生物体内的纳米机器——分子马达

分子马达是生物体内具有马达功能的一类蛋白质大分子纳米机器,可以执行完成生命体内的一切活动,包括肌肉的收缩、细胞内部物质的运输、遗传物质(DNA)的复制、细胞的分裂等等。按照分子马达的不同种类,介绍了各类线性分子马达(如驱动蛋白、动力蛋白和肌球蛋白)的结构、运动方式、主要功能等生物特征,并介绍了旋转分子马达(如ATP合酶)的生物特征,最后进行总结,展望未来。     

纳米机器——分子马达

介绍了细胞内分子马达的能量转化途径,几种纳米分子马达如驱动蛋白、动力蛋白、肌球蛋白和旋转马达的结构和功能,并展望了分子马达对人类的贡献。     

驱动蛋白结构与运动机制

驱动蛋白是一类能够利用ATP水解释放的化学能驱动其所携带的“货物”分子沿着微管（microtubule,MT）定向运动的分子马达,在细胞器运输、有丝分裂、轴突运输等方面有着重要的生理作用。随着驱动蛋白结合ADP、ATP和未结合核苷酸（APO）三种特征状态的晶体结构的解析,驱动蛋白构象变化的研究得到了进一步发展,而在力产生机制和运动模型方面仍然存在较大争议。本文以kinesin-1家族为例,分析了驱动蛋白三种特征状态结构的特点、状态结构间的构象转变,论述了驱动蛋白的力产生机制和整个迈步过程。并探讨了驱动蛋白的运动模型,同时采用分子动力学模拟比较了驱动蛋白的两种迈步方式,为深入研究驱动蛋白提供了一定的理论计算。最后,基于本课题组对复杂体系的研究,对驱动蛋白体系的控制机制提出了新的假设,并对未来的研究方向进行了展望。     

生物分子开关研究进展

分子开关是建立在分子水平上的一个可逆过程,外界条件的改变能使分子的结构或构型会有一些改变,从而表现出一些特殊的性质.其在生物体内的信号传导和信息调控等方面发挥着重要作用,成为近年来国内外的一大研究热点.本文介绍了胚胎干细胞分化的重要分子开关:Oct-3/4、神经突起相关分子开关Rho族GTP酶、控制肌卫星细胞激活的分子开关转录因子RBP-J、慢性疼痛的分子开关蛋白激酶G(PKG)、脂肪控制分子开关Wnt蛋白、调节核糖体翻译功能的分子开关L11蛋白、控制饥饿的分子开关巨噬细胞因子MIC-1、调节人体生理节奏的分子开关BMAL1基因、癌症相关的分子开关Wnt蛋白家族以及糖尿病相关的活性开关蛋白TORC2等生物学领域调控功能分子开关的作用机理以及相关研究进展情况.     

驱动蛋白的能量转换过程——从ATP分子结合到颈链对接

驱动蛋白是一种分子马达,同时也是一种核苷酸酶,它能够将ATP分子所携带的化学能转化为其沿微管蛋白行走的机械能,每消耗一个ATP分子行走一步。对于驱动蛋白如何将ATP的化学能转化为构象变化的机械能的研究一直是生物物理学研究的热点问题。本文从3个方面对此问题的研究进展进行了综述:ATP分子与驱动蛋白结合; ATP结合引起驱动蛋白头部产生转动;驱动蛋白头部转动引起驱动蛋白颈链向头部的对接。将这三个方面的内容合并起来就构成了驱动蛋白的能量传递路径。     

