一种快速准确区分Ⅲ型、Ⅳ型分泌效应蛋白的计算方法

通过Ⅲ型、Ⅳ型、Ⅵ型分泌系统,革兰氏阴性菌可将效应蛋白直接注入宿主体内,并导致宿主感染各种疾病.由于Ⅲ型、Ⅳ型分泌效应蛋白均属非经典分泌蛋白,且它们可能具有相似的序列模体或进化保守性,故两者之间难于区分.基于支持向量机和伪位置特异性得分矩阵,本文提出了一种可快速准确识别革兰氏阴性菌Ⅲ型、Ⅳ型效应蛋白的计算方法.测试集实验结果表明,本方法对Ⅲ型、Ⅳ型效应蛋白具有较好的分类效果,可作为辅助工具用于分泌效应蛋白的进一步研究.     

一种快速准确区分III型、IV型分泌效应蛋白的计算方法

通过Ⅲ型、Ⅳ型、Ⅵ型分泌系统,革兰氏阴性菌可将效应蛋白直接注入宿主体内,并导致宿主感染各种疾病.由于Ⅲ型、Ⅳ型分泌效应蛋白均属非经典分泌蛋白,且它们可能具有相似的序列模体或进化保守性,故两者之间难于区分.基于支持向量机和伪位置特异性得分矩阵,本文提出了一种可快速准确识别革兰氏阴性菌Ⅲ型、Ⅳ型效应蛋白的计算方法.测试集实验结果表明,本方法对Ⅲ型、Ⅳ型效应蛋白具有较好的分类效果,可作为辅助工具用于分泌效应蛋白的进一步研究.     

嗜水气单胞菌Ⅲ型分泌系统研究进展

主要对嗜水气单胞菌Ⅲ型分泌系统、效应蛋白及其致病机理等方面的研究进展进行了综述.细菌分泌系统的发现是近年来细菌致病机制研究的重要进展,许多革兰氏阴性细菌借助于细菌的分泌系统,分泌出毒性因子和效应蛋白,与寄主进行分子交流.     

革兰氏阴性菌中蛋白质亚细胞定位预测

根据革兰氏阴性菌蛋白不同亚细胞位置、其一级结构中氨基酸含量、氨基酸的关联性及亲疏水性的不同，利用最小离散增量的方法，分别以20个氨基酸组份、400个氨基酸二联体组份及氨基酸亲疏水性在蛋白质上的分布为参数构成离散源，对革兰氏阴性菌蛋白的5类亚细胞定位进行预测，分别用self—consistency方法和Jack-knife方法预测，均取得了较高的预测成功率．     

哈维氏弧菌vgrG基因的原核表达及多克隆抗体的制备

VI型分泌系统(T6SS)是革兰氏阴性菌特有的一种多功能分泌系统,它与致病菌的致病性密切相关;而vgrG则是T6SS系统的结构蛋白之一,它与T6SS的功能直接相关.为了获得哈维氏弧菌vgrG基因的外源重组蛋白及制备其多克隆抗体,本研究以哈维氏弧菌QT520菌株的DNA为模板,在扩增获得vgrG基因后,构建了原核表达载体pET32a-vgrG并将其转入大肠杆菌BL21(DE3)中进行原核表达.结果显示,在16℃和1mmol·L^-1 IPTG诱导20 h的条件下,可诱导重组蛋白rvgrG大量表达,条带大小与预期的分子量(88 kDa)一致;通过Ni²⁺亲和层析柱对融合蛋白rvgrG进行纯化,获得了纯度较高的目的蛋白rvgrG;将rvgrG重组蛋白通过腹腔注入小鼠体内后观察其毒性,结果显示rvgrG对小鼠无毒害作用;进一步用纯化蛋白rvgrG制备多克隆抗体,经ELISA检测,vgrG多克隆抗体的效价高达1∶320 000;蛋白免疫印迹(Western blot)检测表明,本实验所制备的vgrG多克隆抗体特异性强,可为后续进行vgrG蛋白的致病性及致...     

巴东冬凌草的抗菌活性研究

对产自巴东的冬凌草鲜叶的水提物进行了抗菌活性研究,以平板菌落计数法进行了抑菌实验.结果表明:巴东冬凌草鲜叶的水提物具有广谱的抗菌活性,对革兰氏阳性菌和革兰氏阴性菌均有抑制作用,对革兰氏阳性菌的抗菌活性要比对革兰氏阴性菌的高;对枯草芽孢杆菌和绿脓杆菌具有杀灭作用.     

关于7,7,8,8-四氰基对苯醌与药物荷移反应的研究

氟罗沙星(FLX)和左旋氧氟沙星(LVFX)是一类临床广泛应用的人工合成抗菌素.该类药物对革兰氏阳性菌和革兰氏阴性菌均有强的抑制作用.     

