人朊病毒蛋白及其突变体的溶剂可及性和模拟分析

朊病毒是一种全新类型的致病因子,以一种全新的致病机制造成许多神经退化性疾病。它的致病性主要由于PrP^C向PrP^Sc构象转变而造成。为了探讨PrP^C向PrP^Sc转变过程中PrP分子构象变化的机制,我们计算分析了人天然PrP分子及不同的单残基突变体(如M166V,S170N,E200K和R220K)的氨基酸残基溶剂可及性,并对天然PrP分子及其突变体进行了结构重叠模拟RMS偏量分析。结果表明：由于166位等单个残基的突变,造成PrP突变体与天然蛋白的局部结构出现较大差别,使得部分残基的溶剂可及表面积发生了较大变化,并且部分残基改变了它们的位置,同时也影响蛋白质表面的电倚分布,这些改变是为了更好地适应次级相互作用的局部环境。分析表明PrP与一般球蛋白在性质上有一定的差异,说明PrP分子并不是一种稳定的球蛋白结构,只是一种折叠中间物。     

朊粒蛋白PrP~(Sc)寡聚体的形成与跨膜毒性

朊粒蛋白(prionprotein,PrP)传染致病机制一直是朊粒(prion)研究领域的焦点.由正常型朊粒蛋白(PrPC)向致病型朊粒蛋白(PrPSc)的转变是致病的关键步骤.本文综述了近年来PrPC向PrPSc转变的结构变化特征、PrPSc由单体形成寡聚体的组装机制、以及PrPSc寡聚体的跨膜机制与细胞毒性间的关系等方面的研究进展.     

PrPC与蛋白质X结合位点可及性的初步探讨

根据PDB提供的PrPC的原子坐标,利用MSMS程序,对PrPC氨基酸残基溶剂可及表面积进行了计算和分析.结果表明:(1) PrPC氨基酸残基可及性具有总体一致性特点;(2) PrPC蛋白质序列中非保守残基与种间屏障有一定关系;(3) 在PrPC向PrP Sc转变过程中,由于蛋白质X的结合,PrPC可能会出现一定的构象变化,这种变化利于PrPC向PrPSc发生转变.     

羊瘙痒病263K毒株感染的仓鼠脑中PrP分子形式多样性的动态测定

正常细胞的朊蛋白(PrPC)代谢和构象的改变是引发动物和人类可传播性海绵状脑病(transmissiblespongiformencephalopathies,TSEs)的根本原因。将羊瘙痒病(scrapie)仓鼠适应株263K颅内接种仓鼠,在接种后的第20、40、50、60、70、80天,通过Westernblot动态检测仓鼠脑中PrP存在的形式。结果在接种后第40天,在感染动物脑组织中即检测到PrPSc分子,比临床症状出现的时间早(平均潜伏期为66 7±1 1天),且无糖基化形式的PrP分子所占百分比在接种后期增加明显。除了标准分子量大小(30kD～35kD)的PrP分子外,在感染动物脑中存在着高分子量和低分子量形式的PrP分子。定量分析显示,随着接种潜伏期的延长,不同形式PrP分子的含量也在增加,其中低分子量形式的PrP分子与临床症状的出现密切相关。蛋白去糖基化实验表明,在感染动物脑组织中,除了标准分子量大小的PrP蛋白外,还存在一条更小分子量的PrP条带,而正常动物脑组织仅存在标准大小的PrP分子。低分子量形式的PrP分子具有与全长PrP分子相类似的糖基化模式。结果提示,scrapie263K感染的仓鼠脑组织中存在不同分子形式的PrPSc,其PrP分子的代谢可能不同于正常动物。     

微管相关蛋白tau与朊蛋白的相互作用

微管相关蛋白tau参与了许多神经退行性疾病的发生, 其中包括一些人类可传播性海绵状脑病. 为了探讨tau与朊蛋白(PrP)之间可能存在的关系, 首先通过GST pull-down和免疫共沉淀等技术发现重组tau蛋白可通过微管结合区与来源于正常叙利亚仓鼠脑组织中的正常细胞膜朊蛋白(PrP^C)和羊瘙痒因子263K感染仓鼠脑组织的异常朊蛋白(PrPSc)相结合. 利用免疫共沉淀实验发现在正常和羊瘙痒因子感染的仓鼠脑组织中存在tau蛋白与PrPC和PrPSc的相互作用, 并且利用激光共聚焦方法检测到PrP和tau蛋白在CHO细胞内具有共定位的关系. 为了确定PrP与tau蛋白相互作用的部位, 构建了不同区域的PrP片段, 从而证明PrP与tau蛋白相互作用的区域位于PrP的N端序列(23~91 aa). PrP与tau蛋白分子间相互作用的直接实验证据提示tau蛋白可能参与PrP的正常生理功能以及朊病毒病的病理过程.     

