Formation of amyloid-like fibrils in COS cells overexpressing part of the Alzheimer amyloid protein precursor

A pathological hallmark of Alzheimer's disease is the deposition of amyloid fibrils in the brain. The principal component of the amyloid fibril is beta/A4 protein, which is derived from a large membrane-bound glycoprotein, Alzheimer amyloid protein precursor (APP). Although the deposition of amyloid is thought to result from the aberrant processing of APP, the detailed molecular mechanisms of amyloidogenesis remain unclear. A C-terminal fragment of APP which spans the beta/A4 and cytoplasmic domains has a tendency to self-aggregate. In an attempt to establish a cultured-cell model for amyloid fibril formation, we have transfected COS-1 cells with complementary DNA encoding the C-terminal 100 residues of APP. In the perinuclear regions of a small population of DNA-transfected cells, we observed inclusion-like deposits which showed a strong immunohistochemical reaction towards an anti-C-terminal APP antibody or an anti-beta/A4 amyloid core-specific antibody. Electron microscope observations of the inclusion-carrying cells revealed an accumulation of amyloid-like fibrils of 8-22 nm diameter near and on the nuclear membrane. The fibrils showed a beaded or helical structure, and reacted positively with the anti-C-terminus antibody by immunoelectron microscopy. These results suggest that the formation of amyloid fibrils is an inherent characteristic of the C-terminal peptide of APP. The present system provides a suitable model for the molecular dissection of the process of brain amyloidogenesis.     

Atomic structures of amyloid cross-beta spines reveal varied steric zippers

Amyloid fibrils formed from different proteins, each associated with a particular disease, contain a common cross-beta spine. The atomic architecture of a spine, from the fibril-forming segment GNNQQNY of the yeast prion protein Sup35, was recently revealed by X-ray microcrystallography. It is a pair of beta-sheets, with the facing side chains of the two sheets interdigitated in a dry 'steric zipper'. Here we report some 30 other segments from fibril-forming proteins that form amyloid-like fibrils, microcrystals, or usually both. These include segments from the Alzheimer's amyloid-beta and tau proteins, the PrP prion protein, insulin, islet amyloid polypeptide (IAPP), lysozyme, myoglobin, alpha-synuclein and beta(2)-microglobulin, suggesting that common structural features are shared by amyloid diseases at the molecular level. Structures of 13 of these microcrystals all reveal steric zippers, but with variations that expand the range of atomic architectures for amyloid-like fibrils and offer an atomic-level hypothesis for the basis of prion strains.     

A camelid antibody fragment inhibits the formation of amyloid fibrils by human lysozyme

Amyloid diseases are characterized by an aberrant assembly of a specific protein or protein fragment into fibrils and plaques that are deposited in various organs and tissues, often with serious pathological consequences. Non-neuropathic systemic amyloidosis is associated with single point mutations in the gene coding for human lysozyme. Here we report that a single-domain fragment of a camelid antibody raised against wild-type human lysozyme inhibits the in vitro aggregation of its amyloidogenic variant, D67H. Our structural studies reveal that the epitope includes neither the site of mutation nor most residues in the region of the protein structure that is destabilized by the mutation. Instead, the binding of the antibody fragment achieves its effect by restoring the structural cooperativity characteristic of the wild-type protein. This appears to occur at least in part through the transmission of long-range conformational effects to the interface between the two structural domains of the protein. Thus, reducing the ability of an amyloidogenic protein to form partly unfolded species can be an effective method of preventing its aggregation, suggesting approaches to the rational design of therapeutic agents directed against protein deposition diseases.     

Amyloid-like fibrils of ribonuclease A with three-dimensional domain-swapped and native-like structure

Amyloid or amyloid-like fibrils are elongated, insoluble protein aggregates, formed in vivo in association with neurodegenerative diseases or in vitro from soluble native proteins, respectively. The underlying structure of the fibrillar or 'cross-beta' state has presented long-standing, fundamental puzzles of protein structure. These include whether fibril-forming proteins have two structurally distinct stable states, native and fibrillar, and whether all or only part of the native protein refolds as it converts to the fibrillar state. Here we show that a designed amyloid-like fibril of the well-characterized enzyme RNase A contains native-like molecules capable of enzymatic activity. In addition, these functional molecular units are formed from a core RNase A domain and a swapped complementary domain. These findings are consistent with the zipper-spine model in which a cross-beta spine is decorated with three-dimensional domain-swapped functional units, retaining native-like structure.     

