昆虫抗茵肽的生理活性及其转基因应用前景

昆虫抗菌肽是昆虫免疫后存在于血淋巴中的一类活性肽.根据分子的结构可分为5类天蚕素类、昆虫防御素、富含脯氨酸或精氨酸的抗菌肽、富含甘氨酸的抗菌肽、抗真菌肽.且具有广谱的抗菌、抗病毒、抑制肿瘤的生物活性.概述了昆虫抗菌肽的基因的克隆与表达及转基因研究方面的进展,并展望了抗菌肽在基因工程中的应用前景.     

昆虫抗菌肽的研究进展

昆虫抗菌肽是昆虫免疫后存在于血淋巴中的一类肽类活性物质.具有相对分子质量小、热稳定性强、无免疫原性、抗菌谱广等特点.结合昆虫抗菌肽的研究现状和发展前景,对昆虫抗菌肽的分类、作用机理、基因工程等方面进行了综述.     

昆虫抗菌肽基因工程及其分子设计研究进展(英文)

昆虫抗菌肽是昆虫免疫后存在于血淋巴中的一类活性肽,具有广谱抗菌的生物活性,因而昆虫抗菌肽基因工程迅速成为生命科学领域的研究热点之一.介绍了昆虫抗菌肽的基因结构和表达,综述了昆虫抗菌肽的人工合成与改造以及转基因工程的研究成果,概述了昆虫抗菌肽的分子设计方法和现状,并展望了昆虫抗菌肽的应用前景.     

两栖类动物皮肤分泌物及其生物学适应意义--大蹼铃蟾皮肤分泌物蛋白质多肽组的启示

以大蹼铃蟾(Bombina maxima)为代表性物种,揭示了两栖类皮肤分泌物蛋白质多肽组丰富的分子和功能多样性。目前在大蹼铃蟾已发现的分子包括3类不同的抗菌肽、缓激肽、缓激肽增强肽和缓激肽拮抗肽、缓激肽基因相关肽、富含脯氨酸铃蟾肽及其基因相关肽、神经调节素U、Bv8肽、三叶型蛋白和蛋白酶抑制剂等。抗菌肽分子多样性及其形成机制、缓激肽及其基因相关肽功能和表达模式的多样性都较好地揭示了在多样的生态条件下,两栖类皮肤活性肽环境适应的分子基础及生物合成的调控机制。富含血红素辅基的白蛋白广泛分布在大蹼铃蟾皮肤外皮层细胞膜上和真皮海绵层内,表明它在皮肤的生理功能,如在呼吸、物质交换和渗透压调节中有重要作用。两栖类皮肤分泌物蛋白质多肽组,由于其分子结构的多样性、新颖性和哺乳类同系物的存在,在生物医学研究和天然药物开发中具有独特和不可替代的价值;同时,两栖类皮肤功能基因组具有多样性丰富、快速重组突变的特征,是探讨生物适应的基因基础、基因形成机制和进化特性等生物学基本问题的优秀模型。     

本期重点推介

昆虫抗菌肽因其重要的可开发利用价值愈来愈受到人们的重视。家蚕Bombyx mori基因组全测序完成后,经注释发现了35条拟抗微生物肽基因序列,但是这些拟抗微生物肽基因是否具有抗菌活性,是否与其他昆虫中鉴定报道的同种抗菌肽具有相似的抗菌谱和抗菌活性,在同一基因组中的同种抗菌     

小菜蛾Gloverin like抗菌肽对蝉拟青霉侵染的响应研究

抗菌肽是昆虫天然免疫的重要成分,在抵御病原物的侵染中发挥着重要的作用。昆虫Gloverin抗菌肽是一类仅在鳞翅目昆虫中存在的富含甘氨酸抗菌肽。本研究在转录组测序的基础上对小菜蛾抗菌肽gloverin like基因进行了克隆,其开放阅读框序列全长519 bp,编码172 aa,信号肽为1-17 aa。序列分析表明其序列中甘氨酸的比例为15%,与鳞翅目昆虫Gloverin抗菌肽具有较高的同源性。qRT-PCR结果表明小菜蛾抗菌肽gloverin like基因在脂肪体和血细胞组织中表达量较高。蝉拟青霉、金黄色葡萄球菌及大肠杆菌均可诱导小菜蛾抗菌肽gloverin like基因上调表达,蝉拟青霉诱导12 h后表达量达到最高。利用LC-MS方法在蝉拟青霉侵染的小菜蛾幼虫血淋巴中鉴定出小菜蛾Gloverin like、Gloverin抗菌肽以及Lysozyme II、Transferrin、Prophenoloxidase等抗菌效应蛋白。与对照相比,蝉拟青霉侵染后小菜蛾Toll通路4个免疫基因(βGBP、Toll、Cactus和Dorsal)表达量上升。本研究在抗菌肽方面为进一步研究小菜蛾抵御病原真菌入侵的分子机制提供基础信息,也为新的害虫防治方法提供了思路和靶标。     

