Novel Bacillus thuringiensis δ-endotoxin active against Locusta migratoria manilensis

A novel δ-endotoxin gene was cloned from a Bacillus thuringiensis strain with activity against Locusta migratoria manilensis by PCR-based genome walking. The sequence of the cry gene was 3,432 bp long, and it encoded a Cry protein of 1,144 amino acid residues with a molecular mass of 129,196.5 kDa, which exhibited 62% homology with Cry7Ba1 in the amino acid sequence. The δ-endotoxin with five conserved sequence blocks in the amino-terminal region was designated Cry7Ca1 (GenBank accession no. {"type":"entrez-nucleotide","attrs":{"text":"EF486523","term_id":"1121787749","term_text":"EF486523"}}EF486523). Protein structure analysis suggested that the activated toxin of Cry7Ca1 has three domains: 227 residues forming 7 α-helices (domain I); 213 residues forming three antiparallel β-sheets (domain II); and 134 residues forming a β-sandwich (domain III). The three domains, respectively, exhibited 47, 44, and 34% sequence identity with corresponding domains of known Cry toxins. SDS-PAGE and Western blot analysis showed that Cry7Ca1, encoded by the full-length open reading frame of the cry gene, the activated toxin 1, which included three domains but without the N-terminal 54 amino acid residues and the C terminus, and the activated toxin 2, which included three domains and N-terminal 54 amino acid residues but without the C terminus, could be expressed in Escherichia coli. Bioassay results indicated that the expressed proteins of Cry7Ca1 and the activated toxins (toxins 1 and 2) showed significant activity against 2nd instar locusts, and after 7 days of infection, the estimated 50% lethal concentrations (LC₅₀s) were 8.98 μg/ml for the expressed Cry7Ca1, 0.87 μg/ml for the activated toxin 1, and 4.43 μg/ml for the activated toxin 2. The δ-endotoxin also induced histopathological changes in midgut epithelial cells of adult L. migratoria manilensis.     

Alpha-melanocyte stimulating hormone protects retinal pigment epithelium cells from oxidative stress through activation of melanocortin 1 receptor–Akt–mTOR signaling

Patients with age related macular degeneration (AMD) will develop vision loss in the center of the visual field. Reactive oxygen species (ROS)-mediated retinal pigment epithelium (RPE) cell apoptosis is an important contributor of AMD. In this study, we explored the pro-survival effect of α-melanocyte stimulating hormone (α-MSH) on oxidative stressed RPE cells. We found that α-MSH receptor melanocortin 1 receptor (MC1R) was functionally expressed in primary and transformed RPE cells. RPE cells were response to α-MSH stimulation. α-MSH activated Akt/mammalian target of rapamycin (mTOR) and Erk1/2 signalings in RPE cells, which were inhibited by MC1R siRNA knockdown. α-MSH protected RPE cells from hydrogen peroxide (H₂O₂)-induced apoptosis, an effect that was almost abolished when MC1R was depleted by siRNA. α-MSH-mediated S6K1 activation and pro-survival effect against H₂O₂ was inhibited by Akt inhibitors (perifosine, MK-2206 and LY294002). Further, mTOR inhibition by rapamycin, or by mTOR siRNA knockdown, diminished α-MSH’s pro-survival effect in RPE cells. Thus, Akt and its downstream mTOR signaling mediates α-MSH-induced survival in RPE cells. In summary, we have identified a new α-MSH–MC1R physiologic pathway that reduces H₂O₂-induced RPE cell damage, and might minimize the risk of developing AMD.     

Soluble expression, purification and functional identification of a disulfide-rich conotoxin derived from Conus litteratus

Conotoxins are a diverse array of small peptides mostly with multiple disulfide bridges. These peptides become an increasing significant source of neuro-pharmacological probes and drugs as a result of the high selectivity for ion channels and receptors. Usually, the analogue of natural conotoxins is produced by means of chemical synthesis. Here, we present a simple and fast strategy of producing disulfide-rich conotoxins via recombinant expression. By fused with thioredoxin and His tag, a novel O-superfamily conotoxin lt7a was successfully expressed in Escherichia coli and purified, resulting in a high yield of recombinant lt7a about 6 mg/l. The purity of target protein is up to 95% as identified by HPLC results. Whole cell patch-clamp recording revealed that the new conotoxin blocked voltage-sensitive sodium channels in rat dorsal root ganglion neurons, indicating it might be a novel microO-conotoxin.     

Three lysine residues in the common β chain of the interleukin-5 receptor are required for Janus kinase (JAK)-dependent receptor ubiquitination, endocytosis, and signaling

Eosinophils are multifunctional leukocytes implicated in the pathogenesis of numerous inflammatory diseases including allergic asthma and hypereosinophilic syndrome. Eosinophil physiology is critically dependent on IL-5 and the IL-5 receptor (IL-5R), composed of a ligand binding α chain (IL-5Rα), and a common β chain, βc. Previously, we demonstrated that the βc cytoplasmic tail is ubiquitinated and degraded by proteasomes following IL-5 stimulation. However, a complete understanding of the role of βc ubiquitination in IL-5R biology is currently lacking. By using a well established, stably transduced HEK293 cell model system, we show here that in the absence of ubiquitination, βc subcellular localization, IL-5-induced endocytosis, turnover, and IL-5R signaling were significantly impaired. Whereas ubiquitinated IL-5Rs internalized into trafficking endosomes for their degradation, ubiquitination-deficient IL-5Rs accumulated on the cell surface and displayed blunted signaling even after IL-5 stimulation. Importantly, we identified a cluster of three membrane-proximal βc lysine residues (Lys(457), Lys(461), and Lys(467)) whose presence was required for both JAK1/2 binding to βc and receptor ubiquitination. These findings establish that JAK kinase binding to βc requires the presence of three critical βc lysine residues, and this binding event is essential for receptor ubiquitination, endocytosis, and signaling.     

