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Unravelling the structural plasticity of stretched DNA under torsional constraint.

ABSTRACT: Regions of the genome are often held under torsional constraint. Nevertheless, the influence of such constraint on DNA-protein interactions during genome metabolism is still poorly understood. Here using a combined optical tweezers and fluorescence microscope, we quantify and explain how torsional constraint influences the structural stability of DNA under applied tension. We provide direct evidence that concomitant basepair melting and helical unwinding can occur in torsionally constrained DNA at forces >∼50 pN. This striking result indicates that local changes in linking number can be absorbed by the rest of the DNA duplex. We also present compelling new evidence that an overwound DNA structure (likely P-DNA) is created (alongside underwound structures) at forces >∼110 pN. These findings substantiate previous theoretical predictions and highlight a remarkable structural plasticity of torsionally constrained DNA. Such plasticity may be required in vivo to absorb local changes in linking number in DNA held under torsional constraint.

SUBMITTER: King GA

PROVIDER: S-EPMC4897764 | biostudies-literature | 2016 Jun

REPOSITORIES: biostudies-literature

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Unravelling the structural plasticity of stretched DNA under torsional constraint.

King Graeme A GA Peterman Erwin J G EJ Wuite Gijs J L GJ

Nature communications 20160606

Regions of the genome are often held under torsional constraint. Nevertheless, the influence of such constraint on DNA-protein interactions during genome metabolism is still poorly understood. Here using a combined optical tweezers and fluorescence microscope, we quantify and explain how torsional constraint influences the structural stability of DNA under applied tension. We provide direct evidence that concomitant basepair melting and helical unwinding can occur in torsionally constrained DNA ...[more]

PMID: 27263853

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Project description:BACKGROUND: An important step in strain optimization is to identify reactions whose activities should be modified to achieve the desired cellular objective. Preferably, these reactions are identified systematically, as the number of possible combinations of reaction modifications could be very large. Over the last several years, a number of computational methods have been described for identifying combinations of reaction modifications. However, none of these methods explicitly address uncertainties in implementing the reaction activity modifications. In this work, we model the uncertainties as probability distributions in the flux carrying capacities of reactions. Based on this model, we develop an optimization method that identifies reactions for flux capacity modifications to predict outcomes with high statistical likelihood. RESULTS: We compare three optimization methods that select an intervention set comprising up- or down-regulation of reaction flux capacity: CCOpt (Chance constrained optimization), DetOpt (Deterministic optimization), and MCOpt (Monte Carlo-based optimization). We evaluate the methods using a Monte Carlo simulation-based method, MCEval (Monte Carlo Evaluations). We present two case studies analyzing a CHO cell and an adipocyte model. The flux capacity distributions required for our methods were estimated from maximal reaction velocities or elementary mode analysis. The intervention set selected by CCOpt consistently outperforms the intervention set selected by DetOpt in terms of tolerance to flux capacity variations. MCEval shows that the optimal flux predicted based on the CCOpt intervention set is more likely to be obtained, in a probabilistic sense, than the flux predicted by DetOpt. The intervention sets identified by CCOpt and MCOpt were similar; however, the exhaustive sampling required by MCOpt incurred significantly greater computational cost. CONCLUSIONS: Maximizing tolerance to variable engineering outcomes (in modifying enzyme activities) can identify intervention sets that statistically improve the desired cellular objective.

Project description:DNA morphology is subjected to environmental conditions and is closely coupled with its function. For example, DNA experiences stretching forces during several biological processes, including transcription and genome transactions, that significantly alter its conformation from that of B-DNA. Indeed, a well-defined 1.5 times extended conformation of dsDNA, known as Σ-DNA, has been reported in DNA complexes with proteins such as Rad51 and RecA. A striking feature in Σ-DNA is that the nucleobases are partitioned into triplets of three locally stacked bases separated by an empty rise gap of ∼5 Å. The functional role of such a DNA base triplet was hypothesized to be coupled with the ease of recognition of DNA bases by DNA-binding proteins (DBPs) and the physical origin of three letters (codon/anti-codon) in the genetic code. However, the underlying mechanism of base-triplet formation and the ease of DNA base-pair recognition by DBPs remain elusive. To investigate, here, we study the diffusion of a protein on a force-induced stretched DNA using coarse-grained molecular dynamics simulations. Upon pulling at the 3' end of DNA by constant forces, DNA exhibits a conformational transition from B-DNA to a ladder-like S-DNA conformation via Σ-DNA intermediate. The resulting stretched DNA conformations exhibit non-uniform base-pair clusters such as doublets, triplets, and quadruplets, of which triplets are energetically more stable than others. We find that protein favors the triplet formation compared to its unbound form while interacting non-specifically along DNA, and the relative population of it governs the ruggedness of the protein-DNA binding energy landscape and enhances the efficiency of DNA base recognition. Furthermore, we analyze the translocation mechanism of a DBP under different force regimes and underscore the significance of triplet formation in regulating the facilitated diffusion of protein on DNA. Our study, thus, provides a plausible framework for understanding the structure-function relationship between triplet formation and base recognition by a DBP and helps to understand gene regulation in complex regulatory processes.

Dataset Information

Unravelling the structural plasticity of stretched DNA under torsional constraint.

Publications

Unravelling the structural plasticity of stretched DNA under torsional constraint.

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