Two Adhesive Sites Can Enhance the Knotting Probability of DNA.
Ontology highlight
ABSTRACT:
SUBMITTER: Najafi S
PROVIDER: S-EPMC4489926 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:During DNA replication in living cells some DNA knots are inadvertently produced by DNA topoisomerases facilitating progression of replication forks. The types of DNA knots formed are conditioned by the 3D organization of replicating DNA molecules. Therefore, by characterizing formed DNA knots it is possible to infer the 3D arrangement of replicating DNA molecules. This topological inference method is highly developed for knotted DNA circles. However, partially replicated DNA molecules have the form of ?-curves. In this article, we use mathematical formalism of ?-curves to characterize the full possibilities of how knotting can occur during replication of DNA molecules in vivo. To do this, we reanalyze earlier experimental studies of knotted, partially replicated DNA molecules and the previously proposed pathway of their formation. We propose a general model of knotting in replication intermediates, and demonstrate that there is an additional, equally important, parallel knotting pathway that also explains how DNA topoisomerases can produce experimentally observed knotted ?-curves. Interestingly, both pathways require intertwining of freshly replicated sister duplexes (precatenanes).
| S-EPMC6158496 | biostudies-literature
Project description:Recent experiments showed that the linear double-stranded DNA in bacteriophage capsids is both highly knotted and neatly structured. What is the physical basis of this organization? Here we show evidence from stochastic simulation techniques that suggests that a key element is the tendency of contacting DNA strands to order, as in cholesteric liquid crystals. This interaction favors their preferential juxtaposition at a small twist angle, thus promoting an approximately nematic (and apolar) local order. The ordering effect dramatically impacts the geometry and topology of DNA inside phages. Accounting for this local potential allows us to reproduce the main experimental data on DNA organization in phages, including the cryo-EM observations and detailed features of the spectrum of DNA knots formed inside viral capsids. The DNA knots we observe are strongly delocalized and, intriguingly, this is shown not to interfere with genome ejection out of the phage.
| S-EPMC2799769 | biostudies-literature
Project description:Dinuclear trihydroxido-bridged osmium-arene complexes are inert and biologically inactive, but we show here that linking dihydroxido-bridged Os(II) -arene fragments by a bridging di-imine to form a metallacycle framework results in strong antiproliferative activity towards cancer cells and distinctive knotting of DNA. The shortened spacer length reduces biological activity and stability in solution towards decomposition to biologically inactive dimers. Significant differences in behavior toward plasmid DNA condensation are correlated with biological activity.
| S-EPMC4982093 | biostudies-literature
Project description:We present a method to obtain numerically accurate values of configurational free energies of semiflexible macromolecular systems, based on the technique of thermodynamic integration combined with normal-mode analysis of a reference system subject to harmonic constraints. Compared with previous free-energy calculations that depend on a reference state, our approach introduces two innovations, namely, the use of internal coordinates to constrain the reference states and the ability to freely select these reference states. As a consequence, it is possible to explore systems that undergo substantially larger fluctuations than those considered in previous calculations, including semiflexible biopolymers having arbitrary ratios of contour length L to persistence length P. To validate the method, high accuracy is demonstrated for free energies of prime DNA knots with L/P = 20 and L/P = 40, corresponding to DNA lengths of 3000 and 6000 base pairs, respectively. We then apply the method to study the free-energy landscape for a model of a synaptic nucleoprotein complex containing a pair of looped domains, revealing a bifurcation in the location of optimal synapse (crossover) sites. This transition is relevant to target-site selection by DNA-binding proteins that occupy multiple DNA sites separated by large linear distances along the genome, a problem that arises naturally in gene regulation, DNA recombination, and the action of type-II topoisomerases.
| S-EPMC4241824 | biostudies-literature
Project description:The E2F1 transcription factor is a well known regulator of cell proliferation and apoptosis, but its role in the DNA damage response is less clear. Using a local UV irradiation technique and immunofluorescence staining, E2F1 is shown to accumulate at sites of DNA damage. Localization of E2F1 to UV-damaged DNA requires the ATM and Rad3-related (ATR) kinase and serine 31 of E2F1 but not an intact DNA binding domain. E2F1 deficiency does not appear to affect the expression of nucleotide excision repair (NER) factors, such as XPC and XPA. However, E2F1 depletion does impair the recruitment of NER factors to sites of damage and reduces the efficiency of DNA repair. E2F1 mutants unable to bind DNA or activate transcription retain the ability to stimulate NER. These findings demonstrate that E2F1 has a direct, non-transcriptional role in DNA repair involving increased recruitment of NER factors to sites of damage.
| S-EPMC2885209 | biostudies-literature
Project description:Most restriction endonucleases bridge two target sites before cleaving DNA: examples include all of the translocating Type I and Type III systems, and many Type II nucleases acting at their sites. A subset of Type II enzymes, the IIB systems, recognise bipartite sequences, like Type I sites, but cut specified phosphodiester bonds near their sites, like Type IIS enzymes. However, they make two double-strand breaks, one either side of the site, to release the recognition sequence on a short DNA fragment; 34 bp long in the case of the archetype, BcgI. It has been suggested that BcgI needs to interact with two recognition sites to cleave DNA but whether this is a general requirement for Type IIB enzymes had yet to be established. Ten Type IIB nucleases were tested against DNA substrates with one or two copies of the requisite sequences. With one exception, they all bridged two sites before cutting the DNA, usually in concerted reactions at both sites. The sites were ideally positioned in cis rather than in trans and were bridged through 3-D space, like Type II enzymes, rather than along the 1-D contour of the DNA, as seen with Type I enzymes. The standard mode of action for the restriction enzymes that excise their recognition sites from DNA thus involves concurrent action at two DNA sites.
