DNA Folding Research Articles

Surprising Twists in Nucleosomal DNA with Implication for Higher-order Folding

While nucleosomes are dynamic entities that must undergo structural deformations to perform their functions, the general view from available high-resolution structures is a largely static one. Even though numerous examples of twist defects have been documented, the DNA wrapped around the histone core is generally thought to be overtwisted. Analysis of available high-resolution structures from the Protein Data Bank reveals a heterogeneous distribution of twist along the nucleosomal DNA, with clear patterns that are consistent with the literature, and a significant fraction of structures that are undertwisted. The subtle differences in nucleosomal DNA folding, which extend beyond twist, have implications for nucleosome disassembly and modeled higher-order structures. Simulations of oligonucleosome arrays built with undertwisted models behave very differently from those constructed from overtwisted models, in terms of compaction and inter-nucleosome contacts, introducing configurational changes equivalent to those associated with 2–3 base-pair changes in nucleosome spacing. Differences in the nucleosomal DNA pathway, which underlie the way that DNA enters and exits the nucleosome, give rise to different nucleosome-decorated minicircles and affect the topological mix of configurational states.

Radical cation transfer in a guanine pair: an insight to the G-quadruplex structure role using constrained DFT/MM

DNA G-Quadruplex is highly sensitive to oxidation as their structures include \(\pi\)-stacked guanine quartets allowing fast hole transfer between the nucleobases. These transfers can be described using vertical energy gap and electronic coupling between the different diabatic states at play in a guanine pair. Using classical molecular dynamics simulation and the constrained DFT/MM implementation in deMon2k, we determine these quantities for all the interacting guanine pairs of six G-quadruplex structures including one to four quartets and corresponding to different DNA folding. We then described an uni-directional transfer within a quartet, with high electronic coupling and vertical energy gap values, which contrasts with the hole transfer between \(\pi\)-stacked guanine, bi-directional and corresponding to smaller charge transfer parameters. The influence of the geometrical parameters on the electronic coupling is explored, while the external or internal position of the guanine may impact its oxidation probability according to the vertical energy gaps.

DNA looping by protamine follows a nonuniform spatial distribution

DNA looping plays an important role in cells in both regulating and protecting the genome. Often, studies of looping focus on looping by prokaryotic transcription factors like lac repressor or by structural maintenance of chromosomes proteins such as condensin. Here, however, we are interested in a different looping method whereby condensing agents (charge ≥+3) such as protamine proteins neutralize the DNA, causing it to form loops and toroids. We considered two previously proposed mechanisms for DNA looping by protamine. In the first mechanism, protamine stabilizes spontaneous DNA fluctuations, forming randomly distributed loops along the DNA. In the second mechanism, protamine binds and bends the DNA to form a loop, creating a distribution of loops that is biased by protamine binding. To differentiate between these mechanisms, we imaged both spontaneous and protamine-induced loops on short-length (≤1 μm) DNA fragments using atomic force microscopy. We then compared the spatial distribution of the loops to several model distributions. A random looping model, which describes the mechanism of spontaneous DNA folding, fit the distribution of spontaneous loops, but it did not fit the distribution of protamine-induced loops. Specifically, it failed to predict a peak in the spatial distribution of loops at an intermediate location along the DNA. An electrostatic multibinding model, which was created to mimic the bind-and-bend mechanism of protamine, was a better fit of the distribution of protamine-induced loops. In this model, multiple protamines bind to the DNA electrostatically within a particular region along the DNA to coordinate the formation of a loop. We speculate that these findings will impact our understanding of protamine’s in vivo role for looping DNA into toroids and the mechanism of DNA condensation by condensing agents more broadly.

Real-time nuclear magnetic resonance spectroscopy in the study of biomolecular kinetics and dynamics.

The review describes the application of nuclear magnetic resonance (NMR) spectroscopy to study kinetics of folding, refolding and aggregation of proteins, RNA and DNA. Time-resolved NMR experiments can be conducted in a reversible or an irreversible manner. In particular, irreversible folding experiments pose large requirements for (i)signal-to-noise due to the time limitations and (ii)synchronising of the refolding steps. Thus, this contribution discusses the application of methods for signal-to-noise increases, including dynamic nuclear polarisation, hyperpolarisation and photo-CIDNP for the study of time-resolved NMR studies. Further, methods are reviewed ranging from pressure and temperature jump, light induction to rapid mixing to induce rapidly non-equilibrium conditions required to initiate folding.

