异戊二烯生物合成研究进展

异戊二烯(isoprene),又名2-甲基-1、3-丁二烯,是最简单的类异戊二烯化合物,是橡胶的重要前体物质,在精细化工如香料、新型农药等方面应用广泛。异戊二烯主要依赖化石燃料合成,但生产成本较高、易污染环境,生物法合成异戊二烯具有巨大的潜在应用价值,本文综述了生物法合成异戊二烯的主要途径与研究进展。     

生物印迹:一个直接的酶修饰技术

生物印迹技术是一个简单、直接的酶修饰技术,已应用到有机合成、生物传感器制备和生物分离等领域,在推动化学品绿色合成方面具有潜在的应用价值。本文在生物印迹概念的提出与发展、分类、应用以及印迹技术策略的发展等方面综述近期研究成果,针对生物印迹研究过程的现实问题,展望其未来的发展趋势,为生物印迹相关研究提供有益的借鉴。     

环境因子对小麦体内镉的生物毒性和植物络合素合成的影响

采用水培方式,研究了不同环境因子对小麦体内Cd的生物毒性与植物络合素(PCs)合成的影响.结果表明,Cd胁迫对小麦产生明显的毒害效应,并显著诱导根合成PCs;pH、Ca和S对小麦体内Cd的吸收和生物毒性具有不同程度的影响,根中PCs的诱导量与Cd的生物毒性变化表现一致;供磷减轻了Cd胁迫的生物毒性,根中PCs的诱导量也显著降低;镁对Cd胁迫的生物毒性影响甚微,根中PCs的诱导量和Cd的吸收量均未见明显变化.本实验结果证明Cd对PCs的诱导能力与植物体内Cd的毒性之间存在一定的相关关系,可将PCs作为Cd胁迫的生物标记物.     

纳米元素硒的生物合成及生物活性

利用生物载体富集和转化硒是获取活性硒形式的合理途径。一定条件下,生物体对硒的代谢可产生红色纳米级胶体元素硒,传统认为,生物体内元素硒的合成是生物体解除硒毒性的有效机制。最近的一些研究发现,人工合成以蛋白质为分散剂的红色纳米元素硒具有抑制肿瘤细胞生长、抗氧化、免疫调节及延缓衰老等生物活性。提示生物来源的纳米元素硒可能具有潜在应用价值。对纳米元素硒及其生物活性的研究进展进行了综述。     

生物喋呤的生物学效应及其在脓毒症中的意义

研究表明,一氧化氮（ＮＯ）的过度产生可能是诱发脓毒性休克的最后共同通路,而生物喋呤为一氧化氮合酶（ＮＯＳ）重要的辅因子。多种免疫刺激因子均可诱导细胞内ＢＨ４合成显著增加,其可瑟ＮＯＳ紧密结合,调控ＮＯ的合成与释放。本文讨论了ＢＨ４的生物学效应,调控机制及其在脓毒症中的作用,并简要介绍其合成抑制剂在脓毒症防治中的潜在意义。     

甲醇酵母代谢工程研究进展

甲醇酵母由于独特优点被认为是绿色生物制造的潜在宿主。特别是其天然甲醇利用性能有望建立甲醇生物转化路线,拓展生物炼制底物,具有重要经济价值和环保意义。文中综述了代谢工程改造甲醇酵母合成蛋白质和化学品的最新研究进展,并比较了其与模式生物酿酒酵母作为细胞工厂的优缺点。随后,分析了甲醇酵母代谢工程改造面临的挑战,并展望了潜在解决方案。随着基因操作工具开发和细胞代谢阐释,甲醇酵母将在未来绿色生物制造发挥越来越重要的作用。     

