Lipid A structural modifications in extreme conditions and identification of unique modifying enzymes to define the Toll-like receptor 4 structure-activity relationship

Scott, Alison; Oyler, Benjamin L.; Goodlett, David R.; Ernst, Robert K.

doi:10.1016/j.bbalip.2017.01.004

Cited by 51 publications

(54 citation statements)

References 136 publications

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“…Changes in lipid A structure can alter the outer membrane permeability barrier and affect resistance to CAMPs, as well as various antibiotics. Furthermore, since lipid A is the bioactive portion of LPS that is recognized by the innate immune system, remodeling of the lipid A structure greatly influences the host inflammatory response (Needham and Trent, ; Scott et al ., ).…”

Section: Introductionmentioning

confidence: 97%

Novel coordination of lipopolysaccharide modifications in Vibrio cholerae promotes CAMP resistance

Herrera

Henderson

Crofts

et al. 2017

Molecular Microbiology

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Summary In the environment and during infection, the human intestinal pathogen Vibrio cholerae must overcome noxious compounds that damage the bacterial outer membrane. The El Tor and classical biotypes of O1 V. cholerae show striking differences in their resistance to membrane disrupting cationic antimicrobial peptides (CAMPs), such as polymyxins. The classical biotype is susceptible to CAMPs, but current pandemic El Tor biotype isolates gain CAMP resistance by altering the net charge of their cell surface through glycine modification of lipid A. Here we report a second lipid A modification mechanism that only functions in the V. cholerae El Tor biotype. We identify a functional EptA ortholog responsible for the transfer of the amino-residue phosphoethanolamine (pEtN) to the lipid A of V. cholerae El Tor that is not functional in the classical biotype. We previously reported that mildly acidic growth conditions (pH 5.8) downregulate expression of genes encoding the glycine modification machinery. In this report, growth at pH 5.8 increases expression of eptA with concomitant pEtN modification suggesting coordinated regulation of these LPS modification systems. Similarly, efficient pEtN lipid A substitution is seen in the absence of lipid A glycinylation. We further demonstrate EptA orthologs from non-cholerae Vibrio species are functional.

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Section: Introductionmentioning

confidence: 97%

Novel coordination of lipopolysaccharide modifications in Vibrio cholerae promotes CAMP resistance

Herrera

Henderson

Crofts

et al. 2017

Molecular Microbiology

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“…Endotoxin is the canonical ligand for Toll-like receptor 4 (TLR4) (3,4) and endotoxin-like lipids are absent from the eukaryotic lipid repertoire. Individual bacterial lipid A structures are unique (5)(6)(7), although modifiable during growth (8,9) to accommodate the diverse conditions encountered (10) during transmission from the environment to a warm-blooded host. Unlike the highly stimulatory endotoxin derived from enteric bacteria (10,11), lipid A derived from Francisella species does not stimulate the canonical endotoxin pattern recognition receptor TLR4 (12) and is an important virulence factor.…”

mentioning

confidence: 99%

“…Individual bacterial lipid A structures are unique (5)(6)(7), although modifiable during growth (8,9) to accommodate the diverse conditions encountered (10) during transmission from the environment to a warm-blooded host. Unlike the highly stimulatory endotoxin derived from enteric bacteria (10,11), lipid A derived from Francisella species does not stimulate the canonical endotoxin pattern recognition receptor TLR4 (12) and is an important virulence factor. Mapping of the protein-based virulence determinant, Burkholderia mallei GroEL was recently demonstrated (13) using mass spectrometry imaging (MSI), highlighting the growing application of MSI to problems in microbial pathogenesis.…”

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confidence: 99%

Host-based lipid inflammation drives pathogenesis in Francisella infection

Scott

Post

Lerner

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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Mass spectrometry imaging (MSI) was used to elucidate host lipids involved in the inflammatory signaling pathway generated at the host-pathogen interface during a septic bacterial infection. Using as a model organism, a bacterial lipid virulence factor (endotoxin) was imaged and identified along with host phospholipids involved in the splenic response in murine tissues. Here, we demonstrate detection and distribution of endotoxin in a lethal murine infection model, in addition to determining the temporally and spatially resolved innate lipid inflammatory response in both 2D and 3D renderings using MSI. Further, we show that the cyclooxygenase-2-dependent lipid inflammatory pathway is responsible for lethality in infection due to overproduction of proinflammatory effectors including prostaglandin E2. The results of this study emphasize that spatial determination of the host lipid components of the immune response is crucial to identifying novel strategies to effectively address highly pathogenic and lethal infections stemming from bacterial, fungal, and viral origins.

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“…However, due to differences in abundance or structure of LPS modifying enzymes, the resultant products from these syntheses can vary greatly, both in structure and function, even within one species [11]. Many non-stoichiometric substitutions of phosphate groups, sugars, amino acids, amines, and other R-groups are also observed in LPS extracts, making accurate structural analyses challenging.…”

Section: Introductionmentioning

confidence: 99%

Top Down Tandem Mass Spectrometric Analysis of a Chemically Modified Rough-Type Lipopolysaccharide Vaccine Candidate

Oyler

Khan

Smith

et al. 2018

J. Am. Soc. Mass Spectrom.

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Recent advances in lipopolysaccharide (LPS) biology have led to its use in drug discovery pipelines, including vaccine and vaccine adjuvant discovery. Desirable characteristics for LPS vaccine candidates include both the ability to produce a specific antibody titer in patients and a minimal host inflammatory response directed by the innate immune system. However, in-depth chemical characterization of most LPS extracts has not been performed; hence, biological activities of these extracts are unpredictable. Additionally, the most widely adopted workflow for LPS structure elucidation includes nonspecific chemical decomposition steps before analyses, making structures inferred and not necessarily biologically relevant. In this work, several different mass spectrometry workflows that have not been previously explored were employed to show proof-of-principle for top down LPS primary structure elucidation, specifically for a rough-type mutant (J5) E. coli-derived LPS component of a vaccine candidate. First, ion mobility filtered precursor ions were subjected to collision induced dissociation (CID) to define differences in native J5 LPS v. chemically detoxified J5 LPS (dLPS). Next, ultra-high mass resolving power, accurate mass spectrometry was employed for unequivocal precursor and product ion empirical formulae generation. Finally, MS analyses in an ion trap instrument showed that previous knowledge about dissociation of LPS components can be used to reconstruct and sequence LPS in a top down fashion. A structural rationale is also explained for differential inflammatory dose-response curves, in vitro, when HEK-Blue hTLR4 cells were administered increasing concentrations of native J5 LPS v. dLPS, which will be useful in future drug discovery efforts. Graphical Abstract ᅟ.

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Lipid A structural modifications in extreme conditions and identification of unique modifying enzymes to define the Toll-like receptor 4 structure-activity relationship

Cited by 51 publications

References 136 publications

Novel coordination of lipopolysaccharide modifications in Vibrio cholerae promotes CAMP resistance

Novel coordination of lipopolysaccharide modifications in Vibrio cholerae promotes CAMP resistance

Host-based lipid inflammation drives pathogenesis in Francisella infection

Top Down Tandem Mass Spectrometric Analysis of a Chemically Modified Rough-Type Lipopolysaccharide Vaccine Candidate

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