Opposite Effects of Potassium Ions on the Thermal Stability of i-Motif DNA in Different Buffer Systems.

i-motifs are noncanonical DNA structures formed via the stack of intercalating hemi-protonated C+: C base pairs in C-rich DNA strands and play essential roles in the regulation of gene expression. Here, we systematically investigated the impacts of K+ on i-motif DNA folding using different buffer systems. We found that i-motif structures display very different Tm values at the same pH and ion strength in different buffer systems. More importantly, K+ disrupts the i-motif formed in the MES and Bis-Tris buffer; however, K+ stabilizes the i-motif in phosphate, citrate, and sodium cacodylate buffers. Next, we selected phosphate buffer and confirmed by single-molecule fluorescence resonance energy transfer that K+ indeed has the stabilizing effect on the folding of i-motif DNA from pH 5.8 to 8.0. Nonetheless, circular dichroism spectra further indicate that the structures formed by i-motif sequences at high K+ concentrations at neutral and alkaline pH are not i-motif but other types of higher-order structures and most likely C-hairpins. We finally proposed the mechanisms of how K+ plays the opposite roles in different buffer systems. The present study may provide new insights into our understanding of the formation and stability of i-motif DNA.

Helicity- and Molecular-Weight-Driven Self-Sorting and Assembly of Helical Polymers towards Two-Dimensional Smectic Architectures and Selectively Adhesive Gels.

Self-sorting plays a crucial role in living systems such as the selective assembly of DNA and specific folding of proteins. However, the self-sorting of artificial helical polymers such as biomacromolecules has rarely been achieved. In this work, single-handed helical poly(phenyl isocyanide)s bearing pyrene (Py) and naphthalene (Np) probes were prepared, which exhibited interesting self-sorting properties driven by both helicity and molecular weight (Mn ) in solution, solid state, gel, and on the gel surface as well. The polymers with the same helix sense and similar Mn can self-sort and assemble into well-defined two-dimensional smectic architectures and form stable gels in organic solvents. In contrast, mixed polymers with opposite handedness or different Mn were repulsive to each other and did not aggregate. Moreover, the gels of helical polymers with the same handedness and similar Mn can recognize themselves and adhere together to form a gel.

Helicity‐ and Molecular‐Weight‐Driven Self‐Sorting and Assembly of Helical Polymers towards Two‐Dimensional Smectic Architectures and Selectively Adhesive Gels

AbstractSelf‐sorting plays a crucial role in living systems such as the selective assembly of DNA and specific folding of proteins. However, the self‐sorting of artificial helical polymers such as biomacromolecules has rarely been achieved. In this work, single‐handed helical poly(phenyl isocyanide)s bearing pyrene (Py) and naphthalene (Np) probes were prepared, which exhibited interesting self‐sorting properties driven by both helicity and molecular weight (Mn) in solution, solid state, gel, and on the gel surface as well. The polymers with the same helix sense and similar Mn can self‐sort and assemble into well‐defined two‐dimensional smectic architectures and form stable gels in organic solvents. In contrast, mixed polymers with opposite handedness or different Mn were repulsive to each other and did not aggregate. Moreover, the gels of helical polymers with the same handedness and similar Mn can recognize themselves and adhere together to form a gel.

DNA Folding by Multivalent Cations

Folding short lengths of DNA into loops is a fundamental area of research in the field of nanoengineering. While physical mechanisms of DNA folding may differ according to the multivalent cations involved in folding, the mechanical task is the same. Here, we compare the folding pathways and resulting loops for DNA folding by multivalent cations that are known to condense DNA. Specifically, we compared the looping of DNA by protamine (+21 and 5 kDA), hexaammine-cobalt (III) (+3 and 0.3 kDa), spermine (+4 and 0.2 kDa), and histone H1 (+42 and 20 kDA).

DNA Folding by the SARS-CoV-2 Nucleocapsid Protein

The SARS-CoV-2 virus has disrupted life worldwide. The nucleocapsid protein (N-protein) of SARS-CoV-2 is responsible for packaging the viral genome. We are interested in the mechanism of viral packaging and DNA condensation. Understanding this mechanism could help in developing an antiviral that targets viral packaging. Recently, we discovered that the protamine protein that condenses DNA in sperm binds and bends DNA into a loop. We wondered if viral N-proteins use the same mechanism for folding, since protamine and the N-protein both have large positive charges.