驱动蛋白的结构与功能

驱动蛋白是一类蛋白质超家族的总称,其中驱动蛋白-1(以下简称驱动蛋白)是目前已知的有机体内最小的马达蛋白.驱动蛋白能够催化三磷酸腺苷(adenosine triphosphate,ATP)分子的水解反应,将贮藏在ATP中的化学能转变为自身机械运动所需的机械能.驱动蛋白能够沿着微管连续定向运动,在细胞的有丝分裂和胞内物质运输中发挥重要作用.在真核细胞中,驱动蛋白主要以二聚体的形式存在,其结构主要包括4个部分,即马达头部、茎部、连接头部与茎部的颈链以及与"货物"相结合的尾部.驱动蛋白二聚体独特的结构特征以及各个组成部分协调的构象变化,保证了其沿微管的连续行走.目前,驱动蛋白的结构与功能之间的关系的研究取得了重要的进展.随着实验和计算水平的不断提高,彻底了解驱动蛋白的运动机理已经为期不远了.     

单分子纳米生物技术

生物分子需要相互协调组成各种分子机器,例如：分子马达、细胞信号处理器、DNA转录机、蛋白质合成器等,来实现他们负责的各种生理功能。一方面,这些分子机器的功能对于环境改变有着很强的适应性,这种适应性对于生物体和生物系统的稳定来说是必须的。另一方面,由于分子机器的大小只有几纳米并且结构具有可变性,它们的运转倾向于受到热扰动的影响具有随机性,此外,分子机器所利用的输入能量的水平与平均热运动能量kBT近似,分子机器可以高效地将热噪声的能量转化利用于行使其功能,对于这些能量的利用效率也很高,这与人造机器运转所需能量远远高于热运动能量相比有着十分显著的差别。因此,分子机器的潜在机理比人造机器复杂了许多。近些年来,随着单分子探测技术和纳米技术在生命科学的各个领域的广泛应用,使得生物分子的动力学性质,以及以往被隐藏在系综平均结果后的单个分子机器运转规律逐渐被揭示。我们研究的目标是希望通过揭示分子机器的独特运转规律进而能够了解、掌握具有适应性的生物系统中包含的工程学原理。本文综述了我们所作的一些关于分子马达、酶促反应、蛋白质动力学和细胞信号转导的单分子探测试验,并且讨论了热涨落（噪声）是怎样在具有独特的运转规律的生物分子机器中起到正效应,进而允许生物系统具有可变性和适应性的。     

单分子层次上相互作用自动检测

任何生命过程都与分子间相互作用有关。这些相互作用决定了生物分子间的＂交流＂方式,组成了生物过程的基本语言。Müller教授研究组发展了一种全自动＂机器人＂（一种全自动原子力显微镜）,可以通过检测细胞上的＂分子机器＂分析分子间相互作用。为了实现这样的目标,该仪器需要将不同的空间尺度联系起来：宏观尺度的悬臂利用其微观尺度的针尖与纳米尺度的蛋白质相接触,进而在亚纳米的尺度上定位与检测分子间相互作用。这项技术能够帮助人们以亚纳米尺度的分辨率定位细胞内分子机器的相互作用位置,并且观察分子间相互作用如何驱动这些分子机器行使功能。在药物筛选研究领域,该技术可以被用来检测配体以及抑制剂与蛋白质结合的位点和强度,还可以检测受体的不同功能状态。     

逃逸速率对分子马达定向运动的影响

在考虑逃逸速率的情况下采用主方程方法研究分子马达,计算了逃逸速率对分子马达的漂移速率、扩散系数、停留时间以及分子马达在轨道上移动的距离．通过计算发现,逃逸速率影响分子马达的定向运动．     

细胞运动、细胞迁移与细胞骨架研究进展

细胞定向运动与细胞骨架的动态循环密切相关。运动细胞在其伪足前沿依靠细胞骨架的不断聚合推动细胞膜的前进,在基部靠近细胞体部位通过细胞骨架的不断解聚收缩拖拉细胞体向前运动,细胞骨架的聚合与解聚通过伪足与支撑表面的吸附与解吸附而偶连。肌动蛋白组成的微丝骨架是大多数运动细胞的主要成分。外界刺激引起微丝细胞骨架动态变化的信号通路已逐步明了。线虫精子细胞的运动行为与阿米巴变形运动相似,但是在线虫精子细胞中没有肌动蛋白,而是以精子主要蛋白为基础形成细胞骨架驱动精子细胞的运动。与肌动蛋白不同,精子主要蛋白没有分子极性、ATP结合位点和马达蛋白。通过比较研究以上两种运动体系将有助于在分子水平上进一步阐明细胞运动的机理。     

