Structural polymorphism of DNA-dendrimer complexes

DNA condensation in vivo relies on electrostatic complexation with small cations or large histones. We report a synchrotron x-ray study of the phase behavior of DNA complexed with synthetic cationic dendrimers of intermediate size and charge. We encounter unexpected structural transitions between columnar mesophases with in-plane square and hexagonal symmetries, as well as liquidlike disorder. The isoelectric point is a locus of structural instability. A simple model is proposed based on competing long-range electrostatic interactions and short-range entropic adhesion by counterion release.     

抗生物素蛋白与DNA相互作用的单分子研究

抗生物素蛋白(avidin)在生物单分子实验中被广泛用于DNA与修饰表面的连接,同时avidin也可作为一种DNA载体用于基因治疗中.本文利用原子力显微镜(AFM)、动态光散射(DLS)、单分子磁镊(MT)技术系统地研究了avidin与DNA之间的相互作用,以及avidin引起DNA凝聚的机理.首先通过AFM对avidin-DNA复合体形貌进行观察,发现不但有avidin导致DNA凝聚的环状形貌,同时也存在avidin自身聚集引起的DNA凝聚现象,通过定量分析,发现其凝聚尺寸越来越小,而当avidin浓度大于2 ng·μL~(-1)时,其凝聚尺寸又突然变大.DLS实验结果也显示了同样的规律,伴随着avidin浓度的升高,DNA的粒径大小从大约170 nm减小到125 nm左右,其电泳迁移率由-2.76(10~(-4)cm~2·V~(-1)·s~(-1))变化到-0.1(10~(-4)cm~2·V~(-1)·~(-1)).此外,通过MT技术的力谱曲线变化,发现avidin导致的DNA凝聚与其他多价离子相比,长度的变化曲线几乎呈线性变化,偶尔存在少而小的阶跃,这种变化趋势与组蛋白的变化曲线更相似.因此可以判断,avidin导致DNA凝聚是由avidin与DNA的静电吸引和avidin自身聚集两种相互作用引起的.     

Brownian dynamics simulation of the cross-talking effect among modified histones on conformations of nucleosomes

Using Brownian dynamics simulation,we studied the effect of histone modifications on conformations of an array of nucleosomes in a segment of chromatin.The simulation demonstrated that the segment of chromatin shows the dynamic behaviour that its conformation can switch between a state with nearly all of the histones being wrapped by DNA and a state with nearly all of the histones being unwrapped by DNA,thus involving the "cross-talking" interactions among the histones.Each state can stay for a sufficiently long time.These conformational states are essential for gene expression or gene silence.The simulation also shows that these conformational states can be inherited by the daughter DNAs during DNA replication,giving a theoretical explanation of the epigenetic phenomenon.     

Role of tension and twist in single-molecule DNA condensation

Using magnetic tweezers, we study in real time the condensation of single DNA molecules under tension. We find that DNA condensation occurs via discrete nucleated events. By measuring the influence of an imposed twist, we show that condensation is initiated by the formation of a plectonemic supercoil. This demonstrates a strong interplay between the condensation transition and externally imposed mechanical constraints.     

Effects of oxaliplatin on DNA condensation

In this paper the interactions between DNA and anti-cancer drug oxaliplatin were investigated by using magnetic tweezers. The dynamics of DNA condensation due to oxaliplatin was traced under various forces. It is found that torsion constraint in DNA enhances the ability of oxaliplatin for shortening DNA. The transplatin helps oxaliplatin combine to DNA and increase the rate of DNA condensation. All these results are consistent to the previously proposed model and are helpful for further investigation of interaction between DNA and oxaliplatin.     

基于变温紫外分光光度法研究温度对DNA凝聚过程影响

荧光单分子实验结果表明温度能够促进DNA凝聚,随着温度升高,精胺(亚精胺)-DNA凝聚体系中DNA凝聚构象趋于更加紧致、有序。为探究温度对不同高价离子-DNA凝聚体系的影响,利用自行搭建的变温紫外分光光度计,通过系统研究荧光单分子实验中采用的精胺(亚精胺)-DNA凝聚体系及去离子水中DNA在260 nm处特征吸收值随温度变化情况,确定了DNA凝聚构象与其260 nm处特征吸收值之间的对应关系为：DNA凝聚构象越紧致、有序,其对应的260 nm处紫外吸收值越低。在此基础上,系统地研究了DNA凝聚实验中常用四种钴胺化合物-DNA凝聚体系随温度变化情况,结果表明三氯六胺合钴与精胺(亚精胺)类似,其对应的DNA凝聚体系在260 nm特征吸收值,随着温度升高而逐渐降低,即DNA凝聚构象趋于更加紧致、有序。而三(乙二胺)氯化钴、反式双(乙二胺)氯化钴、五胺氯化钴的情况却不同,与其对应的DNA凝聚体系在260 nm特征吸收值,随着温度升高,呈现先增加,再降低,然后再增加的规律。     

