Browse
Submit Data
Databases
API
Help

Dataset Information

18 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Mechanical diversity and folding intermediates of parallel-stranded G-quadruplexes with a bulge.

ABSTRACT: A significant number of sequences in the human genome form noncanonical G-quadruplexes (G4s) with bulges or a guanine vacancy. Here, we systematically characterized the mechanical stability of parallel-stranded G4s with a one to seven nucleotides bulge at various positions. Our results show that G4-forming sequences with a bulge form multiple conformations, including fully-folded G4 with high mechanical stability (unfolding forces > 40 pN), partially-folded intermediates (unfolding forces < 40 pN). The folding probability and folded populations strongly depend on the positions and lengths of the bulge. By combining a single-molecule unfolding assay, dimethyl sulfate (DMS) footprinting, and a guanine-peptide conjugate that selectively stabilizes guanine-vacancy-bearing G-quadruplexes (GVBQs), we identified that GVBQs are the major intermediates of G4s with a bulge near the 5' or 3' ends. The existence of multiple structures may induce different regulatory functions in many biological processes. This study also demonstrates a new strategy for selectively stabilizing the intermediates of bulged G4s to modulate their functions.

SUBMITTER: Zhang Y

PROVIDER: S-EPMC8266575 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Parallel G-triplexes and G-hairpins as potential transitory ensembles in the folding of parallel-stranded DNA G-Quadruplexes.

Project description:Guanine quadruplexes (G4s) are non-canonical nucleic acids structures common in important genomic regions. Parallel-stranded G4 folds are the most abundant, but their folding mechanism is not fully understood. Recent research highlighted that G4 DNA molecules fold via kinetic partitioning mechanism dominated by competition amongst diverse long-living G4 folds. The role of other intermediate species such as parallel G-triplexes and G-hairpins in the folding process has been a matter of debate. Here, we use standard and enhanced-sampling molecular dynamics simulations (total length of ∼0.9 ms) to study these potential folding intermediates. We suggest that parallel G-triplex per se is rather an unstable species that is in local equilibrium with a broad ensemble of triplex-like structures. The equilibrium is shifted to well-structured G-triplex by stacked aromatic ligand and to a lesser extent by flanking duplexes or nucleotides. Next, we study propeller loop formation in GGGAGGGAGGG, GGGAGGG and GGGTTAGGG sequences. We identify multiple folding pathways from different unfolded and misfolded structures leading towards an ensemble of intermediates called cross-like structures (cross-hairpins), thus providing atomistic level of description of the single-molecule folding events. In summary, the parallel G-triplex is a possible, but not mandatory short-living (transitory) intermediate in the folding of parallel-stranded G4.

| S-EPMC6698752 | biostudies-literature

Distance measurements between paramagnetic ligands bound to parallel stranded guanine quadruplexes.

Project description:Aside from a double helix, deoxyribonucleic acid (DNA) folds into non-canonical structures, one of which is the guanine quadruplex. Cationic porphyrins bind guanine quadruplexes, but the effects of ligand binding on the structure of guanine quadruplexes with more than four contiguous guanine quartets remains to be fully elucidated. Double electron-electron resonance (DEER) spectroscopy conducted at 9.5 GHz (X-band) using broadband, shaped inversion pulses was used to measure the distances between cationic copper porphyrins bound to model parallel-stranded guanine quadruplexes with increasing numbers of guanine quartets. A single Gaussian component was found to best model the time domain datasets, characteristic of a 2?:?1 binding stoichiometry between the porphyrins and each quadruplex. The measured Cu(2+)-Cu(2+) distances were found to be linearly proportional with the number of guanines. Rather unexpectedly, the ligand end-stacking distance was found to monotonically decreases the overall quadruplex length was extended, suggesting a conformational change in the quadruplex secondary structure dependent upon the number of successive guanine quartets.

| S-EPMC4981883 | biostudies-literature

Propeller-type parallel-stranded G-quadruplexes in the human c-myc promoter.

