contain both ester-and amidebonded fatty acids. 3-Hydroxydodecanoic acid (3-OH C12:0) is found amide linked to C-2 of glucosamine residues in the lipid A component of LPS (9,10). The lipid A also contains ester-bound 3-hydroxydecanoate (3-OH C10:0), dodecanoate (C12:0), 2- (C18i1) acids (14,32,33).…”

mentioning

confidence: 99%

Effect of growth temperature on the lipids, outer membrane proteins, and lipopolysaccharides of Pseudomonas aeruginosa PAO

Kropinski¹,

Lewis²,

Berry³

1987

J Bacteriol

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Growth of Pseudomonas aeruginosa PAO1 at 15 to 45°C in tryptic soy broth resulted in changes in the lipids, lipopolysaccharides (LPSs), and outer membrane proteins of the cells. Cells grown at 15TC contained, relative to those cultivated at 45°C, increased levels of the phospholipid fatty acids hexadecenoate and octadecenoate and reduced levels of the corresponding saturated fatty acids. Furthermore, the lipid A fatty acids also showed thermoadaptation with decreases in dodecanoic and hexadecanoic acids and increases in the level of 3-hydroxydecanoate and 2-hydroxdodecanoate as the growth temperature decreased. In addition, LPS extracted from cells cultivated at the lower temperatures contained a higher content of long-chain S-form molecules than that isolated from cells grown at higher temperatures. On the other hand, the percentage of LPS cores substituted with side-chain material decreased from 37.6 mol% at 45°C to 19.3 mol% at 15°C. The outer membrane protein profiles indicated that at low growth temperatures there was an increase in a polypeptide with an apparent molecular weight of 43,000 and decreases in the content of 21,000 (protein Hl)-and 27,500-molecular-weight proteins.Since Pseudomonas aeruginosa is ubiquitous in nature and is a demonstrated pathogen for plants and insects as well as mamnmals we are interested in the changes that temperature may have on the cellular chemistry, particularly as it affects membrane lipids, lipopolysaccharides (LPSs), and outer membrane proteins. P. aeruginosa strains contain both ester-and amidebonded fatty acids. 3-Hydroxydodecanoic acid (3-OH C12:0) is found amide linked to C-2 of glucosamine residues in the lipid A component of LPS (9, 10). The lipid A also contains ester-bound 3-hydroxydecanoate (3-OH C10:0), dodecanoate (C12:0), 2-hydroxydodecanoate (2-OH C12:0), and a small amount of hexadecanoate (C16:0 141]). The chloroformmethanol-extractable cellular lipids (neutral and phospholipids) contain predominantly hexadecanoic (C16:0), hexadecenoic (C16:1), octadecanoic (C18:0), and octadecenoic (C18i1) acids (14,32,33). In addition, variable amounts of other fatty acids have been observed, including 17-and 19-cyclopropane acids and tetradecanoate (14, 32, 33). While the effects of temperature on the fatty acid profiles of a number of microorganisms, including psychrophilic Pseudomonas species, have been studied in detail (3,4,12,13, 20,21,26,29,31,35), only one study has been done on thermal adaptation of P. aeruginosa. The experiments of Gill and Suisted (13) with P. aeruginosa grown in a chemostat under conditions of nitrogen limitation showed that the extractable lipids of these cells possessed an increased content of unsaturated fatty acids when cultivated at suboptimal temperatures.Studies on the effect of growth temperature on LPS chemistry have been largely restricted to members of the family Enterobacteriaceae. Modifications to the composition of the lipid A component of LPS isolated from Salmonella species (43) We have demonstrated, on the basis of incr...

