The kinetics of force-dependent hybridization and strand-peeling of short DNA fragments

Deoxyribonucleic acid (DNA) carries the genetic information in all living organisms. It consists of two interwound single-stranded (ss) strands, forming a double-stranded (ds) DNA with a right-handed double-helical conformation. The two strands are held together by highly specific basepairing interactions and are further stabilized by stacking between adjacent basepairs. A transition from a dsDNA to two separated ssDNA is called melting and the reverse transition is called hybridization. Applying a tensile force to a dsDNA can result in a particular type of DNA melting, during which one ssDNA strand is peeled away from the other. In this work, we studied the kinetics of strand-peeling and hybridization of short DNA under tensile forces. Our results show that the force-dependent strand-peeling and hybridization can be described with a simple two-state model. Importantly, detailed analysis of the force-dependent transition rates revealed that the transition state consists of several basepairs dsDNA.     

Insights into the Discrepancy between Single Molecule Experiments

Sharp bending as one of the mechanical properties of double-stranded DNA(dsDNA) on the nanoscale is essential for biological functions and processes. Force sensors with optical readout have been designed to measure the forces inside short, strained loops composed of both dsDNA and single-stranded DNA(ssDNA). Recent FRET singlemolecule experiments were carried out based on the same force sensor design, but provided totally contrary results. In the current work, Monte Carlo simulations were performed under three conditions to clarify the discrepancy between the two experiments. The criterion that the work done by the force exerted on dsDNA by ssDNA should be larger than the nearest-neighbor(NN) stacking interaction energy is used to identify the generation of the fork at the junction of dsDNA and ssDNA. When the contour length of dsDNA in the sensor is larger than its critical length, the fork begins to generate at the junction of dsDNA and ssDNA, even with a kink in dsDNA. The forces inferred from simulations under three conditions are consistent with the ones inferred from experiments, including extra large force and can be grouped into two different states, namely, fork states and kink states. The phase diagrams constructed in the phase space of the NN stacking interaction energy and excited energy indicate that the transition between the fork state and kink state is difficult to identify in the phase space with an ultra small or large number of forks, but it can be detected in the phase space with a medium number of forks and kinks.     

Ashkin-Teller Formalism for Elastic Response of DNA Molecule to External Force and Torque

We propose an Ashkin-Teller-like model for elastic response of DNA molecule to external force and torque. The base-stacking interaction is described in a simple and uniform way. We obtain the phase diagram of dsDNA, and in particular, the transition from 13 form to the S state induced by stretching and twisting. The elastic response of the ssDNA is presented also in a unified formalism. The close relation of dsDNA molecule structure with elastic response is shown clearly. The calculated folding angle of the dsDNA molecule is 59.2°.     

Biophysical characterization of DNA binding from single molecule force measurements

Single molecule force spectroscopy is a powerful method that uses the mechanical properties of DNA to explore DNA interactions. Here we describe how DNA stretching experiments quantitatively characterize the DNA binding of small molecules and proteins. Small molecules exhibit diverse DNA binding modes, including binding into the major and minor grooves and intercalation between base pairs of double-stranded DNA (dsDNA). Histones bind and package dsDNA, while other nuclear proteins such as high mobility group proteins bind to the backbone and bend dsDNA. Single-stranded DNA (ssDNA) binding proteins slide along dsDNA to locate and stabilize ssDNA during replication. Other proteins exhibit binding to both dsDNA and ssDNA. Nucleic acid chaperone proteins can switch rapidly between dsDNA and ssDNA binding modes, while DNA polymerases bind both forms of DNA with high affinity at distinct binding sites at the replication fork. Single molecule force measurements quantitatively characterize these DNA binding mechanisms, elucidating small molecule interactions and protein function.     