Project description:The nuclease-hypersensitivity element III1 in the c-myc promoter is a good anticancer target since it largely controls transcriptional activation of the important c-myc oncogene. Recently, the guanine-rich strand of this element has been shown to form an equilibrium between G-quadruplex structures built from two different sets of G-stretches; two models of intramolecular fold-back antiparallel-stranded G-quadruplexes, called "basket" and "chair" forms, were proposed. Here, we show by NMR that two sequences containing these two sets of G-stretches form intramolecular propeller-type parallel-stranded G-quadruplexes in K(+)-containing solution. The two structures involve a core of three stacked G-tetrads formed by four parallel G-stretches with all anti guanines and three double-chain-reversal loops bridging three G-tetrad layers. The central loop contains two or six residues, while the two other loops contain only one residue.

| S-EPMC4692381 | biostudies-literature

Diversity of Parallel Guanine Quadruplexes Induced by Guanine Substitutions.

Project description:Recently, we reported an inhibitory effect of guanine substitutions on the conformational switch from antiparallel to parallel quadruplexes (G4) induced by dehydrating agents. As a possible cause, we proposed a difference in the sensitivity of parallel and antiparallel quadruplexes to the guanine substitutions in the resulting thermodynamic stability. Reports on the influence of guanine substitutions on the biophysical properties of intramolecular parallel quadruplexes are rare. Moreover, such reports are often complicated by the multimerisation tendencies of parallel quadruplexes. To address this incomplete knowledge, we employed circular dichroism spectroscopy (CD), both as stopped-flow-assisted fast kinetics measurements and end-point measurements, accompanied by thermodynamic analyses, based on UV absorption melting profiles, and electrophoretic methods. We showed that parallel quadruplexes are significantly more sensitive towards guanine substitutions than antiparallel ones. Furthermore, guanine-substituted variants, which in principle might correspond to native genomic sequences, distinctly differ in their biophysical properties, indicating that the four guanines in each tetrad of parallel quadruplexes are not equal. In addition, we were able to distinguish by CD an intramolecular G4 from intermolecular ones resulting from multimerisation mediated by terminal tetrad association, but not from intermolecular G4s formed due to inter-strand Hoogsteen hydrogen bond formation. In conclusion, our study indicates significant variability in parallel quadruplex structures, otherwise disregarded without detailed experimental analysis.

| S-EPMC7503932 | biostudies-literature

Single-Molecule Chemo-Mechanical Spectroscopy Provides Structural Identity of Folding Intermediates.

Project description:Single-molecule force spectroscopy has emerged as a powerful tool for studying the folding of biological macromolecules. Mechanical manipulation has revealed a wealth of mechanistic information on transient and intermediate states. To date, the majority of state assignment of intermediates has relied on empirical demarcation. However, performing such experiments in the presence of different osmolytes provides an alternative approach that reports on the structural properties of intermediates. Here, we analyze the folding and unfolding of T4 lysozyme with optical tweezers under a chemo-mechanical perturbation by adding osmolytes. We find that two unrelated protective osmolytes, sorbitol and trimethylamine-n-oxide, function by marginally decelerating unfolding rates and specifically modulating early events in the folding process, stabilizing formation of an on-pathway intermediate. The chemo-mechanical perturbation provides access to two independent metrics of the relevant states during folding trajectories, the contour length, and the solvent-accessible surface area. We demonstrate that the dependence of the population of the intermediate in different osmolytes, in conjunction with its measured contour length, provides the ability to discriminate between potential structural models of intermediate states. Our study represents a general strategy that may be employed in the structural modeling of equilibrium intermediate states observed in single-molecule experiments.

| S-EPMC4816687 | biostudies-literature

Reconciling Intermediates in Mechanical Unfolding Experiments with Two-State Protein Folding in Bulk.

Project description:Most experimentally well-characterized single domain proteins of less than 100 residues have been found to be two-state folders. That is, only two distinct populations can explain both equilibrium and kinetic measurements. Results from single molecule force spectroscopy, where a protein is unfolded by applying a mechanical pulling force to its ends, have largely confirmed this description for proteins found to be two-state in ensemble experiments. Recently, however, stable intermediates have been reported in mechanical unfolding experiments on a cold-shock protein previously found to be a prototypical two-state folder. Here, we tackle this discrepancy using free energy landscapes and Markov state models derived from coarse-grained molecular simulations. We show that protein folding intermediates can be selectively stabilized by the pulling force and that the populations of these intermediates vary in a force-dependent manner. Our model qualitatively captures the experimental results and suggests a possible origin of the apparent discrepancy.

| S-EPMC5597958 | biostudies-literature

NMR solution and X-ray crystal structures of a DNA molecule containing both right- and left-handed parallel-stranded G-quadruplexes.