芜菁UF3GT基因的克隆及表达特性

目的:克隆‘津田’芜菁和‘赤丸’芜菁的UDP-葡萄糖∶类黄酮3-O-葡萄糖基转移酶(UF3GT)基因并研究其表达特性。方法:利用RT-PCR方法克隆‘津田’芜菁BrUF3GT1基因和‘赤丸’芜菁BrUF3GT2基因,并进行生物信息学分析;通过Northern杂交检测BrUF3GT1和BrUF3GT2基因的UV-A诱导表达特性;对BrUF3GT1和BrUF3GT2基因进行原核诱导表达。结果:BrUF3GT1和BrUF3GT2的开放读框为1407 bp,编码468个氨基酸残基;氨基酸序列分析显示,BrUF3GT1和BrUF3GT2与拟南芥UF3GT的同源性为87%,从第16~453位氨基酸残基的肽段具有糖基转移酶家族成员的结构域;BrUF3GT1和BrUF3GT2基因具有高度同源性,核苷酸序列在7个位点存在差异,推导的氨基酸序列在1个位点存在差异;Northern杂交结果显示,UV-A可以诱导BrUF3GT1和BrUF3GT2基因表达,基因的表达量与处理时间相关;原核诱导表达及纯化后可以获得相对分子质量分别约为51.88×103和51.89×103的BrUF3GT1和BrUF3GT2蛋白。结论:克隆了‘津田’芜菁和‘赤丸’芜菁的UF3GT基因,为初步阐明2种芜菁的花青素生物合成机理奠定了实验基础。     

姜黄素类似物抑制肿瘤细胞增殖的细胞学机制研究进展

姜黄素是一种具有多种生物学效应的天然多酚类物质,但其较低的稳定性和生物利用率等因素限制了其实际临床应用,具有同样类似活性的姜黄素类似物或衍生物目前越来越受到关注。已发现多种化学合成姜黄素类似物小分子能在体外和体内抑制肿瘤细胞增殖,具有潜在的抗肿瘤活性。自噬和凋亡是细胞两种面对外界刺激采取的主动自主行为,近来发现大部分姜黄素类似物小分子能通过诱导自噬或(和)凋亡通路途径引起细胞死亡,该文结合笔者课题组前期研究结果就姜黄素类似物诱导的细胞学机制进行综述,以期为未来有关姜黄素类似物的抗肿瘤药理作用研究提供参考。     

多孔材料的生物合成研究进展

多孔材料以其独特的结构和优异的性能而具有潜在的应用,引起了人们广泛的关注。化学法合成多孔材料,往往造成环境问题。为满足高功能和环境友好化工技术时代的要求,生物技术在利用资源和发展绿色技术方面十分重要。本文中,笔者对生物组织模板技术、微生物模板技术和生物分子自组装技术等应用生物技术合成多孔材料进行了综述。     

光遗传学在细菌生产调控中的应用进展

合成生物学的发展使得人们可以根据需求对微生物进行改造,作为“工厂”高效地合成催化所需物质,并通过添加化学诱导物的方式对生命过程进行调控。然而,化学诱导的潜在毒性以及不可逆性等限制其应用。光遗传学技术利用特定波长的光信号实现对细胞生命过程的调控,具有特异性、可逆性、高时空分辨率等特点。近年来,人们对不同来源的光敏蛋白进行改造,开发出各种不同波长、不同效应的光遗传元件用于基因回路的构建,进而实现对细菌蛋白合成、代谢过程的调控。光遗传技术在人与细菌之间搭起了实时的信号沟通桥梁,实现更为精准的物质生产调控:(1)通过光控治疗因子的合成分泌进行药物递送;(2)通过代谢通路的控制提高目的产物的催化效率;(3)通过光诱导控制生物活材料的形成。随着探索的深入,更小体积、更多波长、更高效率的光遗传元件将被开发出来,实现多输入的细菌生命活动调控。     

Gliotoxin promotes Aspergillus fumigatus internalization into type II human pneumocyte A549 cells by inducing host phospholipase D activation