Probing the Contributions to DNA Hybridization Dynamics with Time-Resolved Infrared Spectroscopy

DNA can undergo a diverse set of folding and hybridization transitions of which the energetics and dynamics are highly sensitive to the local solution environment as well as nucleobase chemical modifications. Such sensitivity is used to perform various biological functions and can fuel many nanotechnology applications. However, the physical properties of folding and hybridization reactions that underly these applications often remain poorly understood. DNA folding and hybridization is complicated by many fast structural motions, such as fraying and sliding, and therefore investigation of DNA folding and hybridization requires the ability to probe DNA structure across both fast (ns-µs) and slow (ms-s) timescales. infrared (IR) spectroscopy offers the capability to probe dynamics from ultrafast timescales to seconds with nucleobase-specific information. Along these lines our group has developed temperature-jump (T-jump) IR and two-dimensional IR (2D IR) methods that can monitor DNA interactions from nanoseconds to many seconds. We are investigating DNA unfolding and duplex dehybridization and how they are modulated by nucleobase sequence, chemical modifications, and nucleobase protonation using T-jump IR spectroscopy. Recently, we have found that the modified nucleobases 5-formyl- and 5-carboxylcytosine can alter DNA base pairing and hybridization with great sensitivity to solution pH. Our results provide insight into a complex interplay between sequence, chemical modification, and sequence that may tune DNA hybridization dynamics.

On the thermodynamics of folding of an i-motif DNA in solution under favorable conditions.

Under slightly acidic conditions, cytosine-rich DNA sequences can form non-canonical secondary structures called i-motifs, which occur as four stretches of cytosine repeats form hemi-protonated C·C+ base pairs. The growing interest in the i-motif structures as important components in functional DNA-based nanotechnology or as potential targets of anticancer drugs, increases the need for a deep understanding of the energetics of their structural transitions. Here, a combination of spectroscopic and calorimetric techniques is used to unravel the thermodynamics of folding of an i-motif DNA under favorable conditions. The results give new insights into the energetic aspects of i-motifs and show that thermodynamic and thermal stability are related but not identical properties of such DNA structures.

Applying joint theoretical experimental research to aptamer modeling

The aim of the research. In this work we studied the structure of LC-18 DNA aptamer, which exhibits specifi c binding to lung adenocarcinoma cells. Obtaining the 3D structure of the aptamer is necessary for understanding the mechanism of binding of the aptamer to the target. Th erefore, the aim of the research was modeling of the LC-18 aptamer spatial structure using combination of theoretical methods: DNA folding tools, quantum-chemical calculations and molecular dynamic simulations. Material and methods. Th e secondary structure of the LC-18 aptamer was predicted by using OligoAnalyzer and MFold online soft ware under the conditions typical small-angle X-ray scattering (SAXS) experiment. Th e molecular modeling of the aptamer was carried out using the Avogadro program. For prediction of the structure two computational methods were used: quantum-mechanical method with third-order density-functional tight-binding (DFTB3) and molecular dynamics (MD) with force fi elds. Results. In this paper it was shown that molecular simulations can predict structures from the SAXS experiments. OligoAnalyzer and MFold web servers have been used to generate a set of several likely models. However, more accurate calculations have showed that these models do not predict the relative importance of isomers. Meanwhile, application of quantum-chemical and molecular dynamics calculations have showed reliable molecular structures which have a small deviations from the experimental SAXS curves. Conclusion. Th is study demonstrates the approach for modeling 3D structures of DNA-aptamers in solution using both experimental and theoretical methods. It could be very helpful in designing more effi cient aptamers based on results obtained from molecular simulations.

DNA folds threaten genetic stability and can be leveraged for chemotherapy.

