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Antje Flieger - One of the best experts on this subject based on the ideXlab platform.
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disulfide loop cleavage of legionella pneumophila plaa boosts lysophospholipase a activity
Scientific Reports, 2017Co-Authors: Christina Lang, Miriam Hiller, Antje FliegerAbstract:L. pneumophila, an important facultative intracellular bacterium, infects the human lung and environmental protozoa. At least fifteen phospholipases A (PLA) are encoded in its genome. Three of which, namely PlaA, PlaC, and PlaD, belong to the GDSL lipase family abundant in bacteria and higher plants. PlaA is a lysophospholipase A (LPLA) that destabilizes the phagosomal membrane in absence of a protective factor. PlaC shows PLA and glycerophospholipid: cholesterol acyltransferase (GCAT) activities which are activated by zinc metalloproteinase ProA via cleavage of a disulphide loop. In this work, we compared GDSL enzyme activities, their secretion, and activation of PlaA. We found that PlaA majorly contributed to LPLA, PlaC to PLA, and both substrate-dependently to GCAT activity. Western blotting revealed that PlaA and PlaC are type II-secreted and both processed by ProA. Interestingly, ProA steeply increased LPLA but diminished GCAT activity of PlaA. Deletion of 20 amino acids within a predicted disulfide loop of PlaA had the same effect. In summary, we propose a model by which ProA processes PlaA via disulfide loop cleavage leading to a steep increase in LPLA activity. Our results help to further characterize the L. pneumophila GDSL hydrolases, particularly PlaA, an enzyme acting in the Legionella-containing phagosome.
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zinc metalloproteinase proa directly activates legionella pneumophila plac glycerophospholipid cholesterol acyltransferase
Journal of Biological Chemistry, 2012Co-Authors: Christina Lang, Elena Rastew, Bjoern Hermes, Enrico Siegbrecht, Robert Ahrends, Sangeeta Banerji, Antje FliegerAbstract:Abstract Enzymes secreted by Legionella pneumophila, such as phospholipases A (PLAs) and glycerophospholipid:cholesterol acyltransferases (GCATs), may target host cell lipids and therefore contribute to the establishment of Legionnaires disease. L. pneumophila possesses three proteins, PlaA, PlaC, and PlaD, belonging to the GDSL family of lipases/acyltransferases. We have shown previously that PlaC is the major GCAT secreted by L. pneumophila and that the zinc metalloproteinase ProA is essential for GCAT activity. Here we characterized the mode of PlaC GCAT activation and determined that ProA directly processes PlaC. We further found that not only cholesterol but also ergosterol present in protozoa was palmitoylated by PlaC. Such ester formations were not induced by either PlaA or PlaD. PlaD was shown here to possess lysophospholipase A activity, and interestingly, all three GDSL enzymes transferred short chain fatty acids to sterols. The three single putative catalytic amino acids (Ser-37, Asp-398, and His-401) proved essential for all PlaC-associated PLA, lysophospholipase A, and GCAT activities. A further four cysteine residues are important for the PLA/GCAT activities as well as their oxidized state, and we therefore conclude that PlaC likely forms at least one disulfide loop. Analysis of cleavage site and loop deletion mutants suggested that for GCAT activation deletion of several amino acids within the loop is necessary rather than cleavage at a single site. Our data therefore suggest a novel enzyme inhibition/activation mechanism where a disulfide loop inhibits PlaC GCAT activity until the protein is exported to the external space where it is ProA-activated.