The internalization of Aspergillus fumigatus into lung epithelial cells is critical for the infection process in the host. Gliotoxin is the most potent toxin produced by A. fumigatus. However, its role in A. fumigatus internalization into the lung epithelial cells is still largely unknown. In the present study, the deletion of the gliP gene regulating the production of gliotoxin in A. fumigatus suppressed the internalization of conidia into the A549 lung epithelial cells, and this suppression could be rescued by the exogenous addition of gliotoxin. At lower concentrations, gliotoxin enhanced the internalization of the conidia of A. fumigatus into A549 cells; in contrast, it inhibited the phagocytosis of J774 macrophages in a dose-dependent manner. Under a concentration of 100 ng/ml, gliotoxin had no effect on A549 cell viability but attenuated ROS production in a dose-dependent manner. Gliotoxin significantly stimulated the phospholipase D activity in the A549 cells at a concentration of 50 ng/ml. This stimulation was blocked by the pretreatment of host cells with PLD1- but not PLD2-specific inhibitor. Morphological cell changes induced by gliotoxin were observed in the A549 cells accompanying with obvious actin cytoskeleton rearrangement and a moderate alteration of phospholipase D distribution. Our data indicated that gliotoxin might be responsible for modulating the A. fumigatus internalization into epithelial cells through phospholipase D1 activation and actin cytoskeleton rearrangement.     

Genotoxicity of gliotoxin, a secondary metabolite of Aspergillus fumigatus, in a battery of short-term test systems

The genotoxic effects of gliotoxin, a known fungal secondary metabolite, were studied. Gliotoxin was purified from cultivation medium of Aspergillus fumigatus isolated from the indoor air of a moisture problem house. The genotoxicity of gliotoxin was assessed both in bacterial test systems including bacterial repair assay, Ames Salmonella assay and SOS-chromotest, and in mammalian cells using single cell gel (SCG) electrophoresis assay and sister-chromatid exchange (SCE) test. Gliotoxin was found to be genotoxic in the bacterial repair assay but, not in the Salmonella test or SOS-chromotest. A dose-related increase in DNA damage was observed in mouse RAW264.7 macrophages exposed to gliotoxin for 2 h in plain medium in the SCG assay. In contrast to the positive response in the SCG assay, gliotoxin did not induce any clear, dose-related increase in SCEs in Chinese hamster ovary (CHO) cells.     

Elucidation of the Role of Sugar Chains in Glycosylated-enzymes Using Endo-γ-N-acetylglu-cosaminidase from Flavo-bacterium sp.

In the course of screening for anti-platelet principles produced by micro-organisms, strong anti-platelet activity was detected in the culture broth of Aspergillus fumigatus Fres. The purified active compound was identified as gliotoxin. Gliotoxin inhibited ADP-induced aggregation as well as collagen- or arachidonate-induced aggregation of rabbit platelets (IC₅₀ = about 27 μm) and also accelerated the dissociation of aggregates. Gliotoxin also inhibited the heat hemolysis of rabbit erythrocytes, suggesting that this agent is a membrane-stabilizing anti-aggregant. The disulfide structure in the gliotoxin molecule was responsible for the inhibitory activity, because des- thiogliotoxin had effects on neither platelet aggregation nor heat hemolysis of erythrocytes.     

Use of thin layer chromatography for detection and high performance liquid chromatography for quantitating gliotoxin from rice cultures ofAspergillus fumigatus Fresenius

Gliotoxin, a mycotoxin with antimicrobial and immunosuppressive capabilities, is produced by several genera of fungi including the pathogenic fungusAspergillus fumigatus. The ability of selected isolates ofA. fumigatus to produce gliotoxin on three different media was tested and a thin layer chromatographic and high performance liquid chromatographic method for quantitation of gliotoxin from rice culture was developed and is described. Rice cultures were extracted with chloroform and the resulting extract was partially purified by precipitation with petroleum ether and cleanup by gel permeation chromatography. Gliotoxin was detected by thin layer chromatography and quantitated by high performance liquid chromatography using a U.V. absorbance detector with a 254 nm filter and a mobile phase of methanol-water 4357 (V/V) with a flow rate of 2.0 ml/min. The retention time for gliotoxin was approximately 4.8 min. From rice samples spiked with gliotoxin concentrations of 0.67, 1.33, 2.67, 4.00 and 5.33g/g the average recovery was 83.8%.     