应用分子梳技术对DNA与组蛋白相互作用的研究

利用分子梳技术对λ DNA和组蛋白的相互作用进行了研究. 通过这种简单有效的方法，我们将λ DNA分子拉伸到26—28 μm，相当于其原长（约162 μm）的16—17倍. 当组蛋白与DNA结合后，DNA分子发生凝聚现象，复合体的拉伸长度明显变短，其峰值分布在10—14 μm之间. DNA 组蛋白复合体的拉伸长度与组蛋白的浓度、与碱基对和荧光染料的比例有显著的关系. 关键词： 分子梳 组蛋白 DNA 荧光显微     

Effects of Ionic Dependence of DNA Persistence Length on the DNA Condensation at Room Temperature

DNA persistence length is a key parameter for quantitative interpretation of the conformational properties of DNA and related to the bending rigidity of DNA.A series of experiments pointed out that,in the DNA condensation process by multivalent cations,the condensed DNA takes elongated coil or compact globule states and the population of the compact globule states increases with an increase in ionic concentration.At the same time,single molecule experiments carried out in solution with multivalent cations(such as spermidine,spermine)indicated that DNA persistence length strongly depends on the ionic concentration.In order to revolve the effects of ionic concentration dependence of persistence length on DNA condensation,a model including the ionic concentration dependence of persistence length and strong correlation of multivalent cation on DNA is provided.The autocorrelation function of the tangent vectors is found as an effective way to detect the ionic concentration dependence of toroidal conformations.With an increase in ion concentration,the first periodic oscillation contained in the autocorrelation function shifts,the number of segment contained in the first periodic oscillation decreases gradually.According to the experiments,the average long-axis length is defined to estimate the ionic concentration dependence of condensation process further.The relation between long-axis length and ionic concentration matches the experimental results qualitatively.     

Effect of Torsion on Cisplatin-Induced DNA Condensation

We investigate the effect of torsion on DNA condensation with the covalently closed circular DNA as the torsionconstrained system, using an atomic force microscope. It is found that there are two stages in the DNA condensation process under torsional constraint. At the early stage, the low torsion will accelerate DNA condensation by promoting the formation of micro-loops or intersection structures; while at the later stage, the increasing torsion will slow it down by preventing the crosslinking of cisplatin and DNA since the DNA molecule becomes more rigid. Our results show the important role of torsion in DNA condensation and sheds new light on the mechanism for DNA condensation.     

Processes of DNA condensation induced by multivalent cations： Approximate annealing experiments and molecular dynamics simulations

The condensation of DNA induced by spermine is studied by atomic force microscopy （AFM） and molecular dynamics （MD） simulation in this paper. In our experiments, an equivalent amount of multivalent cations is added to the DNA solutions in different numbers of steps, and we find that the process of DNA condensation strongly depends on the speed of adding cations. That is, the slower the spermine cations are added, the slower the DNA aggregates. The MD and steered molecular dynamics （SMD） simulation results agree well with the experimental results, and the simulation data also show that the more steps of adding multivalent cations there are, the more compact the condensed DNA structure will be. This investigation can help us to control DNA condensation and understand the complicated structures of DNA--cation complexes.     

Single Molecule Fluorescence Imaging and Its Application to the Study of DNA Condensation

Single molecule fluorescence imaging incorporated with optical tweezers and a laminar flow cell has been used to monitor the kinetic process of DNA condensation induced by spermidine. It was found that at least two steps were involved in the condensation process of the hydrodynamically-stretched linear DNA; a lag period followed by a rapid collapse of DNA. The lag time increased with the flow speed and the collapse time remained short within the range of the flow speed studied. The effect of salt concentration on the condensation process was examined, and the results suggest that the longer lag time observed in the higher salt buffer probably results from the displacement of bound cations and rearrangement of spermidine on the DNA. The flow-speed dependence of the lag time suggests that a nucleation event at the free end of the DNA, i.e. formation of a loop, may play a vital role in the kinetic process of condensation.     