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characterization of the major secreted zinc metalloprotease dependent glycerophospholipid cholesterol acyltransferase plac of legionella pneumophila
Infection and Immunity, 2005Co-Authors: Sangeeta Banerji, Mayte Bewersdorff, Bjorn Hermes, Nicholas P Cianciotto, Antje FliegerAbstract:Legionella pneumophila, an intracellular pathogen causing a severe pneumonia, possesses distinct lipolytic activities which have not been completely assigned to specific enzymes so far. We cloned and characterized a gene, plaC, encoding a protein with high homology to PlaA, the major secreted lysophospholipase A of L. pneumophila and to other hydrolytic enzymes belonging to the GDSL family. Here we show that L. pneumophila plaC mutants possessed reduced phospholipase A and lysophospholipase A activities and lacked glycerophospholipid:cholesterol acyltransferase activity in their culture supernatants. The mutants' reduced phospholipase A and acyltransferase activities were complemented by reintroduction of an intact copy of plaC. Additionally, plaC conferred increased lysophospholipase A and glycerophospholipid:cholesterol acytransferase activities to recombinant Escherichia coli. Furthermore, PlaC was shown to be another candidate exported by the L. pneumophila type II secretion system and was activated by a factor present in the bacterial culture supernatant dependent on the zinc metalloprotease. Finally, the role of plaC in intracellular infection of Acanthamoeba castellanii and U937 macrophages with L. pneumophila was assessed, and plaC was found to be dispensable. Thus, L. pneumophila possesses another secreted lipolytic enzyme, a protein with acyltransferase, phospholipase A, and lysophospholipase A activities. This enzyme is distinguished from the previously characterized phospholipases A and lysophospholipases A by its capacity not only to cleave fatty acids from lipids but to transfer them to cholesterol. Cholesterol is an important compound of eukaryotic membranes, and an acyltransferase might be a tool for host cell modification to fit the needs of the bacterium.
Junken Aoki - One of the best experts on this subject based on the ideXlab platform.
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phosphatidylserine specific phospholipase a1 ps pla1 expression in colorectal cancer correlates with tumor invasion and hematogenous metastasis
Anticancer Research, 2015Co-Authors: Yuuki Iida, Eiji Sunami, Hiroharu Yamashita, Masaya Hiyoshi, Soichiro Ishihara, Hironori Yamaguchi, Asuka Inoue, Kumiko Makide, Nelson H Tsuno, Junken AokiAbstract:BACKGROUND: The function of phosphatidylserine-specific phospholipase A1 (PS-PLA1), a phospholipase that acts specifically on phosphatidylserine and produces lysophosphatidylserine, a lysophospholipid mediator, has not been fully elucidated. We evaluated the role of PS-PLA1 in oncogenesis and metastasis of colorectal cancer (CRC). MATERIALS AND METHODS: Specimens from 85 patients with CRC were immunostained with a monoclonal antibody against PS-PLA1. The correlation between PS-PLA1 expression and the clinicopathological variables was analyzed. RESULTS: Tumor depth and hematogenous metastasis independently positively correlated with PS-PLA1 expression. High PS-PLA1 expression was associated with shorter disease-free survival, although it was not an independent predictive factor. CONCLUSION: PS-PLA1 expression in CRC is associated with tumor invasion and metastasis.
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mechanisms of lysophosphatidic acid production
Seminars in Cell & Developmental Biology, 2004Co-Authors: Junken AokiAbstract:Lysophosphatidic acid is one of the most attractive phospholipid mediator with multiple biological functions and is implicated in various human diseases. In the past ten years much has been learned about the physiological roles of LPA through series of studies on LPA actions and its receptors. However, the molecular mechanisms of LPA have been poorly understood. LPA is produced in various conditions both in cells and in biological fluids, where multiple synthetic reactions occur. At least two pathways are postulated. In serum and plasma, LPA is mainly converted from lysophospholipids. By contrast, in platelets and some cancer cells, LPA is converted from phosphatidic acid. In each pathway, at least two phospholipase activities are required: phospholipase A1 (PLA1)/PLA2 plus lysophospholipase D (lysoPLD) activities are involved in the first pathway and phospholipase D (PLD) plus PLA1/PLA2 activities are involved in the second pathway. Now multiple phospholipases are identified that account for PLA1, PLA2, PLD, and lysoPLD activities. In the absence of specific inhibitors and genetically modified animals and individuals, the contribution of each phospholipase to LPA production can not be easily determined. However, apparently certain extracellular phospholipases such as secretory PLA2 (sPLA2-IIA), membrane-associated PA-selective PLA1 (mPA-PLA1), lecithin-cholesterol acyltransferase (LCAT), and lysoPLD are involved in LPA production.
