Crystal structures of Nipah and Hendra virus fusion core proteins

The Nipah and Hendra viruses are highly pathogenic paramyxoviruses that recently emerged from flying foxes to cause serious disease outbreaks in humans and livestock in Australia, Malaysia, Singapore and Bangladesh. Their unique genetic constitution, high virulence and wide host range set them apart from other paramyxoviruses. These characteristics have led to their classification into the new genus Henpavirus within the family Paramyxoviridae and to their designation as Biosafety Level 4 pathogens. The fusion protein, an enveloped glycoprotein essential for viral entry, belongs to the family of class I fusion proteins and is characterized by the presence of two heptad repeat (HR) regions, HR1 and HR2. These two regions associate to form a fusion-active hairpin conformation that juxtaposes the viral and cellular membranes to facilitate membrane fusion and enable subsequent viral entry. The Hendra and Nipah virus fusion core proteins were crystallized and their structures determined to 2.2 A resolution. The Nipah and Hendra fusion core structures are six-helix bundles with three HR2 helices packed against the hydrophobic grooves on the surface of a central coiled coil formed by three parallel HR1 helices in an oblique antiparallel manner. Because of the high level of conservation in core regions, it is proposed that the Nipah and Hendra virus fusion cores can provide a model for membrane fusion in all paramyxoviruses. The relatively deep grooves on the surface of the central coiled coil represent a good target site for drug discovery strategies aimed at inhibiting viral entry by blocking hairpin formation.     

苹果新品种"昌红"的离体培养与规模化快速繁殖

1植物名称苹果(Malus pumila Mill.)新品种"昌红"(图1). 2材料类别茎尖. 3培养条件以MS为基本培养基.(1)启动培养基:MS 6-BA 2.0 mg·L-1 (单位下同) NAA 0.5 3%蔗糖;(2)增殖培养基:MS 6-BA 1.5 NAA 0.05 3%白砂糖;(3)生根培养基:1/2MS IBA 0.7 IAA0.2 2%白砂糖.以上培养基均加入6 g·L-1 琼脂,pH 5.8,在121℃、0.11MPa灭菌20 min.培养温度25～27℃,光照12 h·d-1,光照度2 000 lx.     

弗氏志贺菌磷酸化蛋白的检测

目的:检测弗氏志贺菌全菌蛋白中被磷酸化修饰的蛋白。方法:制取弗氏2a志贺菌2457T野生株全菌磷酸化蛋白样品时加入磷酸化酶抑制剂,随后对样品进行双向电泳,以抗磷酸丝氨酸/苏氨酸/酪氨酸抗体为免疫探针,通过Western印迹找到被磷酸化的蛋白,并进行胶内酶解及MALDI-TOF质谱分析。结果与结论:共检测到13个磷酸化蛋白,其中9个为代谢途径中的酶。     

鉴别检测非洲猪瘟野毒株和基因缺失疫苗株的三重荧光PCR的建立

非洲猪瘟是由非洲猪瘟病毒(African swine fever,ASFV)引起的猪的一种急性、高度接触性传染病.本研究根据ASFV B646L、MGF360-14L和CD2v基因设计了 3套引物和探针,建立了可同时鉴别检测ASFV野毒株和基因缺失疫苗株的三重荧光PCR方法.结果表明,该法特异性强,可同时检测ASFV B646L、MGF360-14L和CD2v基因,与猪圆环病毒2型、猪瘟病毒、猪繁殖与呼吸综合征病毒、口蹄疫病毒、猪A型塞内卡病毒等无交叉反应;两份ASFV MGF360-505R与CD2v基因联合缺失疫苗株DNA三重荧光PCR法检测结果都仅出现一条B646L扩增曲线,与预期一致;灵敏度高,该法对重组质粒pUC-B646L、pUC-360-14L和pUC-CD2v的检出限分别为1.572×103拷贝/mL、1.934×103拷贝/mL 和 1.929×103拷贝/mL,灵敏度比世界动物卫生组织(World organization for animal health,OIE)推荐的荧光PCR高10倍;重复性好,对三种重组质粒检测的Ct值批内和批间变异系数均小于2％;应用该法及OIE推荐的荧光PCR对1 215份样品进行平行检测,结果显示23份样品为阳性,1 192份样品为阴性,两种方法检测结果一致.本研究建立的方法能鉴别检测ASFV野毒株和基因缺失疫苗株(缺失MGF360-505R和/或CD2v基因),对将来基因缺失疫苗的推广应用及ASF疫情防控具有重要的意义.     

