Total Synthesis of the Proposed Structure of Rhodobacter sphaeroides Lipid A Resulting in the Synthesis of New Potent Lipopolysaccharide Antagonists

Christ, William J.; McGuinness, Pamela; Asano, Osamu; Wei, Yuan; Mullarkey, Maureen; Perez, Michel; Hawkins, Lynn D.; Blythe, Todd A.; Dubuc, Gloria R.; Robidoux, Andrea L.

doi:10.1021/ja00087a075

Cited by 86 publications

(56 citation statements)

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“…Chemical syntheses of pure lipid A molecular species (the endotoxin component of LPS) and of various lipid A analogs have been reported (16,(38)(39)(40)(41). The most relevant of these compounds, such as E. coli lipid A 38, are not easy to prepare and are not readily available to the scientific community.…”

Section: Discussionmentioning

confidence: 99%

Kdo2-Lipid A of Escherichia coli, a defined endotoxin that activates macrophages via TLR-4

Raetz

Garrett

Reynolds

et al. 2006

Journal of Lipid Research

209

248

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The LIPID MAPS Consortium (www.lipidmaps. org) is developing comprehensive procedures for identifying all lipids of the macrophage, following activation by endotoxin. The goal is to quantify temporal and spatial changes in lipids that occur with cellular metabolism and to develop bioinformatic approaches that establish dynamic lipid networks. To achieve these aims, an endotoxin of the highest possible analytical specification is crucial. We now report a large-scale preparation of 3-deoxy-D-manno-octulosonic acid (Kdo) 2 -Lipid A, a nearly homogeneous Re lipopolysaccharide (LPS) sub-structure with endotoxin activity equal to LPS. Kdo 2 -Lipid A was extracted from 2 kg cell paste of a heptose-deficient Escherichia coli mutant. It was purified by chromatography on silica, DEAE-cellulose, and C18 reverse-phase resin. Structure and purity were evaluated by electrospray ionization/mass spectrometry, liquid chromatography/mass spectrometry and 1 H-NMR. Its bioactivity was compared with LPS in RAW 264.7 cells and bone marrow macrophages from wild-type and toll-like receptor 4 (TLR-4)-deficient mice. Cytokine and eicosanoid production, in conjunction with gene expression profiling, were employed as readouts. Kdo 2 -Lipid A is comparable to LPS by these criteria. Its activity is reduced by . The LIPID MAPS consortium is developing quantitative methods for evaluating the composition, biosynthesis, and function of all macrophage lipids (1). These amphipathic substances not only are structural components of biological membranes but also play important roles in the pathophysiology of inflammation, atherosclerosis, and growth control. Additional lipid functions should emerge from the comprehensive analysis of macrophage lipids. Electrospray ionization/mass spectrometry (ESI/MS) (2, 3), coupled with prefractionation methods like reversephase liquid chromatography (LC), is being applied systematically to set the stage for the seamless integration of lipid metabolism into the broader fields of genomics, proteomics, and systems biology. To facilitate this endeavor, LIPID MAPS has introduced a new comprehensive classification system for biological lipids, amenable to computer-based data processing and substructure comparison (4). The eight LIPID MAPS categories are 1) fatty acyls, 2) glycerolipids, 3) glycerophospholipids, 4) sphingolipids, 5) sterol lipids, 6) prenol lipids, 7) saccharolipids,

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Section: Discussionmentioning

confidence: 99%

Kdo2-Lipid A of Escherichia coli, a defined endotoxin that activates macrophages via TLR-4

Raetz

Garrett

Reynolds

et al. 2006

Journal of Lipid Research

209

248

View full text Add to dashboard Cite

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“…Currently, such antagonists are chemically synthesized (30,57) or are isolated from bacteria like R. sphaeroides and Rhodobacter capsulatus (5,28,29). Enzymatic and/or in vivo synthesis might increase efficiency, yield, and accessibility.…”

Section: Discussionmentioning

confidence: 99%

“…However, the lipid A molecules from some other organisms, for instance R. sphaeroides, are potent antagonists of that response in mouse and human cells (28 -30). One major difference in the lipid A of R. sphaeroides, as compared with E. coli, is that the 3 and 3Ј O-linked fatty acids are hydroxydecanoates rather than hydroxymyristates (18,30). Thus, subtle features of the structure of lipid A, especially the nature of the fatty acids at the ester-linked positions, appear to be important in triggering the activation of macrophages.…”

Section: Lipopolysaccharide (Lps)mentioning

confidence: 99%

Shortened Hydroxyacyl Chains on Lipid A of Escherichia coli Cells Expressing a Foreign UDP-N-Acetylglucosamine O-Acyltransferase