Effect of Aeration on Gliotoxin Production by Spergillus Fumigatus in its Culture Filtrate

Gliotoxin, one of the mycotoxins produced by Aspergillus fumigatus, has various, potent bioactivities. However, it has not been considered to be a toxic (or virulence) factor because of its slow production. The aim of the present study was to investigate the effects of aeration on the cytotoxicity of A. fumigatus culture filtrate, and to determine the optimal condition for the rapid production of gliotoxin from this fungus. Fungal culture filtrates were made in three different containers under various conditions of aeration and O₂ concentration. These filtrates were compared in terms of their cytotoxicity on murine macrophages and analyzed by gas chromatography. The culture filtrate showed high cytotoxicity when it was made under highly aerated conditions, but it was significantly less cytotoxic when prepared under non-aerated conditions. The cytotoxic activity became evident within 15 h of culture at 20% O₂, when the fungus had already started producing gliotoxin. The culture filtrates also contained some other as yet unidentified substances that might also to some extent contribute to the cytotoxicity. In light of these results, the authors propose that a highly aerated condition is responsible for the rapid production of gliotoxin, and that gliotoxin might play an important role in the respiratory infection by A. fumigatus, with other toxic substances acting additively or synergistically.     

Gliotoxin contamination in and pre- and postfermented corn, sorghum and wet brewer's grains silage in Sao Paulo and Rio de Janeiro State, Brazil

Aims: The aim of this study was to determine total fungal counts and the relative density of Aspergillus fumigatus and related species in silage samples intended for bovines before and after fermentation as well as to monitor the natural occurrence of gliotoxin in silage samples (pre‐ and postfermentation). Methods and methods: The survey was performed in farms located in São Paulo and Rio de Janeiro States in Brazil. In addition, the ability of A. fumigatus strains and related species strains to produce gliotoxin was also evaluated. A total of 300 samples were taken, immediately after opening of the silo (3–5 months) and during the ensiling period. Fungal counts were done by the surface‐spread method. Gliotoxin production ability of isolates and natural contamination were determined by HPLC. Results: All postfermented samples had a total number of moulds exceeding 1 × 10⁴ CFU g^?1, with Aspergillus sp. as the most prevalent genus. Frequency of strains, among A. fumigatus and related species, was able to produce gliotoxin was similar in pre‐ and postfermented samples, except for sorghum, which showed differences between both kinds of samples. The highest toxin levels were produced by strains isolated from postfermented samples. More than 50% of the samples showed gliotoxin contamination levels that exceeded concentrations known to induce immunosuppressive and apoptotic effects in cells. Conclusions: The present data suggest that care should be taken because gliotoxin contamination in feedstuffs could affect productivity and also present a health risk for herds. Significance and Impact of the Study: Gliotoxin was found at quite important concentrations levels in pre‐ and postfermented substrates and its presence could therefore probably affect the productivity and health of herds. Current conservation and management practices do not avoid contamination with A. fumigatus on silage. Therefore, farm workers should be adequately protected during its handling.     

Induction of Gliotoxin Secretion in Aspergillus fumigatus by Bacteria-Associated Molecules

Aspergillus fumigatus is the most common causative agent of mold diseases in humans, giving rise to life-threatening infections in immunocompromised individuals. One of its secreted metabolites is gliotoxin, a toxic antimicrobial agent. The aim of this study was to determine whether the presence of pathogen-associated molecular patterns in broth cultures of A. fumigatus could induce gliotoxin production. Gliotoxin levels were analyzed by ultra-performance liquid chromatography and mass spectrometry. The presence of a bacteria-derived lipopolysaccharide, peptidoglycan, or lipoteichoic acid in the growth media at a concentration of 5 μg/ml increased the gliotoxin concentration in the media by 37%, 65%, and 35%, respectively. The findings reveal a correlation between the concentrations of pathogen-associated molecular patterns and gliotoxin secretion. This shows that there is a yet uncharacterized detection system for such compounds within fungi. Inducing secondary metabolite production by such means in fungi is potentially relevant for drug discovery research. Our results also give a possible explanation for the increased virulence of A. fumigatus during bacterial co-infection, one that is important for the transition from colonization to invasiveness in this pulmonary disease.     