朊蛋白PrP~C与神经元的凋亡和存活

朊病毒是不含核酸的一种蛋白质感染颗粒,感染宿主后可诱导细胞固有的同类朊蛋白(PrPC)构象改变、转化成具有蛋白酶抗性的致病性朊粒蛋白(PrPSc),导致可传播性海绵状脑病的发生.PrPC既作为朊病毒复制和疾病发生的分子基础,又是正常的细胞膜蛋白,作为细胞信号转导的参与者调控多条信号通路.因此,揭示PrPC在各条信号途径中发挥的作用将有助于深入了解PrPC的生理功能,进一步理解疾病发生发展过程,为今后的诊断治疗奠定基础.     

与朊毒体相关的鱼类朊蛋白研究概况

疯牛病(mad cow disease),即牛传染性海绵状脑病(bovine transmissible spongiform encephalopathy,BSE)的俗称,是一种慢性消耗性、致死性、中枢神经系统退行性疾病。疯牛病被认为与朊毒体(Prion)有关,朊毒体是由正常朊蛋白(Prion protein,或者PrPC)发生构象改变后形成的异常蛋白(PrPSc)。疯牛病的发生引起了世界各国政府和科学界的高度重视,PrP的起源及其功能研究已成为研究热点。鱼类PrP相关蛋白的研究正在展开中,由于鱼类PrP相关蛋白与朊蛋白的结构相似,鱼类感染TSE类似病存在理论上的风险。本文全面地综述了疯牛病的概况、朊毒体的特性、朊毒体与哺乳动物朊蛋白、鱼类PrP相关蛋白(PrP1、PrP2和PrP3)及鱼类其他PrP相关蛋白的研究情况,为国内水生动物PrP相关蛋白研究提供参考。     

PrP及其缺失突变体与微管蛋白的体外相互作用的初步研究

为了进一步确定PrP蛋白与微管蛋白是否发生分子间相互作用以及PrP蛋白多肽链中与微管蛋白相互作用的区域,我们表达纯化了全长的PrP以及PrP蛋白缺失突变体,提取了兔脑组织中天然微管蛋白。利用pull-down及免疫共沉淀方法检测全长PrP及PrP蛋白缺失突变体与微管蛋白是否发生分子间相互作用。结果显示,全长His-PrP23-231能与微管蛋白发生体外相互作用,并首次证实了PrP与微管蛋白相互作用的区域位于PrP N端第23位至91位氨基酸。此研究为进一步研究PrP在神经细胞的主动转运机制以及Prion疾病的发病机制提供了一定的理论基础。     

单克隆抗体有望用于治疗朊蛋白病

朊蛋白病是一组致命的神经系统变性疾病 ,其特征之一是正常的细胞朊蛋白 (PrPC)转变为有感染性的疾病相关蛋白PrPSc。目前朊蛋白病尚无有效的治疗方法。最近White等的研究表明 :抗朊蛋白单克隆抗体ICSM 18能抑制朊病毒复制 ,延缓朊蛋白病的发展。动物实验表明 ,持续进行这种被动免疫治疗的小鼠健康生长长达 5 0 0天以上 ,且未出现自身免疫反应的副作用 ;而对照组未接受单克隆抗体治疗小鼠的平均存活期仅为 197天左右。该研究表明 ,ICSM 18几乎不与PrPSc结合 ,而与PrPC 有高亲和力 ,其抑制作用呈剂量依赖性。它通过识别PrP的抗原决…     