副溶血弧菌VopS效应子基因的克隆与原核表达

副溶血弧菌可通过Ⅲ型分泌系统分泌效应毒力蛋白导致真核靶细胞的细胞溶解及细胞凋亡.VopS是Ⅲ型分泌系统中的一个效应毒力因子,能通过抑制NF-κB的活性介导巨噬细胞死亡.实验将效应因子VopS的编码基因克隆到pET28a(+)载体上,成功构建了pET28a(+)-vopS重组质粒,将该质粒转入大肠杆菌BL21 (DE3)中诱导表达,经SDS-PAGE分析,发现在分子量约为43 kDa处有一明显蛋白表达条带.用Ni2+-NTA柱亲和层析获得纯化蛋白,浓度为175 mg/L.为进一步研究副溶血弧菌Ⅲ型分泌系统致病机制提供了材料.     

实验兔巴氏杆菌的血清学(综述)

<正> 多杀性巴氏杆菌是兔的重要细菌病原体。巴氏杆菌病特别是呼吸道病有较高的发病率,目前,巴氏杆菌的血清学和免疫学知识是来自牛、禽,而对兔巴氏杆菌的血清学材料很少。应用革兰氏阴性菌(包括巴氏杆菌)免疫生物学的材料,来提高对兔病的理解,并产生一个预防或控制疾病的基础。本综述的目的有:1.提出革兰氏阴性菌的荚膜和细胞壁的现代理论;2.提出从动物(主要是牛和禽)分离出的巴氏杆菌的血清分类基础;3.将这种分类法应用于兔巴氏杆菌;     

三种木兰科植物茎和叶提取物的抑菌活性研究

木兰科植物木莲(Manglietia fordiana)、含笑(Michelia figo)和北美鹅掌楸(Liriodendrontulipifera)是中国的传统药用植物.本文通过对抑菌圈大小和最小抑菌浓度的测定,研究了这3种植物茎、叶的乙醇粗提物的抑菌活性.结果表明,大部分提取物对革兰氏阳性菌有较好的抑制效果,个别提取物(如木莲茎和含笑茎)能抑制革兰氏阴性菌的生长,所有提取物均无抑制病原真菌的作用.     

桑青枯病病原G12-9内切葡聚糖酶基因的克隆和分类

桑青枯病是一种土传性细菌病害,从广东省桑园发生青枯病的桑树根部分离得到1株病原菌G12-9;经鉴定确定该菌为青枯菌（Ralstonia solanacearum）,该病原菌在TZC固体培养基上呈圆形及不规则圆形,菌落中央呈现淡红色,革兰氏染色成阴性。对G12-9分离株内切葡聚糖酶基因的克隆、序列测定及聚类分析结果表明,桑青枯菌G12-9内切葡聚糖酶属于糖基水解酶家族12。青枯菌最重要的致病性分泌系统为Ⅱ、Ⅲ、Ⅳ型分泌系统,内切葡聚糖酶属于细菌Ⅱ型分泌系统,内切葡聚糖酶对于青枯菌的定植及寄主植物的致病性有着非常重要的作用,明确该酶在糖基水解酶家族的分类地位对桑青枯病的防治具有重要意义。     

土法造纸木素降解之探索：Ⅰ.嫩毛竹自然发酵过程微生物数量的变化

嫩毛竹以水自然发酵，不同间隔时间取样计细菌数，用显微镜直接计数，平皿菌落计数及稀释计数（最大可能值）。计数结果，自始至终均以革兰氏阴性杆菌为主，其次为有芽孢菌，总菌数最高时为第３０－４９ｄ。３种计数法均有其优劣。这些细菌的鉴定及其与木素生物降解的关系将见于后续报告。     

Protein delivery into eukaryotic cells by type III secretion machines

Bacteria that have sustained long-standing close associations with eukaryotic hosts have evolved specific adaptations to survive and replicate in this environment. Perhaps one of the most remarkable of those adaptations is the type III secretion system (T3SS)--a bacterial organelle that has specifically evolved to deliver bacterial proteins into eukaryotic cells. Although originally identified in a handful of pathogenic bacteria, T3SSs are encoded by a large number of bacterial species that are symbiotic or pathogenic for humans, other animals including insects or nematodes, and plants. The study of these systems is leading to unique insights into not only organelle assembly and protein secretion but also mechanisms of symbiosis and pathogenesis.     

