Toxicity, binding and internalization of the pea-A1b entomotoxin in Sf9 cells

1-ACL (articles avec comité de lecture)PA1b (Pea Albumin 1b) is a peptide toxin lethal for certain insects. This paper shows that the cultured insect cells Sf9 are sensitive to the toxin and display a high-affinity binding site for PA1b. Mammalian cells are not sensitive and no binding activity was detected. Signs of apoptosis of the Sf9 cells were observed in response to the toxin. The use of this cellular model also demonstrated that PA1b was internalized in the cells, via the binding site, raising the new question of the role of this toxin within the cell, and of the mechanisms leading to cell death

A folded and functional synthetic PA1b, an interlocked entomotoxic miniprotein

1-ACL (articles avec comité de lecture)PA1b (Pea Albumin 1, subunit b) is a hydrophobic, 37-amino acid niiniprotein isolated from pea seeds (Pivum sativum), crosslinked by three interlocked disulfide bridges, signature of the ICK (inhibitory cystine-knot) family. It acts as an entomotoxic factor against major insect pests in stored crops and vegetables, making it a promising bioinsecticide. Here we report an efficient and simple protocol for the production of large quantities of highly pure, biologically active synthetic PA1b. The features of PA1b oxidative refolding revealed the off-pathway products and competitive aggregation processes. The efficiency of the oxidative folding can be significantly improved by using hydrophobic alcoholic cosolvents and decreasing the temperature. The homogeneity of the synthetic oxidized PA1b was established by reversed-phase HPLC. The correct pairing of the three disulfide bridges, as well as the three-dimensional structure of synthetic PA1b was assessed by NMR. Synthetic PA1b binds to rnicrosomal proteins from Sitophilus oryzae with a Kd of 8 nM, a figure quite similar to that determined for PA1b extracted from its natural source. Moreover, the synthetic miniprotein was as potent as the extracted one towards the sensitive strains of weevils. Our findings will open the way to the production of PA1b analogues by chemical means to an in-depth understanding of the PA1b mechanism of action

Biological activity and binding site characteristics of the PA1b entomotoxin on insects from different orders

1-ACL (articles avec comité de lecture)The aim of this work was to investigate both the biological activity of an entomotoxin, the pea albumin 1b (PA1b), and the presence or absence of its binding site within an array of insect species. The data obtained showed that insect sensitivity was not related to its taxonomic position. Moreover, PA1b was not toxic to several tested microorganisms. However, the binding site was found to be conserved among very different insects, displaying similar thermodynamic constants regardless of the in vivo species sensitivity. The binding site alone was, therefore, not sufficient for toxicity. One exception was the pea weevil, Bruchus pisorum, which was the only tested species without any detectable binding activity. These findings indicate that the binding site probably has an important endogenous function in insects and that adaptation to pea seeds resulted in the elimination of the toxin binding activity in two independent insect lineages. Other mechanisms are likely to interact with the toxin effects, although they are still largely unknown, but there is no evidence of any specific degradation of PA1b in the midgut of insects insensitive to the toxin, such as Drosophila melanogaster or Mamestra brassicae

A new legume bio-insecticide peptide

International audienc

Incorporation of trifluoromethylated pseudoproline in an entomotoxic knottin: effects on oxidative folding, 3D structure and biological activity

International audienc

Incorporation of trifluoromethylated pseudoproline in an entomotoxic knottin: effects on oxidative folding, 3D structure and biological activity

International audienc

Intestinal GCN2 controls Drosophila systemic growth in response to Lactiplantibacillus plantarum symbiotic cues encoded by r/tRNA operons

Symbiotic bacteria interact with their host through symbiotic cues. Here, we took advantage of the mutualism between Drosophila and Lactiplantibacillus plantarum (Lp) to investigate a novel mechanism of host-symbiont interaction. Using chemically defined diets, we found that association with Lp improves the growth of larvae-fed amino acid-imbalanced diets, even though Lp cannot produce the limiting amino acid. We show that in this context Lp supports its host’s growth through a molecular dialogue that requires functional operons encoding ribosomal and transfer RNAs (r/tRNAs) in Lp and the general control nonderepressible 2 (GCN2) kinase in Drosophila’s enterocytes. Our data indicate that Lp’s r/tRNAs are packaged in extracellular vesicles and activate GCN2 in a subset of larval enterocytes, a mechanism necessary to remodel the intestinal transcriptome and ultimately to support anabolic growth. Based on our findings, we propose a novel beneficial molecular dialogue between host and microbes, which relies on a non-canonical role of GCN2 as a mediator of non-nutritional symbiotic cues encoded by r/tRNA operons

Intestinal GCN2 controls Drosophila systemic growth in response to Lactiplantibacillus plantarum symbiotic cues encoded by r/tRNA operons