mTOR complexes -- molecular spiders in molecular networks

PI3K route is one of the most outstanding signal transduction pathways, which has a key role in the decision-making processes and functions of a cell. In this network mTOR (mammalian target of rapamycin) is a well-known integrator. mTOR forms two complexes, and their increased activity is present in many human tumors. Therefore, mTOR inhibitors became more and more important in the targeted therapy, first of all in the treatment of renal cancer, neuroendocrine pancreatic cancer and certain astrocytomas, and many trials are going on in other tumor types. The therapeutic results are obvious, but problems also occur, which lead to new strategies and to the development of new drugs in order to create more individualised cancer therapy.     

Protein scaffold-based molecular probes for cancer molecular imaging

Protein scaffold molecules are powerful reagents for targeting various cell signal receptors, enzymes, cytokines and other cancer-related molecules. They belong to the peptide and small protein platform with distinct properties. For the purpose of development of new generation molecular probes, various protein scaffold molecules have been labeled with imaging moieties and evaluated both in vitro and in vivo. Among the evaluated probes Affibody molecules and analogs, cystine knot peptides, and nanobodies have shown especially good characteristics as protein scaffold platforms for development of in vivo molecular probes. Quantitative data obtained from positron emission tomography, single photon emission computed tomography/CT, and optical imaging together with biodistribution studies have shown high tumor uptakes and high tumor-to-blood ratios for these probes. High tumor contrast imaging has been obtained within 1 h after injection. The success of those molecular probes demonstrates the adequacy of protein scaffold strategy as a general approach in molecular probe development.     

Toward molecular electronics. Self-screening of molecular wires

One-dimensional polyelectrolytes are proposed as molecular devices based on soliton propagation. The use of such "molecular wires" as delay lines or shift registers raises the problem of their electrical screening, because, when a large number of these wires are densely packed, cross-talking between them might occur. In order to overcome such a drawback, a detailed study of the polyelectrolyte behavior in an ionic solution has been developed, under equilibrium conditions. The results bring into evidence a condensation process, around each molecular wire, of the mobile ions in the solution. The consequent self-screening effect could be relevant in reducing cross-talking.     

Contributions of plant molecular systematics to studies of molecular evolution

Dobzhansky stated that nothing in biology makes sense except in the light of evolution. A close corollary, and the central theme of this paper, is that everything makes a lot more sense in the light of phylogeny. Systematics is in the midst of a renaissance, heralded by the widespread application of new analytical approaches and the introduction of molecular techniques. Molecular phylogenetic analyses are now commonplace, and they have provided unparalleled insights into relationships at all levels of plant phylogeny. At deep levels, molecular studies have revealed that charophyte green algae are the closest relatives of the land plants and suggested that liverworts are sister to all other extant land plants. Other studies have suggested that lycopods are sister to all other vascular plants and clarified relationships among the ferns. The impact of molecular phylogenetics on the angiosperms has been particularly dramatic – some of the largest phylogenetic analyses yet conducted have involved the angiosperms. Inferences from three genes (rbcL, atpB, 18S rDNA) agree in the major features of angiosperm phylogeny and have resulted in a reclassification of the angiosperms. This ordinal-level reclassification is perhaps the most dramatic and important change in higher-level angiosperm taxonomy in the past 200 years. At lower taxonomic levels, phylogenetic analyses have revealed the closest relatives of many crops and model organisms for studies of molecular genetics, concomitantly pointing to possible relatives for use in comparative studies and plant breeding. Furthermore, phylogenetic information has contributed to new perspectives on the evolution of polyploid genomes. The phylogenetic trees now available at all levels of the taxonomic hierarchy for angiosperms and other green plants should play a pivotal role in comparative studies in diverse fields from ecology to molecular evolution and comparative genetics.     