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structure and function of phosphatidylserine specific phospholipase a1
Biochimica et Biophysica Acta, 2002Co-Authors: Junken Aoki, Yuki Nagai, Hiroyuki Hosono, Keizo Inoue, Hiroyuki AraiAbstract:Phospholipase A1 (PLA1) is an enzyme that hydrolyzes the sn-1 fatty acids from phospholipids and produces 2-acyl-lysophospholipids. Although PLA1 activities are detected in many tissues and cell lines, a limited number of PLA1s have been purified and cloned so far. These include phosphatidylserine (PS)-specific PLA1 (PS-PLA1) from rat platelets, PLA1 from vespid venom, and phosphatidic acid (PA)-preferential PLA1 (PA-PLA1). Structurally, the former two PLA1s belong to the lipase family, where they form a subfamily among the lipase family. An alignment of the PLA1s with other members of the lipase family revealed two molecular characteristics of PLA1: the presence of extremely short lids and deleted beta9 loops. The two surface loops have been implicated in the ligand recognition in human pancreatic lipase (PL) and guinea pig PL-related protein 2. Under physiological conditions, accessibility of PS-PLA1 to its substrate is limited as it is a secreted enzyme and PS is normally located in the inner leaflet of the lipid bilayer. However, PS-PLA1 efficiently hydrolyzes PS exposed on the surface of cells such as apoptotic cells and activated platelets, and produces 2-acyl-lysophosphatidylserine (lysoPS), which is a lipid mediator for mast cells, T cells and neural cells. Identification of PS-PLA1 reveals the presence of PLA1 subfamily within the lipase family and suggests that PLA1 has a role in the production of lysophospholipid mediators.
Lisong Dong - One of the best experts on this subject based on the ideXlab platform.
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impact of d isomer content on the microstructure and mechanical properties of uniaxially pre stretched poly lactic acid
Polymer, 2020Co-Authors: Yunjing Chen, Ling Zhao, Hongwei Pan, Shiling Jia, Lijing Han, Lisong DongAbstract:Abstract Poly(lactic acid) with 2%, 4%, and 12% d -isomer content were selected to investigate the influence of stereoregularity on mechanical properties of uniaxially pre-stretched PLAs (ps-PLAs). The mobility of molecular chains was activated by pre-stretching because the network structures composed of cohesional entanglements were destroyed, leading to a brittle-ductile transition. The elongation at break increased to 145%, 175% and 190% at pre-stretching ratio of 0.5 for ps-PLA2, ps-PLA4 and ps-PLA12, respectively. The network structures of cohesional entanglements were easier destroyed in PLAs with more d -isomer due to the stereoregularity defects, leading to higher elongation at break. Then the elongation at break decreased with increasing pre-stretching ratio owing to the development of mesophase which might initiate crazing. The more d -isomer the less strain-induced mesophase resulting from the more active chain relaxation, and then the slower decrease of elongation at break. In addition, the modulus and strength of ps-PLAs were all increased with increasing pre-stretching ratio.
Wolfgang Koenig - One of the best experts on this subject based on the ideXlab platform.
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Lipoprotein-Associated Phospholipase A2 Activity Is a Marker of Risk But Not a Useful Target for Treatment in Patients With Stable Coronary Heart Disease.