Interactions between CdSe/CdS quantum dots and DNA through spectroscopic and electrochemical methods

The interaction of CdSe/CdS quantum dots (QDs) with Herring sperm-DNA (hs-DNA) has been studied by UV-vis spectroscopy and electrochemical method. Cu(phen)₂^2+/1+ (phen = 1, 10-phenanthroline) was used as an indicator for electroactive dsDNA or ssDNA. The apparent association constant has been deduced (4.94 × 10³ M⁻¹ and 2.39 × 10² M⁻¹) from the absorption spectral changes of the dsDNA-QDs and ssDNA-QDs. The results of dissociation method suggest that Cu(phen)₂^2+/1+ is more easily dissociated from dsDNA or ssDNA modified gold electrode (dsDNA/Au or dsDNA/Au) in presence of QDs. The dissociation rate constant (k) of Cu(phen)₂^2+/1+ on dsDNA/Au is 4.48 times higher than that in absence of QDs, while k is 2.34 times higher than that in absence of QDs on ssDNA/Au in Tris buffer with low ionic strength (pH 7.0, 0.5 mM NaCl). The results illuminate that hs-DNA has high affinity for QDs due to electrostatic force, hydrogen bonds, and van der Waals interactions, and the binding force of QDs with dsDNA is stronger than ssDNA.     

Direct Observation Method of Individual Single-Stranded DNA Molecules Using Fluorescent Replication Protein A

Direct observation studies of single molecules have revealed molecular behaviors usually hidden in the ensemble and time-averaging of bulk experiments. Direct single DNA molecule analysis of DNA metabolism reactions such as DNA replication, repair, and recombination is necessary to fully understand these essential processes. Intercalation of fluorescent dyes such as YOYO-1 and SYTOX Orange has been the standard method for observing single molecules of double-stranded DNA (dsDNA), but effective fluorescent dyes for observing single molecules of single-stranded DNA (ssDNA) have not been found. To facilitate direct single-molecule observations of DNA metabolism reactions, it is necessary to establish methods for discriminating ssDNA and dsDNA. To observe ssDNA directly, we prepared a fusion protein consisting of the 70 kDa DNA-binding domain of replication protein A and enhanced yellow fluorescent protein (RPA-YFP). This fusion protein had ssDNA-binding activity. In our experiments, dsDNA was stained by SYTOX Orange and ssDNA by RPA-YFP, and we succeeded in staining ssDNA and dsDNA by using RPA-YFP and SYTOX Orange simultaneously.     

Monomer dynamics in double- and single-stranded DNA polymers

We present the first measurements of the kinetics of random motion of individual monomers within large polymer coils. We use double- and single-stranded DNA (dsDNA and ssDNA) as models of semiflexible and flexible polymers, respectively. Fluorescence fluctuations of DNA fragments labeled specifically at a single position reveal the time dependence of the DNA monomer's mean-square displacement . The monomer motions within dsDNA and ssDNA coils are characterized by two qualitatively different kinetic regimes: close to proportional to t(2/3) for ssDNA and proportional to sqrt[t] for dsDNA. While the kinetic behavior of ssDNA is consistent with the generally accepted Zimm theory of polymer dynamics, the kinetic behavior of dsDNA monomers is in good agreement with the Rouse model.     

Trivial and non-trivial superconductivity in dsDNA

A double-stranded DNA (dsDNA) is modeled by two coupled one-dimensional Kitaev's chain and the topological superconductivity is studied. It is shown that the zero energy mode exists under some specific conditions. The wave function of zero mode is calculated and it is shown that the Majorana quasi-particles exist on the ends of each strand. By calculating the winding number, we show that the topological phase transition can happen if the hopping integral between two strands is very smaller than the pairing potential between the Cooper pairs. It means that the dsDNA behaves as a trivial superconductor, commonly, but single-stranded DNA (or two coupled ssDNA with very small hopping between them) may behave as a non-trivial superconductor. Finally, we suggest an experimental setup for probable detection of Majorana quasi-particle in DNA.     

Pairing Mismatched ssDNA to dsDNA Studied with Reflectometric Interference Spectroscopy Sensor

The interaction between two single-stranded DNA(ssDNA)molecules as pairing to a double-stranded DNA(dsDNA)molecule is studied by the reflectometric interference spectroscopy(RIFS)technology.A nano-porous anode alumina membrane coated an Au layer is employed as the sensor substrate.The results indicate that when there are mismatched nucleotide bases,the effective optical thicknesses(OT_(eff))have obvious difference,and the changes of OT_(eff)are connected with the sensor layer thickness and the effective refractive index.It is also demonstrated that the RIFS technique can be used to precisely detect the ssDNA molecules with individual base mismatched as pairing to dsDNA.     

Elasticity of strongly stretched ssDNA

We present a simple model which describes elastic response of single-stranded DNA (ssDNA) to stretching, including the regime of very high force (up to 1000 pN). ssDNA is modelled as a discrete persistent chain, whose ground state is a zigzag rather than a straight line configuration. This mimics the underlying molecular architecture and helps to explain the experimentally observed saturation of the stretching curve at very high force.     