Odegaard¹,

Kaltashov²,

Cotter³

et al. 1997

Journal of Biological Chemistry

106

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The first reaction of lipid A biosynthesis in Gramnegative bacteria is catalyzed by UDP-N-acetylglucosamine (UDP-GlcNAc) O-acyltransferase, the product of the lpxA gene. The reaction involves the transfer of an acyl chain from hydroxyacyl-acyl carrier protein (ACP) to the glucosamine 3-OH position of UDP-GlcNAc. The lipid A isolated from Escherichia coli contains (R)-3-hydroxymyristate at the 3 and 3 positions. Accordingly, LpxA of E. coli is highly selective for (R)-3-hydroxymyristoyl-ACP over ACP thioesters of longer or shorter acyl chains. We now demonstrate that the lpxA gene from Neisseria meningitidis encodes a similar acyltransferase that selectively utilizes 3-hydroxylauroyl-ACP. Strains of E. coli harboring the temperature-sensitive lpxA2 mutation make very little lipid A and lose viability rapidly at 42°C. We have created an E. coli strain in which the chromosomal lpxA2 mutation is complemented by the N. meningitidis lpxA gene introduced on a plasmid. This strain, RO138/pTO6, grows similarly to wild type cells at 42°C and produces wild type levels of lipid A. However, the lipid A isolated from RO138/pTO6 contains mostly hydroxylaurate and hydroxydecanoate in the 3 and 3 positions. The strain RO138/pTO6 is more susceptible than wild type to certain antibiotics at 42°C. This is the first report of an E. coli strain growing with shortened hydroxyacyl chains on its lipid A. The lpxA gene product appears to be a critical determinant of the length of the ester-linked hydroxyacyl chains found on lipid A in living cells.

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“…11,12 The most widely studied analog is a synthetic analog based on the lipid A of Rhodobacter sphaeroides or R. capsulatus, two species having very similar lipid A structures. [13][14][15][16] Although the lipid As of R. sphaeroides/R. capsulatus and E. coli have the same bis-1,4′-phosphorylated glucosamine disaccharide backbone their fatty acyl patterns differ considerably.…”

Section: Introductionmentioning

confidence: 99%

Agonistic and antagonistic properties of a Rhizobium sin-1 lipid A modified by an ether-linked lipid

2007

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LPS from Rhizobium sin-1 (R. sin-1) can antagonize the production of tumor necrosis factor alpha (TNF-α) by E. coli LPS in human monocytic cells. Therefore these compounds provide interesting leads for the development of therapeutics for the prevention or treatment of septic shock. Detailed structure activity relationship studies have, however, been hampered by the propensity of these compounds to undergo β-elimination to give biological inactive enone derivatives. To address this problem, we have chemically synthesized in a convergent manner a R. sin-1 lipid A derivative in which the β-hydroxy ester at C-3 of the proximal sugar unit has been replaced by an ether linked moiety. As expected, this derivative exhibited a much-improved chemical stability. Furthermore, its ability to antagonize TNF-α production induced by enteric LPS was only slightly smaller than that of the parent ester modified derivative demonstrating that the ether-linked lipids affect biological activities only marginally. Furthermore, it has been shown for the first time that R. sin-1 LPS and the ether modified lipid A are also able to antagonize the production of the cytokine interferoninducible protein 10, which arises from the TRIF-dependent pathway. The latter pathway was somewhat more potently inhibited than the MyD88-dependent pathway. Furthermore, it was observed that the natural LPS possesses much greater activity than the synthetic and isolated lipid As, which indicates that di-KDO moiety is important for optimal biological activity. It has also been found that isolated R. sin-1 LPS and lipid A agonize a mouse macrophage cell line to induce the production of TNF-α and interferon beta in a Toll-like receptor 4-dependent manner demonstrating species specific properties.

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Total Synthesis of the Proposed Structure of Rhodobacter sphaeroides Lipid A Resulting in the Synthesis of New Potent Lipopolysaccharide Antagonists

Cited by 86 publications

References 0 publications

Kdo2-Lipid A of Escherichia coli, a defined endotoxin that activates macrophages via TLR-4

Kdo2-Lipid A of Escherichia coli, a defined endotoxin that activates macrophages via TLR-4

Shortened Hydroxyacyl Chains on Lipid A of Escherichia coli Cells Expressing a Foreign UDP-N-Acetylglucosamine O-Acyltransferase

Agonistic and antagonistic properties of a Rhizobium sin-1 lipid A modified by an ether-linked lipid

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