Stereoselective Accumulation of Propranolol Enantiomers in K562 and K562/ADR Cells

The stereoselective uptake of propranolol enantiomers was investigated by using the K562 and K562 adriamycin‐resistant cell line (K562/ADR) as a model. An enantioselective RP‐HPLC method was applied to determine the accumulation of propranolol (PPL) stereoisomers in K562 and K562/ADR cells. The concentration, time and temperature dependent studies showed that the accumulation of S‐(?)‐PPL was higher than R‐(+)‐PPL in K562 cells and uptake of R‐(+)‐PPL was significantly higher than that of S‐(?)‐PPL in K562/ADR cells. The results indicate the enantioselective accumulation of propranolol enantiomers in K562 and K562 / ADR cells. Chirality 25:361–364, 2013. © 2013 Wiley Periodicals, Inc.     

基于蛋白表达分析的干酪乳杆菌ATCC393交互胁迫保护机制

为考察干酪乳杆菌典型株ATCC393在交互胁迫环境下的生理应答机制,应用二维电泳和iTRAQ技术在蛋白水平上比较了交互胁迫前后干酪乳杆菌蛋白质组的变化情况。在对不同处理条件下细胞全蛋白的二维电泳分析中发现,干酪乳杆菌的主要蛋白分布在等电点pI4～7的范围,经酸预适应处理后细胞的蛋白表达产生了较大的变化。通过iTRAQ技术对细胞在酸适应前后以及相应致死条件下蛋白表达的定性及相对定量分析得知,酸诱导所产生的热胁迫应激蛋白（dnaK,dnaJ等）、氧胁迫应激蛋白（mutS,YeaO）以及与代谢相关的酶类上调可能是提高细胞对交互胁迫耐受能力的主要原因,而酸适应后GTP环化水解酶I和GMP合成酶的高表达可能与这一过程的诱导有关。上述研究结果为提高工业生产菌株在发酵生产及加工过程中对外界不利环境的抵御能力,进而通过调控与微生物生理应答机制密切相关的功能元器件实现生产菌株的性能强化提供了重要的生物信息和可借鉴的研究思路。     

多杀菌素和氟虫睛对瓢虫、寄生蜂和菜缢管蚜选择性的研究

采用药膜法研究了多杀菌素和氟虫睛对3种瓢虫、2种寄生蜂和菜缢管蚜的选择性。与菜缢管蚜相比,2种杀虫剂对3种瓢虫的毒性较低(LC50比值=1.73-41.9),尤其是氟虫睛对3种瓢虫以及多杀菌素对稻红瓢虫均具有较好的选择性(LC50比值大于5)。菜蚜茧蜂和蚜虫宽缘金小蜂对2种杀虫剂的敏感性远高于3种瓢虫和菜缢管蚜,2种杀虫剂对菜蚜茧蜂和蚜虫宽缘金小蜂均是高毒的(LC50比值=0.006-0.05)。3种瓢虫对多杀菌素和氟虫睛具有较高的耐药性,这将有助于其在生物防治中的应用。     

昆虫体内的天然屏障——围食膜

昆虫围食膜是由昆虫中肠上皮细胞分泌的非细胞薄膜状结构,主要成份是几丁质、蛋白质和多糖,是昆虫抵御外界侵害的第一道天然屏障,能够保护中肠上皮细胞不受机械损伤并且能够抵御病毒、细菌及其他有害物质,防止化学损伤.昆虫病毒增效蛋白、荧光增白剂和几丁质酶等生物防治促进因子通过与围食膜上特异位点的结合,能够破坏围食膜结构,加速病原微生物对害虫的感染进程.就围食膜组分、结构、功能以及与害虫防治的关系等方面的研究进展进行综述,并且论述了以围食膜为害虫生物防治靶标的应用前景.     

聚丙烯酰胺凝胶电泳的快速脱色方法

以牛血清白蛋白为材料进行聚丙烯酰胺凝胶电泳(PAGE)和十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE),凝胶固定后,用考马斯亮蓝R-250染色后比较传统脱色液(冰乙酸-甲醇溶液)和不同盐溶液(NACL、KCL、CUCL2)的脱色效果的结果表明:PAGE和SDS-PAGE胶,0.25和0.5MOL·L-1NACL,在70℃(PAGE)、50℃(SDS-PAGE)下脱色,约2￣4H,效果好,灵敏度高,背景低。