Improvement of xylose utilization in Clostridium acetobutylicum via expression of the talA gene encoding transaldolase from Escherichia coli

Clostridium acetobutylicum ATCC 824 was metabolically engineered for improved xylose utilization. The gene talA, which encodes transaldolase from Escherichia coli K-12, was cloned and overexpressed in C. acetobutylicum ATCC 824. Compared with C. acetobutylicum ATCC 824 (824-WT), the transformant bearing the E. coli talA gene (824-TAL) showed improved ability on xylose utilization and solvents production using xylose as the sole carbon source. During the fermentation of xylose and glucose mixtures with three xylose/glucose ratios (approximately 1:2, 1:1 and 2:1), the rate of xylose consumption and final solvents titers of 824-TAL were all higher than those of 824-WT, despite glucose repression on xylose uptake still existing. These results suggest that the insufficiency of transaldolase in the pentose phosphate pathway (PPP) of C. acetobutylicum is one of the bottlenecks for xylose metabolism and therefore, overexpressing the gene encoding transaldolase is able to improve xylose utilization and solvent production.     

Antitumor mechanism of Se-containing polysaccharide, a novel organic selenium compound

Recent studies on the inhibition of tumor growth by Se-containing polysaccharide were reviewed. Meanwhile, the possible molecular mechanisms of the inhibition of tumor cell growth through antioxidation, induction of tumor cell apoptosis, blockade of cell cycle, and enhancement of immunity by Se-containing polysaccharide were proposed. In the end, the potential application of Se-containing polysaccharide in the prevention and treatment of tumor was elucidated.     

鹿衔草化学成分的研究：羟基肾叶鹿蹄草甙的结构鉴定

从鹿衔草（Ｐｙｒｏｌａ ｃａｌｌｉａｎｔｈａ Ｈ．Ａｎｄｒｅｓ）分离得到两个化合物。经光谱分析,确定其中１个化合物为新化合物,命名为羟基肾叶鹿蹄草甙（１, ｈｙｄｒｏｘｙｌｒｅｎｉｆｏｌｉｎ）,另一化合物为儿茶素（３）。     

蓝斑核对上呼吸道阻力肌—颏舌肌功能的影响

本实验在３８只经氨基甲酸乙酯麻醉的健康家兔上进行,观察了电、化学刺激蓝斑核（ＬＣ）对颏舌肌功能的影响。结果如下：（１）长串电脉冲刺激蓝斑核使颏舌肌肌电活动明显增强,表现为长吸性肌电积分幅度升高,膈肌亦出现与颏舌肌同步的肌电活动增强。（２）ＬＣ内微量注射胞体兴奋剂谷氨酸钠,也引起明显的颏舌肌和膈肌肌电活动增强。（３）上述电、化学刺激的区域对照和盐水对照实验,均未出现有意义的肌电改变。提示：电、化学刺激ＬＣ可特异性增强上呼吸道阻力肌──颏舌肌的紧张性活动,具有减小上呼吸道阻力的作用,这对于某些上呼吸道阻塞性疾病发病机制的研究可能具有重要的意义。     

铁蛋白介导的地衣多糖酶胞内自发聚集及其高效制备

酶分离纯化、固定化及催化性能提升一直是生物催化领域的研究热点和前沿,也是众多研究者致力解决的难点.研究和开发新型的纯化、固定化及提升催化性能的方法,降低纯化及储存等设备的要求及生产成本,对酶大规模应用具有重要意义.文中将铁蛋白(ferritin)与目标酶(地衣多糖酶)基因融合,经高效表达后,在细胞内 自发形成无载体固定...     

东亚中更新世古人类下颔第二臼齿釉质-齿质连接面三维形状和釉质厚度分布

东亚中更新世古人类在头骨、下颌骨、牙齿等解剖部位表现出不同程度的形态多样性,中期成员代表为直立人,而晚期成员的演化地位具有较大争议。为进一步了解东亚中更新世古人类内部的形态变异特点和为东亚中更新世晚期古人类分类提供依据,本文使用微分同胚的表面匹配（Diffeomorphic Surface Matching, DSM）和形态测量图（Morphometric map）对下颌第二臼齿（M₂）釉质-齿质连接面的形状和齿冠侧面釉质厚度分布模式进行了量化分析。结果显示：1）东亚中更新世古人类与晚期人属成员（尼安德特人和现代人）存在较明显的形态差别;2）该时段晚期的东亚古人类相对中期直立人在侧面釉质厚度分布规律上具有独特性,并在釉质-齿质连接面的三维形状上与晚期人属成员更加接近。本文在以往对东亚中更新世古人类牙齿内外结构单个性状研究的基础上,使用三维形态测量方法进一步量化了M₂釉质-齿质连接面三维形状和侧面釉质厚度分布模式两项重要特征的变异特点,这对未来该时段同类型牙齿的形态鉴定以及解决东亚中更新世晚期古人类的分类地位具有一定意义。