Journal of the American Heart Association, 2016Co-Authors: Lars Wallentin, Claes Held, Richard Y Davies, Robert A Harrington, Judith S. Hochman, Paul W. Armstrong, Christopher P. Cannon, Christopher B. Granger, Emil Hagström, Wolfgang KoenigAbstract:Background We evaluated lipoprotein‐associated phospholipase A2 (Lp‐PLA2) activity in patients with stable coronary heart disease before and during treatment with darapladib, a selective Lp‐PLA2 inhibitor, in relation to outcomes and the effects of darapladib in the STABILITY trial. Methods and Results Plasma Lp‐PLA2 activity was determined at baseline (n=14 500); at 1 month (n=13 709); serially (n=100) at 3, 6, and 18 months; and at the end of treatment. Adjusted Cox regression models evaluated associations between Lp‐PLA2 activity levels and outcomes. At baseline, the median Lp‐PLA2 level was 172.4 μmol/min per liter (interquartile range 143.1–204.2 μmol/min per liter). Comparing the highest and lowest Lp‐PLA2 quartile groups, the hazard ratios were 1.50 (95% CI 1.23–1.82) for the primary composite end point (cardiovascular death, myocardial infarction, or stroke), 1.95 (95% CI 1.29–2.93) for hospitalization for heart failure, 1.42 (1.07–1.89) for cardiovascular death, and 1.37 (1.03–1.81) for myocardial infarction after adjustment for baseline characteristics, standard laboratory variables, and other prognostic biomarkers. Treatment with darapladib led to a ≈65% persistent reduction in median Lp‐PLA2 activity. There were no associations between on‐treatment Lp‐PLA2 activity or changes of Lp‐PLA2 activity and outcomes, and there were no significant interactions between baseline and on‐treatment Lp‐PLA2 activity or changes in Lp‐PLA2 activity levels and the effects of darapladib on outcomes. Conclusions Although high Lp‐PLA2 activity was associated with increased risk of cardiovascular events, pharmacological lowering of Lp‐PLA2 activity by ≈65% did not significantly reduce cardiovascular events in patients with stable coronary heart disease, regardless of the baseline level or the magnitude of change of Lp‐PLA2 activity. Clinical Trial Registration URL: . Unique identifier: NCT00799903.
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relationship of lipoprotein associated phospholipase a2 mass and activity with incident vascular events among primary prevention patients allocated to placebo or to statin therapy an analysis from the jupiter trial
Clinical Chemistry, 2012Co-Authors: Paul M Ridke, Jea G Macfadye, Robe L Wolfe, Wolfgang KoenigAbstract:BACKGROUND: Although lipoprotein-associated phospholipase A2 (Lp-PLA2) levels are associated with cardiovascular events, Lp-PLA2 is physically linked to LDL cholesterol (LDL-C). Whether measures of Lp-PLA2 mass or activity continue to predict risk after LDL-C reduction by statin therapy is uncertain. METHODS: Lp-PLA2 mass concentration and activity were evaluated at baseline and after treatment in the Justification for the Use of Statins in Prevention: an Intervention Trial Evaluating Rosuvastatin (JUPITER) trial comparing rosuvastatin 20 mg to placebo among 17 802 men and women without cardiovascular disease or diabetes at study entry. The relationships of Lp-PLA2 mass and activity with risk of future vascular events were evaluated in the placebo and rosuvastatin groups. RESULTS: Before randomization, levels of Lp-PLA2 mass and activity correlated moderately with each other and with LDL-C. The magnitude of these correlations increased after statin therapy. Rosuvastatin reduced Lp-PLA2 mass by 33.8%, Lp-PLA2 activity by 33.2%, and LDL-C by 48.7% (all P < 0.0001). Among those study participants allocated to placebo, increasing quartiles of Lp-PLA2 activity ( P trend = 0.04) but not Lp-PLA2 mass ( P trend = 0.92) were associated with incident cardiovascular events after adjustment for LDL-C and conventional risk factors. Comparable analyses conducted among those allocated to rosuvastatin revealed no significant relationship between Lp-PLA2 levels and subsequent vascular events. The ability of rosuvastatin to reduce vascular events was not significantly modified by baseline Lp-PLA2 level. CONCLUSIONS: Among JUPITER trial participants allocated to placebo, levels of Lp-PLA2 activity, but not mass, were associated with cardiovascular risk. However, Lp-PLA2 no longer predicted risk or modified clinical outcomes when participants were treated with rosuvastatin.