Solubility and charge inversion of complexes of DNA and basic proteins

The basic proteins, protamines and histones H1, are known to condense DNA in vivo. We examine here their ability to condense and solubilize in vitro linear DNA [and a synthetic polyanion, Poly(Styrene-Sulfonate) or PSS] at low ionic concentrations by varying the charge concentration ratio. Phase separation is observed in a very narrow range of ratios for short DNA and PSS; on both sides of this range, polydisperse and charged complexes are formed. A charge inversion is detected. For long DNA chains however, a different behavior is observed: the complexes are not soluble in excess of proteins.     

利用纳米孔研究DNA

脱氧核糖核酸 (DNA)和蛋白质是构成生命体最为重要的两类生物大分子 .随着科学技术的快速发展 ,越来越多的纳米技术被用来研究这些生物大分子 .文章详细介绍了近来利用纳米孔技术研究DNA的一些进展 .结合作者近期利用聚焦离子束 (FIB)制作纳米孔的工作 ,提出了利用纳米孔解离核小体的设想 .如果能够利用纳米孔将双螺旋DNA从组蛋白八聚体上剥离下来 ,并探测这一过程 ,将揭示核小体中包含的许多生物化学、物理信息 .文章对此进行了较为详细的分析 :处于电场中的核小体在电场的作用下 ,DNA分子穿越纳米孔 ,同时由于纳米孔的阻挡力 ,使组蛋白不能穿越 ,从而诱使DNA从组蛋白八聚体上分离下来 .通过准确检测DNA分子穿孔过程中引起的电流阻塞效应 ,可将DNA与组蛋白的相互作用的一些性质反映出来     

Polymer reptation and nucleosome repositioning

We consider how beads can diffuse along a chain that wraps them, without becoming displaced from the chain; our proposed mechanism is analogous to the reptation of "stored length" in more familiar situations of polymer dynamics. The problem arises in the case of globular aggregates of proteins (histones) that are wound by DNA in the chromosomes of plants and animals; these beads (nucleosomes) are multiply wrapped and yet are able to reposition themselves over long distances, while remaining bound by the DNA chain.     

Reentrant condensation of proteins in solution induced by multivalent counterions

Negatively charged globular proteins in solution undergo a condensation upon adding trivalent counterions between two critical concentrations C and C, C 相似文献   

Low-force DNA condensation and discontinuous high-force decondensation reveal a loop-stabilizing function of the protein Fis

We report single-DNA-stretching experiments showing that the protein Fis, an abundant bacterial chromosome protein of E. coli, mediates a dramatic DNA condensation to zero length. This condensation occurs abruptly when DNA tension is reduced below a protein-concentration-dependent threshold f* < 1 pN. Following condensation, reopening under larger forces proceeds via a series of discrete jumps, indicating that Fis is able to stabilize DNA crossings. Our experiments suggest that Fis may play a role in vivo stabilizing the "loop-domain" structure of the bacterial chromosome.     

Theory of DNA condensation in the presence of stretching force: Zimm-Bragg model

A theory of DNA condensation by multivalent cations in the presence of an external stretching force is presented. It is shown that in the mean-field approximation the system is described by the Zimm-Bragg model with effective parameters of growth of ordered phase and cooperativity. Within the frames of the proposed model the experimental results on stretching of a double-stranded DNA of λ-phage are interpreted. Possible scenarios of homo- and heteropolymeric behavior of DNA during condensation are analyzed. A possible mechanism restricting the growth of linear size of DNA is proposed.     

一类检测DNA凝聚过程中“铰链结构”的氟化物探针

Bloomfield很早指出,在DNA凝聚过程中,急剧弯折会导致DNA分子中出现“铰链(kink)”结构,但未见实验证实。基于紫外可见分光光度法,分别在精胺、亚精胺与DNA凝聚体系中加入氟化物以及氯化物,发现二者虽同为卤族,但对精胺-DNA凝聚体系的效应不同。氟离子使精胺-DNA凝聚体系中出现了“蓝移增色”现象,氯离子仅使该体系出现微弱的增色效应, 而二者对亚精胺-DNA凝聚体系的效应基本相同,均出现微弱增色效应。基于氟离子特殊性质,设计一种能够有效探测DNA凝聚过程中“铰链(kink)”结构氟化物探针,而且证实,第二类“铰链(kink)”结构出现在精胺-DNA凝聚体系中。     

Kinetic accessibility of buried DNA sites in nucleosomes

Using a theoretical model for spontaneous partial DNA unwrapping from histones, we study the transient exposure of protein-binding DNA sites within nucleosomes. We focus on the functional dependence of the rates for site exposure and reburial on the site position, which is measurable experimentally and pertinent to gene regulation. We find the dependence to be roughly described by a random walker model. Close inspection reveals a surprising physical effect of flexibility-assisted barrier crossing, which we characterize within a toy model, the "semiflexible Brownian rotor."