昆虫抗菌肽和抗真菌肽结构与功能的关系及分子设计

在对GenBank和EMBL数据库中登录的昆虫抗微生物肽 (antimicrobial peptide, AMP), 即昆虫抗菌肽 (antibacterial peptide) 和抗真菌肽 (antifungal peptide, AFP) 进行归类整理的基础上,对天蚕素族(cecropins )抗菌肽结构与功能的关系及人工改造的分子设计策略,特别是对目前新发现的一些昆虫抗真菌肽的已知结构与功能关系的研究进展、存在问题等进行了简要介绍和分析,为从事昆虫抗微生物肽的理论研究和发展新型抗生素药物提供了必要的信息。     

昆虫抗菌肽的结构和功能研究现状

昆虫抗菌肽是昆虫血淋巴中的一类活性肽。在昆虫感染病菌或体壁受到损伤的情况下其脂肪体能迅速合成一系列抗菌活性物质并释放到血淋巴中发挥作用,抑杀多种致病菌、真菌、病毒等,但不破坏生物体的正常细胞。对昆虫抗菌肽的结构特征、生物活性及抗菌肽分子的作用模式的最新研究进展作了综述。     

融合有酸性片段的抗菌肽串连重复基因的构建

为提高抗菌肽的表达,设计在抗菌肽基因的N端融合一段编码酸性肽的片段以及减轻表达产物对宿主的毒性,通过含有酶切位点的接头将该融合肽基因以同向串连的方式连接成多拷贝基因,克隆至pUC19载体。为此,分段设计合成了编码天蚕素A-蜂毒素杂合肽和酸性肽的DNA片段。首先将其连接成融合肽全基因,然后分别与含相同粘性末端的前后接头连接。通过控制基因和接头加入的量及次序,可得到两侧有EcoRI和SalI酶切位点的同向串连的多拷贝基因。选取合适拷贝数的基因,将其克隆至pUC19载体,PCR扩增和DNA测序证明多拷贝基因构建成功且基因方向相同。结果表明,该方法能简捷高效地获得所需的多拷贝基因,为提高表达产物的量打下基因。     

昆虫抗真菌肽

随着对昆虫抗菌肽理论和应用研究的不断深入,已陆续发现了10多种昆虫抗真菌肽。文章就昆虫抗真菌肽的类型、结构和功能的关系、作用机理、应用等方面的新进展进行简要介绍和分析,为昆虫抗真菌肽的理论研究和发展新型抗生素等提供了必要的信息。     

昆虫抗菌肽分类及在医学中应用

昆虫是地球上种类最为繁多的生物,其抗菌肽的种类和应用范围也远多于其他生物产生的抗菌肽。随着越来越多昆虫抗菌肽的发现及对其深入的研究,昆虫抗菌肽的结构和作用机制也被逐一阐明,并广泛应用于畜牧、食品工业及医药等领域。然而,由于某些限制因素,昆虫抗菌肽还未应用于临床。为了加快昆虫抗菌肽在临床中的应用,本文将从昆虫抗菌肽的结构分类、潜在的医学应用以及昆虫抗菌肽的生产研究现状等方面作一综述,以期为昆虫抗菌肽在抗细菌、抗病毒、抗肿瘤及抗寄生虫药物等方面的医用研发提供文献支持。     

The inducible antibacterial peptides of insects

Insects respond to bacterial challenge by the rapid and transient synthesis of a large number of potent antibacterial peptides that are active against many different bacteria. Two families of inducible antibacterial peptides are well characterized: the cecropins and the insect defensins. A rapidly increasing number of proline- and glycine-rich peptides are reported from various insect species together with cecropins and insect defensins. In this review, Stéphane Cociancich, Philippe Bulet, Charles Hetru and Jules A. Hoffmann give an update of our current information on the induced antibacterial peptides.     

昆虫抗菌肽及其基因工程研究进展

昆虫抗菌肽是昆虫免疫后存在于血淋巴中的一类活性肽,具有广谱的抗菌、抗病毒、抑制肿瘤的生物活性,具有很高的应用潜力。本文主要介绍昆虫抗菌肽的类型、生理活性、基因的克隆与表达及转基因工程研究进展。 Insect Antibacterial Peptides and Its Progress in Advance on Genetic Engineering GONG Xia,YUE Guo-wei,SHI Yong-hui Laboratory for Animal Nutrition Research,College of Food Science,Southern Yangtze University,Wuxi 214036,China Abstract:Insect antibacterial peptides produced are kinds of bioactive peptides in hameolymph hen insect has been immunized.These peptides have the wide range of antibacterial,antiviral spectrum and inhibiting growth of tumor cell.The types of antibacterial peptides and its physiological activities,cloning and expression of the genes and transgenetic research are summarized.Its application on medicine and gene engineering are prospected. Key words：insect antibacterial peptides; physiological activity; gene     

Antibacterial peptides isolated from insects.