Sangeeta Banerji - One of the best experts on this subject based on the ideXlab platform.
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zinc metalloproteinase proa directly activates legionella pneumophila plac glycerophospholipid cholesterol acyltransferase
Journal of Biological Chemistry, 2012Co-Authors: Christina Lang, Elena Rastew, Bjoern Hermes, Enrico Siegbrecht, Robert Ahrends, Sangeeta Banerji, Antje FliegerAbstract:Abstract Enzymes secreted by Legionella pneumophila, such as phospholipases A (PLAs) and glycerophospholipid:cholesterol acyltransferases (GCATs), may target host cell lipids and therefore contribute to the establishment of Legionnaires disease. L. pneumophila possesses three proteins, PlaA, PlaC, and PlaD, belonging to the GDSL family of lipases/acyltransferases. We have shown previously that PlaC is the major GCAT secreted by L. pneumophila and that the zinc metalloproteinase ProA is essential for GCAT activity. Here we characterized the mode of PlaC GCAT activation and determined that ProA directly processes PlaC. We further found that not only cholesterol but also ergosterol present in protozoa was palmitoylated by PlaC. Such ester formations were not induced by either PlaA or PlaD. PlaD was shown here to possess lysophospholipase A activity, and interestingly, all three GDSL enzymes transferred short chain fatty acids to sterols. The three single putative catalytic amino acids (Ser-37, Asp-398, and His-401) proved essential for all PlaC-associated PLA, lysophospholipase A, and GCAT activities. A further four cysteine residues are important for the PLA/GCAT activities as well as their oxidized state, and we therefore conclude that PlaC likely forms at least one disulfide loop. Analysis of cleavage site and loop deletion mutants suggested that for GCAT activation deletion of several amino acids within the loop is necessary rather than cleavage at a single site. Our data therefore suggest a novel enzyme inhibition/activation mechanism where a disulfide loop inhibits PlaC GCAT activity until the protein is exported to the external space where it is ProA-activated.
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characterization of the major secreted zinc metalloprotease dependent glycerophospholipid cholesterol acyltransferase plac of legionella pneumophila
Infection and Immunity, 2005Co-Authors: Sangeeta Banerji, Mayte Bewersdorff, Bjorn Hermes, Nicholas P Cianciotto, Antje FliegerAbstract:Legionella pneumophila, an intracellular pathogen causing a severe pneumonia, possesses distinct lipolytic activities which have not been completely assigned to specific enzymes so far. We cloned and characterized a gene, plaC, encoding a protein with high homology to PlaA, the major secreted lysophospholipase A of L. pneumophila and to other hydrolytic enzymes belonging to the GDSL family. Here we show that L. pneumophila plaC mutants possessed reduced phospholipase A and lysophospholipase A activities and lacked glycerophospholipid:cholesterol acyltransferase activity in their culture supernatants. The mutants' reduced phospholipase A and acyltransferase activities were complemented by reintroduction of an intact copy of plaC. Additionally, plaC conferred increased lysophospholipase A and glycerophospholipid:cholesterol acytransferase activities to recombinant Escherichia coli. Furthermore, PlaC was shown to be another candidate exported by the L. pneumophila type II secretion system and was activated by a factor present in the bacterial culture supernatant dependent on the zinc metalloprotease. Finally, the role of plaC in intracellular infection of Acanthamoeba castellanii and U937 macrophages with L. pneumophila was assessed, and plaC was found to be dispensable. Thus, L. pneumophila possesses another secreted lipolytic enzyme, a protein with acyltransferase, phospholipase A, and lysophospholipase A activities. This enzyme is distinguished from the previously characterized phospholipases A and lysophospholipases A by its capacity not only to cleave fatty acids from lipids but to transfer them to cholesterol. Cholesterol is an important compound of eukaryotic membranes, and an acyltransferase might be a tool for host cell modification to fit the needs of the bacterium.