Arg26和Lys27突变对海南捕鸟蛛毒素-Ⅳ生物活性影响

海南捕鸟蛛毒素Ⅳ(hainantoxin-Ⅳ,HNTX-Ⅳ)是一种新型的从海南捕鸟蛛粗毒中分离纯化的作用于河豚毒素敏感型(tetrodotoxinsensitive,TTXS)钠通道阻断剂.采用2D1HNMR技术解析HNTXⅣ的空间结构为胱氨酸抑制剂结模体,为进一步阐述HNTX-Ⅳ结构与功能的关系,应用固相Fmoc方法化学合成了用丙氨酸代替海南捕鸟蛛毒素Ⅳ第26位精氨酸的单残基突变体R26AHNTXⅣ和第27位赖氨酸的单残基突变体K27AHNTXⅣ.合成的突变体用谷胱甘肽法氧化复性并通过反相高效液相色谱(RPHPLC)纯化.通过MALDITOF质谱测定突变体的分子量.通过核磁共振谱仪测定突变体的空间结构.通过全细胞膜片钳实验比较天然HNTX-Ⅳ(nHNTX-Ⅳ)和两个突变体分子的生物学活性.结果发现,nHNTX-Ⅳ的R26或K27被突变后的空间结构没有发生明显变化.R26AHNTX-Ⅳ能明显抑制TTXS钠电流,K27AHNTX-Ⅳ对TTXS钠电流无明显影响.说明第26位的精氨酸与HNTX-Ⅳ的生物学活性无关,而第27位赖氨酸则是HNTX-Ⅳ的关键残基.     

Serology and systematics of the Ebenales and the Theales

The systematic position ofthe Ebenaceae, Sapotaceae, Styracaceae, Ochnaceae, Stachyuraceae, Dipterocarpaceae, Clusiaceae and Hypericaceae has been investigated using serological comparisons of sets of antigenic determinants. The results show that the Sytracaceae and Sapotaceae are undoubtedly more closely associated with the Actinidiaeceae and Theaceae, respectively, than with each other. We found no corresponding determinants betnween antigen systems from the Ebenaceae and systems from any other family whose relations to this family have been proposed. As discovered previously, investigations of antigen systems from the Ochnaceae, Dipterocarpaceae, Stachyuraceae, Clusiaceae and Hypericaceae are against the idea of a natural order “Theales” in which these families, or at least some of them, are combined with the Theaceae and Actinidiaceae. This paper completes our previous investigations which largely support a superorder Ericanae sensu Ehrendorfer and Takhtajan. We propose to include the Actinidiaceae and Theaceae in this superorder, assigning them a central position laong with the Sapotaceae and Sytracaeae on one side and the Primulales and Ericales on the other. Another most interesting finding is that there are corresponding determinants between antigen systems from the members of the Ericanae and representatives of the Polemoniaceae and Loasaceae.     

The Choroid Plexuses and the Barriers Between the Blood and the Cerebrospinal Fluid

1. The fluid homeostasis of the brain depends both on the endothelial blood–brain barrier and on the epithelial blood–cerebrospinal fluid (CSF) barrier located at the choroid plexuses and the outer arachnoid membrane.2. The brain has two fluid environments: the brain interstitial fluid, which surrounds the neurons and glia, and the CSF, which fills the ventricles and external surfaces of the central nervous system.3. CSF acts as a fluid cushion for the brain and as a drainage route for the waste products of cerebral metabolism.4. Recent findings suggest that CSF may also act as a third circulation conveying substances secreted into the CSF rapidly to many brain regions.     

The efficacy of ropivacaine and bupivacaine in the caesarean section and the effect on the vital signs and the hemodynamics of the lying-in women

ObjectiveTo investigate the efficacy of ropivacaine and bupivacaine in caesarean section and vital signs and the hemodynamics of the lying-in women.MethodsA total of 480 lying-in women who were admitted to this hospital for treatment between December 2017 and June 2018 were enrolled into this study as the subjects, which were divided into the experiment group and the control group, with 240 subjects in each group. In the experiment group, subjects received the local anesthesia by infusion of 1.5 mL ropivacaine (0.75%), while those in the control group also took the local anesthesia by infusion of 1.5 mL bupivacaine (0.75%). Thereafter, we observed the differences in the anesthetic efficiency, vital signs and hemodynamics of the lying-in women between two groups.ResultsThe excellent and good rates of the anesthesia in two groups were 92.1% and 87.9%, showing no obvious difference; in the experiment group, the average arterial pressures and systolic pressures at 5 min and 10 min after combined spinal and epidural analgesia (CSEA) were all elevated when comparing to the control group (all P < 0.05); in the experiment group, the onset time was obviously extended, while duration of sensory and motor block and the duration of motor block were all shorter than those in the control group (all P < 0.05). During anesthesia, the incidence rate of the adverse reactions in the control group was 2.50%, significantly higher than 0.83% in the experiment group (P < 0.05).ConclusionDespite that ropivacaine and bupivacaine are efficient in anesthesia in the CSEA in the caesarean section, ropivacaine is more recommended for little influence on the hemodynamics, shorter duration of sensory block and motor block and low incidence rate of adverse reactions, which are conducive to the recovery and also safe to the patients.     