Production of gliotoxin during the pathogenic state in turkey poults byAspergillus fumigatus Fresenius

Turkey poults were given either of two different dosages of two different gliotoxin-producing strains ofAspergillus fumigatus. Infected lung tissue was examined postmortem for the presence of gliotoxin. Gliotoxin was found in lung tissue of ten poults infected with one strain and in seven of ten poults infected with the other strain. Concentrations of gliotoxin in the tissue exceeded 6 ppm in some of the infected tissues. The concentration of gliotoxin found in infected tissue did not appear to be correlated with the dosage of organism given. Considering the pathologic changes observed in turkey poults with aspergillosis and the production of gliotoxin during the pathogenic state in turkey poults, gliotoxin is considered likely to be involved in avian aspergillosis. Disclaimer: Names are necessary to report factually on available data; however, the USDA neither guarantees nor warrants the standard of the product, and the use of the name by USDA implies no approval of the product to the exclusion of others that may also be suitable.     

Gliotoxin inhibits transformation and its cytotoxic to turkey peripheral blood lymphocytes

Gliotoxin, an epipolythiodioxopiperizine mycotoxin, has been shown to be produced by, among other fungi,Aspergillus fumigatus Fresenius. This organism is the major causative agent of the respiratory disease aspergillosis in avian species, especially turkeys. Because gliotoxin has been shown to be immunosuppressive and has the potential for being involved in the pathogenesis of aspergillosis, the in vitro activity of this compound with avian lymphocytes was investigated. Immunosuppression was investigated using peripheral blood lymphocytes from turkeys in a lymphoblastogenesis assay and a cytotoxicity assay using conversion of the tetrazolium salt MTT to MTT formazan by the mitochondrial succinate dehydrogenase enzyme elaborated only by living cells. Gliotoxin appeared to have a threshold level in both tests because little or no response or stimulation was evident when cells were exposed to concentrations of the toxin below 100 ng/ml, but at 100 ng/ml, all cells appeared to be dead. Using T-2 mycotoxin as a known cytotoxic agent, the response in the MTT bioassay using turkey peripheral lymphocytes was linear with increasing concentrations of toxin. Gliotoxin may potentially cause immunosuppression in turkey poults through action on the lymphocytes or if this toxin were present in low concentrations stimulation could possibly occur.     

Natural occurrence of gliotoxin in turkeys infected with Aspergillus fumigatus,Fresenius

Thirteen samples of infected turkey lung tissue from cases of airsacculitis were collected either at the processing plant or from a local turkey farm and subjected to cultural and gliotoxin analysis. Aspergillus fumigatus was isolated from 6 of the 13 samples; all isolates were determined to be gliotoxin producers when grown in laboratory culture and assayed by HPLC procedures. Gliotoxin was isolated from 5 of the 13 tissues but was not isolated from all tissues that were infected with A. fumigatus. Gliotoxin was isolated from two tissues from which no A. fumigatus was isolated and it was not detected in three tissues from which gliotoxin-producing isolates of A. fumigatus were obtained. The ability of this pathogenic fungus to produce this immunomodulating compound in naturally infected turkeys provides further evidence that gliotoxin may be involved in the pathogenesis of the disease, aspergillosis of turkeys. Disclaimer: Names are necessary to report factually on available data; however, the USDA neither guarantees nor warrants the standard of the products, and the use of the name by USDA implies no approval of the product to the exclusion of others that may also be suitable.