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“…The chemical structures of lipid A from various bacteria such as Escherichia coli, Pseudomonas aeruginosa, Proteus mirabilis, Salmonella minnesota, Salmonella typhimurium, Shigella sonnei, Yersinia pestis, Er-winia carotovora, and Haemophilus ducreyi are remarkably identical in that they have a backbone of 3-1',6-glucosamine (GlcNAc) (19,32). Heterogeneity of lipid A arises because of variation in the fatty acid composition, which appears to vary depending on the bacterial strain and growth conditions (6,11,21,25,39,40,41,49,52). The composition of the core oligosaccharide is also variable; this oligosaccharide is generally composed of 2-keto-3-deoxyoctonic acid, heptose, glucose, and galactose.…”

mentioning

confidence: 99%

Phosphorylation of lipopolysaccharides in the Antarctic psychrotroph Pseudomonas syringae: a possible role in temperature adaptation

1994

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Phosphorylation of lipopolysaccharide (LPS) from a psychrotrophic bacterium, Pseudomonas syringae, from Antarctica was studied by using sucrose gradient-separated membrane fractions. The bacterium was found to possess an LPS kinase which could phosphorylate more LPS postsynthetically at higher temperatures. The phosphorylation was low at a lower temperature and was also found to occur in vivo. After phosphorylation of LPS in vitro, it was found that the major part of the radioactivity (>85%) was associated with the core oligosaccharide region of the LPS. The phosphate groups of this region are probably involved in the binding of metal ions, which could be removed by EDTA. The cells grown at the lower temperature probably contained fewer divalent cations because of the smaller amount of phosphate and thereby were more sensitive to EDTA. The cells were also more sensitive to cationic antibiotics at the lower temperature. A possible role of this differential phosphorylation of LPS in modulating the function of the outer membrane as a permeability barrier in the psychrotroph is discussed.The outermost layer of the outer membrane bilayer of gram-negative bacteria is composed of lipopolysaccharides (LPS) which contain a hydrophobic moiety, lipid A, and a polysaccharide moiety consisting of the so-called core oligosaccharide and 0 polysaccharide. The chemical structures of lipid A from various bacteria such as Escherichia coli, Pseudomonas aeruginosa, Proteus mirabilis, Salmonella minnesota, Salmonella typhimurium, Shigella sonnei, Yersinia pestis, Er-winia carotovora, and Haemophilus ducreyi are remarkably identical in that they have a backbone of 3-1',6-glucosamine (GlcNAc) (19,32). Heterogeneity of lipid A arises because of variation in the fatty acid composition, which appears to vary depending on the bacterial strain and growth conditions (6,11,21,25,39,40,41,49,52). The composition of the core oligosaccharide is also variable; this oligosaccharide is generally composed of 2-keto-3-deoxyoctonic acid, heptose, glucose, and galactose. The structure of the 0 polysaccharide is most variable, and the 0 polysaccharide may contain various unusual sugars (32,38). The other striking feature of LPS is the presence of a high level of phosphorus (2 to 5.6%), which is attached either to the core sugars (such as 2-keto-3-deoxyoctonic acid and heptose) or to GlcNAc of lipid A as ethanolamine phosphate, ethanolamine PPi, PP1, or simply phosphate (13,25,38,51).Although much information about LPS composition and structure has accumulated during the last 25 years, very little is known about the physiological function of LPS. LPS have been studied as toxins and virulence factors of gram-negative bacteria that cause pathogenesis in animals and plants (19,32). However, the function of LPS with respect to the physiology of the bacteria is poorly understood. In bacteria, LPS has been implicated in providing a kind of permeation barrier for hydrophobic compounds to move across the membrane. The evidence for this has come from the study of...

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Studies of lipid A fractions from the lipopolysaccharides of Pseudomonas aeruginosa and Pseudomonas alcaligenes

Cited by 35 publications

References 32 publications

Chemical validation of molecular mimicry: interaction of cholera toxin with Campylobacter lipooligosaccharides

Chemical validation of molecular mimicry: interaction of cholera toxin with Campylobacter lipooligosaccharides

Effect of growth temperature on the lipids, outer membrane proteins, and lipopolysaccharides of Pseudomonas aeruginosa PAO

Phosphorylation of lipopolysaccharides in the Antarctic psychrotroph Pseudomonas syringae: a possible role in temperature adaptation

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