Symbiotic bacteria interact with their host through symbiotic cues. Here, we took advantage of the mutualism between Drosophila and Lactiplantibacillus plantarum (Lp) to investigate a novel mechanism of host-symbiont interaction. Using chemically defined diets, we found that association with Lp improves the growth of larvae-fed amino acid-imbalanced diets, even though Lp cannot produce the limiting amino acid. We show that in this context Lp supports its host’s growth through a molecular dialogue that requires functional operons encoding ribosomal and transfer RNAs (r/tRNAs) in Lp and the general control nonderepressible 2 (GCN2) kinase in Drosophila ’s enterocytes. Our data indicate that Lp’s r/tRNAs are packaged in extracellular vesicles and activate GCN2 in a subset of larval enterocytes, a mechanism necessary to remodel the intestinal transcriptome and ultimately to support anabolic growth. Based on our findings, we propose a novel beneficial molecular dialogue between host and microbes, which relies on a non-canonical role of GCN2 as a mediator of non-nutritional symbiotic cues encoded by r/tRNA operons

Fabrication of Flexible, Transparent and Conducting Carbon Nanotube Films

奈米碳管由於本身具有的優異導電性與力學可撓性為目前新興發展的熱門材料，而當中的多層奈米碳管，更由於其低廉的材料成本與簡易的製造設備可望直接取代氧化銦錫 ( ITO ) 薄膜而應用於各類螢幕顯示器。然而至今為止，奈米碳管受限於管狀石墨管壁的特性，而難以有效組裝近而獲得品質優良的奈米碳管薄膜。在本論文中，結合超音波霧化與旋轉塗佈成功製備奈米碳管薄膜於聚對苯二甲二乙酯 ( PET ) 基板上，後續，更藉由管壁銀/鉑合金奈米顆粒的附著、聚乙烯醇 ( PVA ) 的表面塗佈保護、熱壓等方法，近一步提高奈米碳管薄膜的透光導電效能與耐撓性。而在本論文中所獲得的多層奈米碳管薄膜可達到 550nm 波長光穿透率 85%、片電阻值 150 歐姆平方、並且可以忍受高達500次以上的高角度折彎。Mechanical flexibility is essential for carbon nanotube ( CNT) films used in touch screens. So far, a robust network composed of individual CNTs is difficult to fabricate because their cohesion was limited by weak Van der Waals forces. Here we create hybrid composite films constructed from multi-walled carbon nanotube (MWCNT)-supported Ag/Pt alloy nanoparticles. Through the combination of ultrasonic atomization and spin coating methods, the Ag/Pt-MWCNT hybrid network on the flexible PET substrate have been achieved at room temperature. The hybrid network with 80% transparency at 550 nm exhibits a 154-W/sq sheet resistance, which is superior to that of single-walled CNT. Importantly, the corresponding sheet conductance exhibits no degradation even after the film was flexed and folded more than 500 times. This study may offer a direct alternative to indium tin oxide ( ITO ) and other transparent conducting oxides.序論.............................................................................................................1 1-1. 奈米碳管介紹.............................................................................2 1-2. 傳統透光導電薄膜...................................................................36 1-3. 奈米碳管用於透光導電薄膜...................................................47 1-4. 奈米碳管透光導電薄膜分類與製備.......................................99 1-5. 奈米碳管透光導電薄膜應用.................................................168 結果與討論.............................................................................................190 2-1. 研究流程.................................................................................190 2-2. 奈米碳管前處理.....................................................................195 2-2-1 機械式預分散處理........................................................210 2-2-2 強酸強鹼純化處理........................................................218 2-2-3 熱處理............................................................................232 2-3. 奈米碳管分散.........................................................................246 2-4. 透光導電薄膜.........................................................................328 2-4-1. 薄膜製備.......................................................................328 2-4-2. 薄膜檢測.......................................................................360 2-4-3. 薄膜相關參數影響.......................................................397 2-4-4. 多次轉印.......................................................................513 2-5. 奈米金屬粒子複合透光導電薄膜.........................................546 2 2-5-1. 製備奈米碳管單金屬奈米顆粒複合物.......................576 2-5-2. 製備奈米碳管雙金屬奈米顆粒複合物.......................586 2-5-3. 奈米碳管金屬複合透光導電薄膜製備.......................602 2-5-4. 奈米碳管金屬複合透光導電薄膜後處理...................612 2-2-5. 奈米碳管金屬複合薄膜薄膜檢測...............................637 結論與未來展望....................................................................................645 實驗部份.................................................................................................647 3-1. 藥品與樣品材料.....................................................................647 3-2. 儀器.........................................................................................663 3-3. 設備.........................................................................................667 3-4. 樣品製備與檢測步驟.............................................................676 引用文獻.................................................................................................68