Seven-helix bundles: molecular modeling via restrained molecular dynamics.

Simulated annealing via restrained molecular dynamics (SA/MD) has been used to model compact bundles of seven approximately (anti)parallel alpha-helices. Seven such helix bundles occur, e.g., in bacteriorhodopsin, in rhodopsin, and in the channel-forming N-terminal domain of Bacillus thuringiensis delta-endotoxin. Two classes of model are considered: (a) those consisting of seven Ala20 peptide chains; and (b) those containing a single polypeptide chain, made up of seven Ala20 helices linked by GlyN interhelix loops (where N = 5 or 10). Three different starting C alpha templates for SA/MD are used, in which the seven helices are arranged (a) on a left-handed circular template, (b) on a bacteriorhodopsin-like template, or (c) on a zig-zag template. The ensembles of models generated by SA/MD are analyzed in terms of their geometry and energetics, and the most stable structures from each ensemble are examined in greater detail. Structures resembling bacteriorhodopsin and structures resembling delta-endotoxin are both represented among the most stable structures. delta-Endotoxin-like structures arise from both circular and bacteriorhodopsin-like C alpha templates. A third helix-packing mode occurs several times among the stable structures, regardless of the C alpha template and of the presence or absence of interhelix loops. It is characterized by a "4 + 1" core, in which four helices form a distorted left-handed supercoil around a central, buried helix. The remaining two helices pack onto the outside of the core. This packing mode is comparable with that proposed for rhodopsin on the basis of two-dimensional electron crystallographic and sequence analysis studies.     

Non-linear molecular pattern classification using molecular beacons with multiple targets

In vitro pattern classification has been highlighted as an important future application of DNA computing. Previous work has demonstrated the feasibility of linear classifiers using DNA-based molecular computing. However, complex tasks require non-linear classification capability. Here we design a molecular beacon that can interact with multiple targets and experimentally shows that its fluorescent signals form a complex radial-basis function, enabling it to be used as a building block for non-linear molecular classification in vitro. The proposed method was successfully applied to solving artificial and real-world classification problems: XOR and microRNA expression patterns.     

Computer assisted simulations and molecular graphics methods in molecular design

A survey is presented of computer-assisted statistical mechanical methods. The general theoretical background is described and special methods are discussed in detail. Practical procedures allowing for the calculation of binding energies are examined. A recent perturbation-relaxation procedure is summarized.     

Wavelength-shifting molecular beacons

We describe wavelength-shifting molecular beacons, which are nucleic acid hybridization probes that fluoresce in a variety of different colors, yet are excited by a common monochromatic light source. The twin functions of absorption of energy from the excitation light and emission of that energy in the form of fluorescent light are assigned to two separate fluorophores in the same probe. These probes contain a harvester fluorophore that absorbs strongly in the wavelength range of the monochromatic light source, an emitter fluorophore of the desired emission color, and a nonfluorescent quencher. In the absence of complementary nucleic acid targets, the probes are dark, whereas in the presence of targets, they fluoresce-not in the emission range of the harvester fluorophore that absorbs the light, but rather in the emission range of the emitter fluorophore. This shift in emission spectrum is due to the transfer of the absorbed energy from the harvester fluorophore to the emitter fluorophore by fluorescence resonance energy transfer, and it only takes place in probes that are bound to targets. Wavelength-shifting molecular beacons are substantially brighter than conventional molecular beacons that contain a fluorophore that cannot efficiently absorb energy from the available monochromatic light source. We describe the spectral characteristics of wavelength-shifting molecular beacons, and we demonstrate how their use improves and simplifies multiplex genetic analyses.