Molecular dynamic study of temperature dependence of mechanical properties and plastic inception of CoCrCuFeNi high-entropy alloy

Molecular dynamics simulations are performed to study mechanical characteristics and homogeneous plastic inception of CoCrCuFeNi high-entropy alloy at various temperatures under uniaxial tension. It is found that the elastic modulus and ultimate tensile strength increase with temperature decreasing. A notable softening effect is observed at the elastic deformation stage caused by the decrease of the interatomic force gradient. Extrinsic stacking faults and deformation twins are extensively observed, which are formed via intrinsic stacking faults overlap.     

Sequence dependent rigidity of single stranded DNA

Single stranded DNA (ssDNA) equilibrium dynamics are investigated using a fluorophore/quencher-labeled hairpin structure which thermally fluctuates between open and closed states. Temporal correlations of the fluorescence fluctuations are used to determine the energy barrier to conformational change. We find that ssDNA distortion is purely entropic for poly(T) but requires an additional enthalpy of +0.5 kcal x mol(-1) x base(-1) for poly(A), consistent with the disruption of base stacking. Such sequence dependent dynamics challenge the classical model of ssDNA as a completely flexible coil.     

Comparison of the measured phase diagrams in the force-temperature plane for the unzipping of two different natural DNA sequences

In this work, we consider the critical force required to unzip two different naturally occurring sequences of double-stranded DNA (dsDNA) at temperatures ranging from 20 ^°C to 50 ^°C, where one of the sequences has a 53% average guanine-cytosine (GC) content and the other has a 40% GC content. We demonstrate that the force required to separate the dsDNA of the 53% GC sequence into single-stranded DNA (ssDNA) is approximately 0.5 pN, or approximately 5% greater than the critical force required to unzip the 40% GC sequence at the same temperature. In the temperature range between 20 and 40 ^°C the measured critical forces correspond reasonably well to predictions based on a simple theoretical homopolymeric model, but at temperatures above 40 ^°C the measured critical forces are much smaller than the predicted forces. The correspondence between theory and experiment is not improved by using Monte Carlo simulations that consider the heteropolymeric nature of the sequences.     

Stationary solutions for a modified Peyrard-Bishop DNA model with up to third-neighbor interactions

We investigate a DNA model that takes into account stacking interactions with neighbors up to three bases away. The model is a generalization of the well-known Peyrard-Bishop (PB) model and is motivated by studies that suggest that nearest-neighbor models for base-pair interaction in a DNA chain might not be enough to capture the mechanism and dynamics of DNA base-pair opening. We study stationary solutions of the modified model and investigate their stability. A comparison with the PB model reveals that under a wide range of parameter values the main characteristics of the original model --such as the hyperbolicity of the equilibrium at the origin-- are preserved, but new types of stationary solutions emerge.     

Voltammetric studies on the interaction of epinephrine with DNA base on a glassy carbon electrode modified with Au nanoparticles

Ionics - The interaction of epinephrine (EP) with DNA, including double-strand DNA (dsDNA) and single-strand DNA (ssDNA) were investigated both in Mcllvaine buffer solution and on a glassy carbon...     

Stretching single stranded DNA,a model polyelectrolyte

The elastic properties of single stranded (ss)DNA, studied by pulling on an isolated molecule, are shown to agree with a recent model of ssDNA that takes into account base pairings and screened electrostatic repulsion of the phosphodiester backbone. By an appropriate physicochemical treatment, the pairing interactions were suppressed and ssDNA used as an experimental model for a generic polyelectrolyte. The elastic behavior of such an altered ssDNA deviates strongly from the behavior of an ideal polymer. This deviation is shown to result from the elasticity of the chain and its electrostatic self-avoiding interactions.     

Colorimetric detection of ssDNA in a solution

A solution containing ssDNA was detected by using colloidal gold nanoparticles upon a color change of solution. First of all, we prepared colloidal gold nanoparticles with a size of less than 20 nm by means of citrate reduction of HAuCl₄. Since colloidal gold nanoparticles modified with citrate anions have a negative charge, they are very well dispersed in the solution due to the negative charge repulsion, showing red color. If the electrostatic repulsion is screened by additives such as NaCl, the nanoparticles start to aggregate, leading to a color alteration. We found that the color alteration is retarded when the solution contains ssDNA, which plays a role in preventing the nanoparticles from aggregation. This allows the determination of whether or not ssDNA is present in a solution. However, the color alteration is not retarded when the solution contains dsDNA.     