Project description:Analogous to the B- and Z-DNA structures in double-helix DNA, there exist both right- and left-handed quadruple-helix (G-quadruplex) DNA. Numerous conformations of right-handed and a few left-handed G-quadruplexes were previously observed, yet they were always identified separately. Here, we present the NMR solution and X-ray crystal structures of a right- and left-handed hybrid G-quadruplex. The structure reveals a stacking interaction between two G-quadruplex blocks with different helical orientations and displays features of both right- and left-handed G-quadruplexes. An analysis of loop mutations suggests that single-nucleotide loops are preferred or even required for the left-handed G-quadruplex formation. The discovery of a right- and left-handed hybrid G-quadruplex further expands the polymorphism of G-quadruplexes and is potentially useful in designing a left-to-right junction in G-quadruplex engineering.

| S-EPMC6735952 | biostudies-literature

Double-stranded flanking ends affect the folding kinetics and conformational equilibrium of G-quadruplexes forming sequences within the promoter of KIT oncogene.

Project description:G-quadruplexes embedded within promoters play a crucial role in regulating the gene expression. KIT is a widely studied oncogene, whose promoter contains three G-quadruplex forming sequences, c-kit1, c-kit2 and c-kit*. For these sequences available studies cover ensemble and single-molecule analyses, although for kit* the latter were limited to a study on a promoter domain comprising all of them. Recently, c-kit2 has been reported to fold according to a multi-step process involving folding intermediates. Here, by exploiting fluorescence resonance energy transfer, both in ensemble and at the single molecule level, we investigated the folding of expressly designed constructs in which, alike in the physiological context, either c-kit2 or c-kit* are flanked by double stranded DNA segments. To assess whether the presence of flanking ends at the borders of the G-quadruplex affects the folding, we studied under the same protocols oligonucleotides corresponding to the minimal G-quadruplex forming sequences. Data suggest that addition of flanking ends results in biasing both the final equilibrium state and the folding kinetics. A previously unconsidered aspect is thereby unravelled, which ought to be taken into account to achieve a deeper insight of the complex relationships underlying the fine tuning of the gene-regulatory properties of these fascinating DNA structures.

| S-EPMC8464035 | biostudies-literature

Frustrated folding of guanine quadruplexes in telomeric DNA.

Project description:Human chromosomes terminate in long, single-stranded, DNA overhangs of the repetitive sequence (TTAGGG)n. Sets of four adjacent TTAGGG repeats can fold into guanine quadruplexes (GQ), four-stranded structures that are implicated in telomere maintenance and cell immortalization and are targets in cancer therapy. Isolated GQs have been studied in detail, however much less is known about folding in long repeat sequences. Such chains adopt an enormous number of configurations containing various arrangements of GQs and unfolded gaps, leading to a highly frustrated energy landscape. To better understand this phenomenon, we used mutagenesis, thermal melting, and global analysis to determine stability, kinetic, and cooperativity parameters for GQ folding within chains containing 8-12 TTAGGG repeats. We then used these parameters to simulate the folding of 32-repeat chains, more representative of intact telomeres. We found that a combination of folding frustration and negative cooperativity between adjacent GQs increases TTAGGG unfolding by up to 40-fold, providing an abundance of unfolded gaps that are potential binding sites for telomeric proteins. This effect was most pronounced at the chain termini, which could promote telomere extension by telomerase. We conclude that folding frustration is an important and largely overlooked factor controlling the structure of telomeric DNA.

| S-EPMC8034632 | biostudies-literature

Changes of protein stiffness during folding detect protein folding intermediates.

Project description:Single-molecule force-quench atomic force microscopy (FQ-AFM) is used to detect folding intermediates of a simple protein by detecting changes of molecular stiffness of the protein during its folding process. Those stiffness changes are obtained from shape and peaks of an autocorrelation of fluctuations in end-to-end length of the folding molecule. The results are supported by predictions of the equipartition theorem and agree with existing Langevin dynamics simulations of a simplified model of a protein folding. In the light of the Langevin simulations the experimental data probe an ensemble of random-coiled collapsed states of the protein, which are present both in the force-quench and thermal-quench folding pathways.

| S-EPMC3923959 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data