Damaging DNA is a current and efficient strategy to fight against cancer cell proliferation. Numerous mechanisms exist to counteract DNA damage, collectively referred to as the DNA damage response (DDR) and which are commonly dysregulated in cancer cells. Precise knowledge of these mechanisms is necessary to optimise chemotherapeutic DNA targeting. New research on DDR has uncovered a series of promising therapeutic targets, proteins and nucleic acids, with application notably via an approach referred to as combination therapy or combinatorial synthetic lethality. In this review, we summarise the cornerstone discoveries which gave way to the DNA being considered as an anticancer target, and the manipulation of DDR pathways as a valuable anticancer strategy. We describe in detail the DDR signalling and repair pathways activated in response to DNA damage. We then summarise the current understanding of non-B DNA folds, such as G-quadruplexes and DNA junctions, when they are formed and why they can offer a more specific therapeutic target compared to that of canonical B-DNA. Finally, we merge these subjects to depict the new and highly promising chemotherapeutic strategy which combines enhanced-specificity DNA damaging and DDR targeting agents. This review thus highlights how chemical biology has given rise to significant scientific advances thanks to resolutely multidisciplinary research efforts combining molecular and cell biology, chemistry and biophysics. We aim to provide the non-specialist reader a gateway into this exciting field and the specialist reader with a new perspective on the latest results achieved and strategies devised.

A machine learning framework for the prediction of chromatin folding in Drosophila using epigenetic features.

Technological advances have lead to the creation of large epigenetic datasets, including information about DNA binding proteins and DNA spatial structure. Hi-C experiments have revealed that chromosomes are subdivided into sets of self-interacting domains called Topologically Associating Domains (TADs). TADs are involved in the regulation of gene expression activity, but the mechanisms of their formation are not yet fully understood. Here, we focus on machine learning methods to characterize DNA folding patterns in Drosophila based on chromatin marks across three cell lines. We present linear regression models with four types of regularization, gradient boosting, and recurrent neural networks (RNN) as tools to study chromatin folding characteristics associated with TADs given epigenetic chromatin immunoprecipitation data. The bidirectional long short-term memory RNN architecture produced the best prediction scores and identified biologically relevant features. Distribution of protein Chriz (Chromator) and histone modification H3K4me3 were selected as the most informative features for the prediction of TADs characteristics. This approach may be adapted to any similar biological dataset of chromatin features across various cell lines and species. The code for the implemented pipeline, Hi-ChiP-ML, is publicly available: https://github.com/MichalRozenwald/Hi-ChIP-ML

Diamond B23 CD Imaging of Thin Films of Chiral Materials or Achiral Polymers Coated with Chiral Molecules

The novel vertical sample chamber, developed at the B23 beamline for synchrotron radiation circular dichroism (SRCD), has enabled the Diamond User community to conduct different types of experiments from high throughput CD of protein and DNA folding using 96-well multiplates to CD imaging at high spatial resolution. Here, we present the application of CD imaging to large areas of achiral polymer PVA films doped with D-dopa to assess the chiral homogeneity of the film preparation with potential antimicrobial property. Synopsis: CDi application of Diamond B23 SRCD beamline.

Real-Time Conformational Change Monitoring of G-Quadruplex Using Capillary-Based Biocompatible Whispering Gallery Mode Microresonator

In this paper, a capillary-based microfluidic whispering gallery mode (WGM) microresonator is proposed to dynamically monitor the conformation change of G-quadruplex driven by K+ ion. We have theoretically analyzed the resonance wavelength shift mechanism for the DNA folding process, and the resonance mode with the largest wavelength sensitivity of 995.63 nm/RIU according to our simulation result is selected for monitoring of the DNA folding as well as the PGQS immobilization processes. The experimentally observed resonance wavelength shift caused by the immobilization of streptavidin is in accordance with our theoretical analysis on the WGM properties of the proposed microresonator. Moreover, experimental results indicate that PGQS could be folded with a KCl concentration down to 1 μM and a resonance wavelength shift rate of 18.85 × 10 <sup xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">-5</sup> nm/s, which decreases to 2.03 × 10 <sup xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">-5</sup> nm/s when the KCl concentration increases to 10 mM due to the reduction of available PGQS folding sites. Our proposed WGM-microresonator-based measurement platform possesses many desirable merits such as high sensitivity, high integration, microfluidic, label-free and real-time monitoring operation, which make it a promising candidate for DNA sensing, measurement of trivial amount biochemical analyte and related applications.

