Browsing by Subject "Lipid analysis"
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Item Helicobacter pylori versus the Host: Remodeling of the Bacterial Outer Membrane Is Required for Survival in the Gastric Mucosa(Public Library of Science, 2011-12-22) Cullen, Thomas W.; Giles, David K.; Wolf, Lindsey N.; Ecobichon, Chantal; Boneca, Ivo G.; Trent, M. StephenModification of bacterial surface structures, such as the lipid A portion of lipopolysaccharide (LPS), is used by many pathogenic bacteria to help evade the host innate immune response. Helicobacter pylori, a gram-negative bacterium capable of chronic colonization of the human stomach, modifies its lipid A by removal of phosphate groups from the 1- and 4′-positions of the lipid A backbone. In this study, we identify the enzyme responsible for dephosphorylation of the lipid A 4′-phosphate group in H. pylori, Jhp1487 (LpxF). To ascertain the role these modifications play in the pathogenesis of H. pylori, we created mutants in lpxE (1-phosphatase), lpxF (4′-phosphatase) and a double lpxE/F mutant. Analysis of lipid A isolated from lpxE and lpxF mutants revealed lipid A species with a 1 or 4′-phosphate group, respectively while the double lpxE/F mutant revealed a bis-phosphorylated lipid A. Mutants lacking lpxE, lpxF, or lpxE/F show a 16, 360 and 1020 fold increase in sensitivity to the cationic antimicrobial peptide polymyxin B, respectively. Moreover, a similar loss of resistance is seen against a variety of CAMPs found in the human body including LL37, β-defensin 2, and P-113. Using a fluorescent derivative of polymyxin we demonstrate that, unlike wild type bacteria, polymyxin readily associates with the lpxE/F mutant. Presumably, the increase in the negative charge of H. pylori LPS allows for binding of the peptide to the bacterial surface. Interestingly, the action of LpxE and LpxF was shown to decrease recognition of Helicobacter LPS by the innate immune receptor, Toll-like Receptor 4. Furthermore, lpxE/F mutants were unable to colonize the gastric mucosa of C57BL/6J and C57BL/6J tlr4 -/- mice when compared to wild type H. pylori. Our results demonstrate that dephosphorylation of the lipid A domain of H. pylori LPS by LpxE and LpxF is key to its ability to colonize a mammalian host.Item Polymer applications for improved biofuel production from algae(2011-12) Jones, Jessica Naomi; Poenie, Martin F.; Brand, Jerry; Brodbelt, Jennifer; Georgiou, George; Roy, Krishnendu; Seibert, FrankBiofuel is a renewable and sustainable energy source with near-neutral carbon footprint. Algae are an ideal feedstock for biofuel production because they reproduce quickly and have high oil. Algae can be cultivated in non-arable land, and would not impact the food supply. Unfortunately, processing algae into biofuel is more expensive than land crops due to the large volumes of dilute algal suspension that must be harvested and concentrated. In order to improve algae-based biofuel economics, resins were developed that reduce costs associated with water pumping and transport. Hydrophobic resins were developed for binding oil out of an algal suspension so that the residual biomass could be recovered without solvent contamination. Binding behavior displayed lipid species specificity, and binding capacity was improved by ethanol treatment of the biomass. Algae was harvested by binding to anion exchange resin and directly converted into biodiesel. One-step, room temperature in situ transesterification of algae yielded nearly as much biodiesel as two-step, heated transesterification of dried biomass. Elution with transesterification reagent also regenerated the resin for subsequent algal binding. Functionalized resins were developed with high algal binding capacity at neutral pH. Binding was easily reversed, as treatment with buffer with pH higher than pKa of the resin functional group removed the algae and regenerated the resin for subsequent use. The resin bound 10% of its weight in algae and released it as a 100-fold concentrated suspension. The polymers developed can be scaled up for commercial processes and reduce algal harvesting and concentration costs.