Oxidation degree dependent adsorption of ssDNA onto graphene-based surface

DNA/GO composite plays a significant role in the research field of biotechnology and nanotechnology, and attracts a great deal of interest. However, it is still unclear how the oxidation degree of the graphene-based surface affects the adsorption process of single-strand DNA(ss DNA). In this paper, based on the molecular dynamics simulations, we find that ss DNA molecule is absorbed on the GO surface in the most stable state with the oxidation degree around 15%. The microscopic mechanism is attributed to the van Der Walls and the electrostatic interactions between the ss DNA molecule and the graphene-based surface, which is accompanied with the π–π stacking and hydrogen bond formation. The number of π–π stacking between ss DNA and GO reaches the maximum value when the oxidation degree is around 15% among all the GO surfaces. Our simulation results also reveal the coexistence of stretched and curved configurations as well as the adsorption orientation of ss DNA on the GO surface. Furthermore, it is found that the absorbed ss DNA molecules are more likely to move on the graphene-based surface of low oxidation degree, especially on pristine graphene. Our work provides the physics picture of ss DNA's physisorption dynamics onto graphene-based surface and it is helpful in designing DNA/GO nanomaterials.     

基于片层光照明的新型单分子横向磁镊

磁镊是一种高精度的单分子技术,它用磁场对连有生物大分子的超顺磁球产生磁力,通过追踪磁球的位置来测量生物大分子的长度信息.磁镊包括横向磁镊和纵向磁镊.纵向磁镊空间精度高,但昂贵;横向磁镊简单便宜,但由于受其成像原理的限制,一般情况下只能连接较长的DNA等生物大分子,且其空间精度较差,进而限制了其应用范围.为了解决这个问题,本文改进了横向磁镊,用片层光照明的方法使光线主要被磁球散射,从而能够直接观察到吸附在样品槽侧壁上的磁球,这使得测量短连接的底物成为可能.对于实际应用的检测,首先测试了包含270 bp发卡结构的0.5μm双链DNA,用其中发卡结构的"折叠-去折叠"跳变过程证明了改进后的横向磁镊的确可以追踪短DNA等生物大分子.然后,进一步用16μm的λ-DNA检验了实验系统.最后,将新型横向磁镊与普通横向磁镊及纵向磁镊在小力和大力条件下拉伸不同长度DNA的噪声进行了比较,发现改进后的横向磁镊在空间精度上明显优于普通横向磁镊,与纵向磁镊相比也无明显差异.以上结果证明了改进后的横向磁镊的精度优势,并扩展了横向磁镊的应用范围.     

Statistical theory of force-induced unzipping of DNA

The unzipping transition under the influence of external force of a dsDNA molecule has been studied using the Peyrard-Bishop Hamiltonian. The critical force F_c(T) for unzipping calculated in the constant force ensemble is found to depend on the potential parameter k which measures the stiffness associated with a single strand of DNA and on D, the well depth of the on-site potential representing the strength of hydrogen bonds in a base pair. The dependence on temperature of F_c(T) is found to be (T_D - T)^1/2 (T_D being the thermal denaturation temperature) with F_c(T_D) = 0 and F_c(0) = . We used the constant extension ensemble to calculate the average force F(y) required to stretch a base pair a y distance apart. The value of F(y) needed to stretch a base pair located far away from the ends of a dsDNA molecule is found twice the value of the force needed to stretch a base pair located at one of the ends to the same distance for y 1.0 . The force F(y) in both cases is found to have a very large value for y 0.2 compared to the critical force found from the constant force ensemble to which F(y) approaches for large values of y. It is shown that the value of F(y) at the peak depends on the value of k which measures the energy barrier associated with the reduction in DNA strand rigidity as one passes from dsDNA to ssDNA and on the value of the depth of the on-site potential. The effect of defects on the position and height of the peak in the F(y) curve is investigated by replacing some of the base pairs including the one being stretched by defect base pairs. The formation and behaviour of a loop of Y shape when one of the ends base pair is stretched and a bubble of ssDNA with the shape of an eye when a base pair far from ends is stretched are investigated.