Searching time operation reduced IPV6 matching through dynamic DNA routing table for less memory and fast IP processing

The internet technology needs high-speed (Internet Protocol) IP addressing system and IP matching in router. The IP addressing speed depends on the size of address and destination. Until now, IPV4 address matching performs with various algorithms such as binary trie, LC trie, prefix tree, priority trie, DTBM, and multi-prefix tree. The above algorithms suit for IPV4 address matching and never suit for IPV6 address matching. The IPV6 consists of 128 bits with different header structures when compared to IPV4. The IPV6 address matching needs a robust algorithm with high processing speed IP matching and uses low memory and high accuracy in IP address matching with less operation time. In this paper, we propose DNA-based sequence matching lookup (DSML) algorithm for IPV6 address matching with reduced searching time. The DSML algorithm performs better than existing algorithms such as disjoint prefix tree and multi-index suffix trie because of direct computation of IPv6 matching through DNA folding sequence. The proposed DSML algorithm does the direct computation due to hit score method and never splits the prefix of IPV6. The splitting of prefix in the existing algorithm consumes more time and reduces the accuracy in IPv6 matching. From the experimentation, when compared to existing algorithms the proposed DSML algorithm provides 80% result in terms of speed and scalability.

Origin of heat capacity increment in DNA folding: The hydration effect

BackgroundUnderstanding DNA folding thermodynamics is crucial for prediction of DNA thermal stability. It is now well established that DNA folding is accompanied by a decrease of the heat capacity ∆cp, F, however its molecular origin is not understood. In analogy to protein folding it has been assumed that this is due to dehydration of DNA constituents, however no evidence exists to support this conclusion. MethodsHere we analyze partial molar heat capacity of nucleic bases and nucleosides in aqueous solutions obtained from calorimetric experiments and calculate the hydration heat capacity contribution ∆cphyd. ResultsWe present hydration heat capacity contributions of DNA constituents and show that they correlate with the solvent accessible surface area. The average contribution for nucleic base dehydration is +0.56 J mol−1 K−1 Å−2 and can be used to estimate the ∆cp, F contribution for DNA folding. ConclusionsWe show that dehydration is one of the major sources contributing to the observed ∆cp, F increment in DNA folding. Other possible sources contributing to the overall ∆cp, F should be significant but appear to compensate each other to high degree. The calculated ∆cphyd for duplexes and noncanonical DNA structures agree excellently with the overall experimental ∆cp, F values. By contrast, empirical parametrizations developed for proteins result in poor ∆cphyd predictions and should not be applied to DNA folding. General significanceHeat capacity is one of the main thermodynamic quantities that strongly affects thermal stability of macromolecules. At the molecular level the heat capacity in DNA folding stems from removal of water from nucleobases.

A new method towards the detection of DNA mutation by Surface-Enhanced Raman Spectroscopy

It is generally believed that the self-folding of single-stranded DNA depends on the hydrophobic effect of its internal bases, but the folding of a single-stranded DNA in a solution was not disordered and would be affected by the stacking effect of adjacent bases. In this work, we developed a new method to explore the stacking between adjacent bases using Surface-Enhanced Raman Spectroscopy (SERS) for the first time. Acidic titanium ions were introduced into silver nanoparticles as an aggregating agent (Ag@ITNPs), and obtained a symmetrical spectrum by normalizing the peak to deoxyribose at 955 cm−1. Based on the influence of adjacent base stacking on the spectrum, we first identified the point mutation sites accurately by SERS. Also, the base content and the DNA frameshift mutations in ssDNA were precisely analyzed. This new method has a simple experimental process and can accurately capture the changes in the base ring breathing peak intensity caused by different adjacent bases, and thus will provide potential application value in the field of gene diagnosis.

C-HiC: A High-Resolution Method for Unbiased Chromatin Conformation Capture Targeting Small Locus.

Within the nucleus, precise DNA folding and organization is mandatory for a tight control of gene expression. In the past 20years, a wealth of molecular approaches has unraveled the existence of DNA territories. With the emergence of affordable deep-sequencing approaches, "Cs" techniques such as 4C, 5C, and HiC, to name a few, are now routinely performed by the scientific community in a large number of model systems. We have modified the HiC approach to a capture probe-based version named C-HiC. This updated assay has resulted in an improved throughput analysis, reduced input material, and good repeatability. The protocol described below details our procedure and notes for a C-HiC approach, designed to target only specific portion of a given genome.