On the structure and dynamics of the complex of the nucleosome and the linker histone

Several different models of the linker histone (LH)–nucleosome complex have been proposed, but none of them has unambiguously revealed the position and binding sites of the LH on the nucleosome. Using Brownian dynamics-based docking together with normal mode analysis of the nucleosome to account for the flexibility of two flanking 10 bp long linker DNAs (L-DNA), we identified binding modes of the H5-LH globular domain (GH5) to the nucleosome. For a wide range of nucleosomal conformations with the L-DNA ends less than 65 Å apart, one dominant binding mode was identified for GH5 and found to be consistent with fluorescence recovery after photobleaching (FRAP) experiments. GH5 binds asymmetrically with respect to the nucleosomal dyad axis, fitting between the nucleosomal DNA and one of the L-DNAs. For greater distances between L-DNA ends, docking of GH5 to the L-DNA that is more restrained and less open becomes favored. These results suggest a selection mechanism by which GH5 preferentially binds one of the L-DNAs and thereby affects DNA dynamics and accessibility and contributes to formation of a particular chromatin fiber structure. The two binding modes identified would, respectively, favor a tight zigzag chromatin structure or a loose solenoid chromatin fiber.     

Biophysics and Structure of the Patch and the Gigaseal

Interpreting channel behavior in patches requires an understanding of patch structure and dynamics, especially in studies of mechanosensitive channels. High resolution optical studies show that patch formation occurs via blebbing that disrupts normal membrane structure and redistributes in situ components including ion channels. There is a 1-2 μm region of the seal below the patch where proteins are excluded and this may consist of extracted lipids that form the gigaseal. Patch domes often have complex geometries with inhomogeneous stresses due to the membrane-glass adhesion energy (E_a), cytoskeletal forces, and possible lipid subdomains. The resting tension in the patch dome ranges from 1-4 mN/m, a significant fraction of the lytic tension of a bilayer (∼10 mN/m). Thus, all patch experiments are conducted under substantial, and uneven, resting tension that may alter the kinetics of many channels. E_a seems dominated by van der Waals attraction overlaid with a normally repulsive Coulombic force. High ionic strength pipette saline increased E_a and, surprisingly, increased cytoskeletal rigidity in cell-attached patches. Low pH pipette saline also increased E_a and reduced the seal selectivity for cations, presumably by neutralizing the membrane surface charge. The seal is a negatively charged, cation selective, space with a resistance of ∼7 gigohm/μm in 100 mM KCl, and the high resistivity of the space may result from the presence of high viscosity glycoproteins. Patches creep up the pipette over time with voltage independent and voltage dependent components. Voltage-independent creep is expected from the capillary attraction of E_a and the flow of fresh lipids from the cell. Voltage-dependent creep seems to arise from electroosmosis in the seal. Neutralization of negative charges on the seal membrane with low pH decreased the creep rate and reversed the direction of creep at positive pipette potentials.     

Epigenetics and aging: the targets and the marks

'Aging epigenetics' is an emerging field that promises exciting revelations in the near future. Here we focus on the functional and biological significance of the epigenetic alterations that accumulate during aging and are important in tumorigenesis. Paradigmatic examples are provided by the global loss of DNA methylation in aging and cancer and by the promoter hypermethylation of genes with a dual role in tumor suppression and progeria, such as the Werner syndrome (WRN) and lamin A/C genes. Another twist is provided by sirtuins, a family of NAD-dependent deacetylases that act on Lys16 of histone H4, which are emerging as a link between cellular transformation and lifespan.     

Initiating and promoting the aquatic ecosystem health concept: the Society and the Journal

Since the 1970's the management of aquatic habitats has changed from piecemeal monitoring to the ecosystem approach; this was initiated in the North American Great Lakes, comprising social, economic, and environmental aspects. The information included in this paper is based on the presentation made at the Seminar On Ecosystem Approach To Water Management held in Oslo, Norway during 1991. Recently, the multidisciplinary, holistic, and integrated concept of ecosystem health has emerged, and is being advanced for the implementation of an ecosystem approach to environmental management, which has resulted in the formation of an international society (Aquatic Ecosystem Health & Management Society) and the publication of a primary journal (Journal of Aquatic Ecosystem Health). The information has been updated to incorporate new developments and recent progress about the Society and the journal since the Oslo Seminar.     

Phymosomatoid echinoids from the campanian and the maastrichtian of Belgium and the Netherlands

Phymosomatoid echinoids from the Campanian and the Maastrichtian of Belgium and the Netherlands are systematically revised. Ten species, belonging to four genera are identified and described. One new species is introduced:Gauthieria?mosae sp. nov.