| S-EPMC1892151 | biostudies-literature
Project description:Nerve terminals contain multiple sites specialized for the release of neurotransmitters. Release usually occurs with low probability, a design thought to confer many advantages. High-probability release sites are not uncommon, but their advantages are not well understood. Here, we test the hypothesis that high-probability release sites represent an energy-efficient design. We examined release site probabilities and energy efficiency at the terminals of two glutamatergic motor neurons synapsing on the same muscle fiber in Drosophila larvae. Through electrophysiological and ultrastructural measurements, we calculated release site probabilities to differ considerably between terminals (0.33 versus 0.11). We estimated the energy required to release and recycle glutamate from the same measurements. The energy required to remove calcium and sodium ions subsequent to nerve excitation was estimated through microfluorimetric and morphological measurements. We calculated energy efficiency as the number of glutamate molecules released per ATP molecule hydrolyzed, and high-probability release site terminals were found to be more efficient (0.13 versus 0.06). Our analytical model indicates that energy efficiency is optimal (∼0.15) at high release site probabilities (∼0.76). As limitations in energy supply constrain neural function, high-probability release sites might ameliorate such constraints by demanding less energy. Energy efficiency can be viewed as one aspect of nerve terminal function, in balance with others, because high-efficiency terminals depress significantly during episodic bursts of activity.
| S-EPMC5253075 | biostudies-literature
Project description:Proteins interacting at multiple sites on DNA via looping play an important role in many fundamental biochemical processes. Restriction endonucleases that must bind at two recognition sites for efficient activity are a useful model system for studying such interactions. Here we used single DNA manipulation to study sixteen known or suspected two-site endonucleases. In eleven cases (BpmI, BsgI, BspMI, Cfr10I, Eco57I, EcoRII, FokI, HpaII, NarI, Sau3AI and SgrAI) we found that substitution of Ca2+ for Mg2+ blocked cleavage and enabled us to observe stable DNA looping. Forced disruption of these loops allowed us to measure the frequency of looping and probability distributions for loop size and unbinding force for each enzyme. In four cases we observed bimodal unbinding force distributions, indicating conformational heterogeneity and/or complex binding energy landscapes. Measured unlooping events ranged in size from 7 to 7500 bp and the most probable size ranged from less than 75 bp to nearly 500 bp, depending on the enzyme. In most cases the size distributions were in much closer agreement with theoretical models that postulate sharp DNA kinking than with classical models of DNA elasticity. Our findings indicate that DNA looping is highly variable depending on the specific protein and does not depend solely on the mechanical properties of DNA.
| S-EPMC1474071 | biostudies-literature
Project description:Crystal structures of classical cadherins have revealed two dimeric configurations. In the first, N-terminal beta-strands of EC1 domains 'swap' between partner molecules. The second configuration (the 'X dimer'), also observed for T-cadherin, is mediated by residues near the EC1-EC2 calcium binding sites, and N-terminal beta-strands of partner EC1 domains, though held adjacent, do not swap. Here we show that strand-swapping mutants of type I and II classical cadherins form X dimers. Mutant cadherins impaired for X-dimer formation show no binding in short-time frame surface plasmon resonance assays, but in long-time frame experiments, they have homophilic binding affinities close to that of wild type. Further experiments show that exchange between monomers and dimers is slowed in these mutants. These results reconcile apparently disparate results from prior structural studies and suggest that X dimers are binding intermediates that facilitate the formation of strand-swapped dimers.
| S-EPMC2872554 | biostudies-literature
Project description:The cell microenvironment is composed of extracellular matrix (ECM), which contains specific binding sites that allow the cell to adhere to its surroundings. Cells employ focal adhesion proteins, which must be able to resist a variety of forces to bind to ECM. Current techniques for detecting the spatial arrangement of these adhesions, however, have limited resolution and those that detect adhesive forces lack sufficient spatial characterization or resolution. Using a unique application of force spectroscopy, we demonstrate here the ability to determine local changes in the adhesive property of a fibronectin substrate down to the resolution of the fibronectin antibody-functionalized tip diameter, ~20 nm. To verify the detection capabilities of force spectroscopy mapping (FSM), changes in loading rate and temperature were used to alter the bond dynamics and change the adhesion force. Microcontact printing was also used to pattern fluorescein isothiocyanate-conjugated fibronectin in order to mimic the discontinuous adhesion domains of native ECM. Fluorescent detection was used to identify the pattern while FSM was used to map cell adhesion sites in registry with the initial fluorescent image. The results show that FSM can be used to detect the adhesion domains at high resolution and may subsequently be applied to native ECM with randomly distributed cell adhesion sites.
| S-EPMC2997741 | biostudies-literature