湄洲湾天然菌群对石油烃的降解作用

从湄洲湾海域内湾及中湾的３个不同站位采集水样,测定它们对大庆原油的自然降解率,结果表明,营养盐及原油的初始浓度对降解作用有很大的影响,将采集的水样经富集培养后,分离、纯化、挑选出３８个能在以原油为唯一碳源的平板上生长的菌落,经鉴定这些菌株多数为革兰氏阴性菌,从微生物细胞的表面结构及其与界面的作用分析海洋烃降解菌革兰氏阴性菌占优势的原因。     

Chaperone release and unfolding of substrates in type III secretion

Type III protein secretion systems are essential virulence factors of many bacteria pathogenic to humans, animals and plants. These systems mediate the transfer of bacterial virulence proteins directly into the host cell cytoplasm. Proteins are thought to travel this pathway in a largely unfolded manner, and a family of customized cytoplasmic chaperones, which specifically bind cognate secreted proteins, are essential for secretion. Here we show that InvC, an ATPase associated with a Salmonella enterica type III secretion system, has a critical function in substrate recognition. Furthermore, InvC induces chaperone release from and unfolding of the cognate secreted protein in an ATP-dependent manner. Our results show a similarity between the mechanisms of substrate recognition by type III protein secretion systems and AAA + ATPase disassembly machines.     

海星内生细菌的分离与初步鉴定

从罗氏海盘车（Aster rollestoni Bell）的胃、幽门盲囊和生殖腺内共分离到20株菌株．其中从其胃中分离到14株,幽门盲囊中分离到5株,生殖腺中分离到1株．根据菌落及菌体形态特征和生理生化实验结果初步认定海星胃中的W1^#、幽门盲囊中的Y1^#以及生殖腺中的S1^#为同一菌株,另外海星胃中的两株菌株（W13^#、W2^#）分别与幽门盲囊中的两株菌株（Y3^#、Y4^#）相同．所得菌株中只有一株为革兰氏阳性球菌,其余为革兰氏阴性杆菌．实验表明海星内生细菌中有62．5％的菌株分属于假单胞菌属（Pseudomonas）和弧菌属（Vibrio）．     

Type VI secretion requires a dynamic contractile phage tail-like structure

Type VI secretion systems are bacterial virulence-associated nanomachines composed of proteins that are evolutionarily related to components of bacteriophage tails. Here we show that protein secretion by the type VI secretion system of Vibrio cholerae requires the action of a dynamic intracellular tubular structure that is structurally and functionally homologous to contractile phage tail sheath. Time-lapse fluorescence light microscopy reveals that sheaths of the type VI secretion system cycle between assembly, quick contraction, disassembly and re-assembly. Whole-cell electron cryotomography further shows that the sheaths appear as long tubular structures in either extended or contracted conformations that are connected to the inner membrane by a distinct basal structure. These data support a model in which the contraction of the type VI secretion system sheath provides the energy needed to translocate proteins out of effector cells and into adjacent target cells.     

Type VI secretion delivers bacteriolytic effectors to target cells

Peptidoglycan is the major structural constituent of the bacterial cell wall, forming a meshwork outside the cytoplasmic membrane that maintains cell shape and prevents lysis. In Gram-negative bacteria, peptidoglycan is located in the periplasm, where it is protected from exogenous lytic enzymes by the outer membrane. Here we show that the type VI secretion system of Pseudomonas aeruginosa breaches this barrier to deliver two effector proteins, Tse1 and Tse3, to the periplasm of recipient cells. In this compartment, the effectors hydrolyse peptidoglycan, thereby providing a fitness advantage for P. aeruginosa cells in competition with other bacteria. To protect itself from lysis by Tse1 and Tse3, P. aeruginosa uses specific periplasmically localized immunity proteins. The requirement for these immunity proteins depends on intercellular self-intoxication through an active type VI secretion system, indicating a mechanism for export whereby effectors do not access donor cell periplasm in transit.     

Signal sequence mutations disrupt feedback between secretion of an exported protein and its synthesis in E. coli

Recent studies in a eukaryotic system indicate that a block in secretion can lead to a block in the translation of secretory proteins. This feedback on protein synthesis is thought to be a result of an interaction of the signal recognition particle with the signal sequences of nascent proteins. Genetic studies in the prokaryote Escherichia coli suggest that a complex secretion machinery and a similar feedback mechanism exist. In addition, mutations affecting two genes, secA and secC, thought to encode components of the bacterial secretion machinery, selectively interfere with the synthesis of exported proteins. This selective interference with translation may be a result of recognition by the secretion machinery of signal sequences. If so, alteration of the signal sequence of a particular protein by mutation should eliminate the block in synthesis for that protein. We show here that signal sequence mutants for an exported protein, maltose binding protein, prevent the block in synthesis of this protein in a secA mutant.