Long-term dendritic spine stability in the adult cortex

The structural dynamics of synapses probably has a crucial role in the development and plasticity of the nervous system. In the mammalian brain, the vast majority of excitatory axo-dendritic synapses occur on dendritic specializations called 'spines'. However, little is known about their long-term changes in the intact developing or adult animal. To address this question we developed a transcranial two-photon imaging technique to follow identified spines of layer-5 pyramidal neurons in the primary visual cortex of living transgenic mice expressing yellow fluorescent protein. Here we show that filopodia-like dendritic protrusions, extending and retracting over hours, are abundant in young animals but virtually absent from the adult. In young mice, within the 'critical period' for visual cortex development, approximately 73% of spines remain stable over a one-month interval; most changes are associated with spine elimination. In contrast, in adult mice, the overwhelming majority of spines (approximately 96%) remain stable over the same interval with a half-life greater than 13 months. These results indicate that spines, initially plastic during development, become remarkably stable in the adult, providing a potential structural basis for long-term information storage.     

Experience-dependent and cell-type-specific spine growth in the neocortex

Functional circuits in the adult neocortex adjust to novel sensory experience, but the underlying synaptic mechanisms remain unknown. Growth and retraction of dendritic spines with synapse formation and elimination could change brain circuits. In the apical tufts of layer 5B (L5B) pyramidal neurons in the mouse barrel cortex, a subset of dendritic spines appear and disappear over days, whereas most spines are persistent for months. Under baseline conditions, new spines are mostly transient and rarely survive for more than a week. Transient spines tend to be small, whereas persistent spines are usually large. Because most excitatory synapses in the cortex occur on spines, and because synapse size and the number of alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors are proportional to spine volume, the excitation of pyramidal neurons is probably driven through synapses on persistent spines. Here we test whether the generation and loss of persistent spines are enhanced by novel sensory experience. We repeatedly imaged dendritic spines for one month after trimming alternate whiskers, a paradigm that induces adaptive functional changes in neocortical circuits. Whisker trimming stabilized new spines and destabilized previously persistent spines. New-persistent spines always formed synapses. They were preferentially added on L5B neurons with complex apical tufts rather than simple tufts. Our data indicate that novel sensory experience drives the stabilization of new spines on subclasses of cortical neurons. These synaptic changes probably underlie experience-dependent remodelling of specific neocortical circuits.     

内固定器对人体颈椎运动生物力学性能的影响

在失稳颈椎内安装固定器后,其稳定性的生物力学评价是医学研究的一个重要课题。该文提出了一种双目三维光学精密运动测量方法对其进行评价。采用该方法分别对保存良好的人体颈椎标本在失稳、内固定以及疲劳3种状态下的三维运动情况(包括前屈、后伸、左侧弯、右侧弯4种运动)进行了测量,并以每种运动情况下的颈椎C 1-C 2节相互旋转角度衡量其内固定的效果。实验结果表明,内固定后颈椎的旋转角度明显小于失稳时的旋转角度;内固定后模拟远期效果的疲劳加载后测试结果证实内固定的效果仍然是可靠的。     

颈椎融合器的力学测试及有限元验证

目的:设计一款聚乳酸材质融合器,完成力学测试并判定融合器的可行性.方法:分别对健康的和植入融合器的C5-C6节段羊颈椎进行压缩、前屈、后伸、侧弯实验,得出载荷-位移曲线,比较融合器植入前后颈椎的力学性能;再用万能电子试验机检验融合器的疲劳强度;最后对融合器进行有限元仿真观察应力分布.结果:加入融合器后颈椎的稳定性有所提高;融合器的疲劳强度合格,应力分布合理,未产生应力集中.结论:聚乳酸材质融合器有足够的力学强度,满足力学要求.     