Insects are amazingly resistant to bacterial infections. To combat pathogens, insects rely on cellular and humoral mechanisms, innate immunity being dominant in the latter category. Upon detection of bacteria, a complex genetic cascade is activated, which ultimately results in the synthesis of a battery of antibacterial peptides and their release into the haemolymph. The peptides are usually basic in character and are composed of 20-40 amino acid residues, although some smaller proteins are also included in the antimicrobial repertoire. While the proline-rich peptides and the glycine-rich peptides are predominantly active against Gram-negative strains, the defensins selectively kill Gram-positive bacteria and the cecropins are active against both types. The insect antibacterial peptides are very potent: their IC50 (50% of the bacterial growth inhibition) hovers in the submicromolar or low micromolar range. The majority of the peptides act through disintegrating the bacterial membrane or interfering with membrane assembly, with the exception of drosocin, apidaecin and pyrrhocoricin which appear to deactivate a bacterial protein in a stereospecific manner. In accordance with their biological function, the membrane-active peptides form ordered structures, e.g. alpha-helices or beta-pleated sheets and often cast permeable ion-pores. Their cytotoxic properties were exploited in in vivo studies targeting tumour progression. Although the native peptides degrade quickly in biological fluids other than insect haemolymph, structural modifications render the peptides resistant against proteases without sacrificing biological activity. Indeed, a pyrrhocoricin analogue shows lack of toxicity in vitro and in vivo and protects mice against experimental Escherichia coli infection. Careful selection of lead molecules based on the insect antibacterial peptides may extend their utility and produce viable alternatives to the conventional antimicrobial compounds for mammalian therapy.     

Insect immunity. Isolation from the lepidopteran Heliothis virescens of a novel insect defensin with potent antifungal activity

Lepidoptera have been reported to produce several antibacterial peptides in response to septic injury. However, in marked contrast to other insect groups, no inducible antifungal molecules had been described so far in this insect order. Surprisingly, also cysteine-rich antimicrobial peptides, which predominate in the antimicrobial defense of other insects, had not been discovered in Lepidoptera. Here we report the isolation from the hemolymph of immune induced larvae of the lepidopteran Heliothis virescens of a cysteine-rich molecule with exclusive antifungal activity. We have fully characterized this antifungal molecule, which has significant homology with the insect defensins, a large family of antibacterial peptides directed against Gram-positive strains. Interestingly, the novel peptide shows also similarities with the antifungal peptide drosomycin from Drosophila. Thus, Lepidoptera appear to have built their humoral immune response against bacteria on cecropins and attacins. In addition, we report that Lepidoptera have conferred antifungal properties to the well conserved structure of antibacterial insect defensins through amino acid replacements.     

Isolation, characterization and chemical synthesis of a new insect defensin from Chironomus plumosus (Diptera)

Injection of low doses of bacteria into the aquatic larvae of the dipteran insect Chironomus plumosus induces the appearance in their hemolymph of a potent antibacterial activity. We have isolated two 36-residue peptides from this hemolymph which are active against Gram-positive bacteria. The peptides are novel members of the insect defensin family and their sequences present marked differences with those of insect defensins isolated from other dipteran species. We have developed a method for efficient renaturation of this cysteine-rich molecule and obtained a highly pure synthetic Chironomus defensin.     

Novel antibacterial peptides induced by probiotics in Hermetia illucens (Diptera: Stratiomyidae) larvae

There is a need to discover new therapeutic substances due to the emergence of deadly infectious diseases and various antibiotic resistance. We focused on the larvae that are utilized as a medical insect for the treatment of skin damage in Europe and America. This study was to investigate the pharmacological activities of novel antibacterial peptides isolated from Hermetia illucens larvae against the Klebsiella pneumoniae and Shigella dysenteriae. The larvae were immunized by probiotics (Lactobacillus casei) for 24 h. The hemolymph from the immunized larvae was fractionated through reverse‐phase chromatography. Peptides were purified using HPLC and the coomassie blue staining, and identified using Nano‐LC‐ESI‐MS/MS system. Antibacterial activities of the peptides were evaluated by turbidometric assay, liquid broth dilution assay, resazurin assay, and agar disk diffusion method. The minimum inhibitory concentrations (MICs) of the peptides were measured as 150 μg/mL through the turbidometric, liquid broth dilution, and resazurin assays. The peptides effectively inhibited their growth/proliferation as well as the survival rate of the tested bacteria. Furthermore, the immunized larvae exhibited overexpression of the peptides compared to non‐immunized larvae. These results demonstrate that the peptides induced by H. illucens exert strong antibacterial activity against Gram‐negative bacteria. The results suggest that the activation of the humoral immunity induced by immunization functioned to enhance the production of antibacterial peptides from the insect and their antibacterial properties. This study indicates the potential of the peptides produced from larvae as antibacterial peptide substance for the development of novel antibacterial drugs.