钢筋混凝土双重网格楼盖结构构造特点的研究

钢筋混凝土双重网格结构在钢筋混凝土空腹夹层板基础上发展而来,但它的网格组成形式、结构构造发生了改变,对其构造特点进行研究分析,说明该结构结构型式合理,建筑造型新颖美观,适用于跨度和空间较大的公共建筑.     

论农民专业合作社法人治理结构的冲突与完善

我国农民专业合作社特殊的产权结构使其在治理结构上存在着社员民主控制与管理层专业化经营的冲突、平权式治理结构与少数人控制的冲突、劳动控制资本与成员利益诉求异质性的冲突。这些冲突实质上是农民专业合作社发展过程中守成与创新的冲突。如何权衡合作社发展过程的守成与创新是解决农民专业合作社治理结构冲突的突破口。     

椎间盘与韧带对腰椎变形影响的生物力学研究

利用CT扫描图像和三维重建软件,重建羊腰椎L1-L4的三维实体模型,建立包括椎体、椎间盘和韧带的有限元模型,数值计算得到椎体,椎间盘和韧带的应力分布规律.数值计算结果表明:施加单一牵引载荷和侧向弯曲载荷时,考虑间盘与韧带时腰椎的位移场与未考虑间盘与韧带时腰椎的位移场有明显的区别,对椎体的等效应力影响较小.文中结果表明,椎间盘与韧带明显影响腰椎变形的,研究腰椎变形需要考虑椎间盘与韧带.     

恒星结构的稳定性研究

恒星是一个热核反应体系,已有很多学者对其结构稳定性做过研究.作为粒子数可变的体系,宏观性质可以用下述动力学方程组描述:(1) 分析系统的定态解是否稳定以及在什么条件下由稳定转变为不稳定,一个有效而简便的方法是小扰动简正模方法.     

Structure of the acrosomal bundle

In the unactivated Limulus sperm, a 60- micro m-long bundle of actin filaments crosslinked by the protein scruin is bent and twisted into a coil around the base of the nucleus. At fertilization, the bundle uncoils and fully extends in five seconds to support a finger of membrane known as the acrosomal process. This biological spring is powered by stored elastic energy and does not require the action of motor proteins or actin polymerization. In a 9.5-A electron cryomicroscopic structure of the extended bundle, we show that twist, tilt and rotation of actin-scruin subunits deviate widely from a 'standard' F-actin filament. This variability in structural organization allows filaments to pack into a highly ordered and rigid bundle in the extended state and suggests a mechanism for storing and releasing energy between coiled and extended states without disassembly.     

资本结构理论与乡镇企业治理结构变迁

以资本结构理论对乡镇企业治理结构的变迁进行了解释,提出乡镇企业资本结构是其治理结构演进的决定因素,优化乡镇企业资本结构是改进治理结构提高企业效率和效益的关键.     

含均三嗪结构苯并噻唑类化合物的合成和晶体结构

以三聚氯氰等为原料合成了一种含均三嗪结构苯并噻唑类化合物（HY）,并通过X射线单晶衍射对其结构进行了表征。结果表明：HY属于单斜晶体,空间群为P2（1）／”。晶胞参数为：Z=4,a=1．4278（17）nm,b=1．6690（2）nm,c=1．44842（17）nm,α=90°,β=102．707（2）°,）,γ=90°,V=3．3655（7）nm^3;R1=0．0755;wR2=0．2161。     

σ型半群的结构

研究一类拟适当半群，即σ型半群。引进了适当半群和左正则带的拟织积，建立σ型半群的拟织积结构，作为应用，给出了σ型调和半群和右逆半群的结构。     

Structure of the actin-myosin interface

The topography of the rigor complex between F-actin and myosin heads (S1) has been investigated by carbodiimide zero-length cross-linking. The results demonstrate for the first time that the 95,000-molecular weight (95K) heavy chain of the myosin head enters into van der Waals contact with two neighbouring actin monomers; one is bound to the 50K domain and the other to the 20K domain of the myosin chain. The covalent F-actin-S1 complex can be isolated; it shows a vastly elevated Mg2+-ATPase. Each pair of actin subunits in the thin filament seems to act as a functional unit for specific binding of a myosin head and stimulation of its Mg2+-ATPase activity.