A Molecular Ruler for Measuring Quantitative Distance Distributions

Mathew-Fenn, Rebecca S.; Das, Rhiju; Silverman, Julian; Walker, Peter A.; Harbury, Pehr B.

doi:10.1371/journal.pone.0003229

Cited by 60 publications

(143 citation statements)

References 30 publications

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“…Similar small stretch numbers are shown in Ref. 3 as well. If we now consider that the linear transformation ratio for the helicoidal strip sensor 4 is equal to…”

Section: Calculation Resultssupporting

confidence: 89%

An effective model of DNA like helicoidal structure: with length fluctuation nonlinearity

Tseytlin

2011

AIP Advances

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One of the natural helicoidal nanostructure, which thermomechanical features are studied carefully with the help of different mechanical models, is a DNA cell / molecule. Our study proves that the experimentally determined nonlinear fluctuations of the molecular length of DNA can be better understood by modeling the molecule as a helicoidal pretwisted nanostrip sensor with nonlinear function. The calculations presented here are in good agreement with the experimental data within 10%. Other used by many researchers mechanical models such as an elastic rod, wormlike chain (WLC), accordion bellows, or an elastic core wrapped with rigid wires do not show the possible variance nonlinearity of thermomechanical DNA molecular length fluctuations. We have found that the nonlinear variance of the length fluctuations is an intrinsic property of the micro-nano-sensors with helicoidal shape. This model allows us to estimate the persistence length and twist-stretch coupling of a DNA molecule as well. It also shows the molecule's overwinding possibility at initial stretching with correct numerical representation

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“…Similar small stretch numbers are shown in Ref. 3 as well. If we now consider that the linear transformation ratio for the helicoidal strip sensor 4 is equal to…”

Section: Calculation Resultssupporting

confidence: 89%

An effective model of DNA like helicoidal structure: with length fluctuation nonlinearity

Tseytlin

2011

AIP Advances

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“…The Au-Au center-to-center distance distributions deduced from the Au-Au scattering profiles in C, following the same color code as C. The minor peaks at the short and long distances, outside of the main distribution, are generally noise that is samplepreparation-dependent, as described in ref. 23. (E) The mean Au-Au distance values determined from X-ray interferometry (gray) and predicted from the literature average structure models generated by smFRET (orange) (30).…”

Section: Resultsmentioning

confidence: 99%

“…X-ray interferometry can be used to determine site-tosite distance distributions instantaneously because it relies on atomic scattering (17,(19)(20)(21)(22)(23)(24). Standard small-angle X-ray scattering (SAXS) measures the sum of the scattering and scattering interference from all atoms in a macromolecule (25).…”

Section: Helix-junction-helix | Saxsmentioning

confidence: 99%

From a structural average to the conformational ensemble of a DNA bulge

Shi¹,

Beauchamp

Harbury

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Direct experimental measurements of conformational ensembles are critical for understanding macromolecular function, but traditional biophysical methods do not directly report the solution ensemble of a macromolecule. Small-angle X-ray scattering interferometry has the potential to overcome this limitation by providing the instantaneous distance distribution between pairs of gold-nanocrystal probes conjugated to a macromolecule in solution. Our X-ray interferometry experiments reveal an increasing bend angle of DNA duplexes with bulges of one, three, and five adenosine residues, consistent with previous FRET measurements, and further reveal an increasingly broad conformational ensemble with increasing bulge length. The distance distributions for the AAA bulge duplex (3A-DNA) with six different Au-Au pairs provide strong evidence against a simple elastic model in which fluctuations occur about a single conformational state. Instead, the measured distance distributions suggest a 3A-DNA ensemble with multiple conformational states predominantly across a region of conformational space with bend angles between 24 and 85 degrees and characteristic bend directions and helical twists and displacements. Additional X-ray interferometry experiments revealed perturbations to the ensemble from changes in ionic conditions and the bulge sequence, effects that can be understood in terms of electrostatic and stacking contributions to the ensemble and that demonstrate the sensitivity of X-ray interferometry. Combining X-ray interferometry ensemble data with molecular dynamics simulations gave atomic-level models of representative conformational states and of the molecular interactions that may shape the ensemble, and fluorescence measurements with 2-aminopurinesubstituted 3A-DNA provided initial tests of these atomistic models. More generally, X-ray interferometry will provide powerful benchmarks for testing and developing computational methods. helix-junction-helix | SAXSA grand challenge in biology is to understand the complex free-energy landscape of macromolecules and to decipher the resulting conformational ensembles. To perform their biological functions, macromolecules must adopt a multiplicity of conformations. Balancing and controlling different conformational states is central to biological processes including protein folding, allostery and signaling, and the stepwise assembly and function of macromolecular machines. To understand these complex molecules requires characterization of their free-energy landscapes-i.e., their equilibrium conformational ensembles. Precise measurements of conformational ensembles could allow quantitative modeling of the folding and function of biological macromolecules, would provide valuable experimental data to test current computational models and assumptions, and might facilitate the rational design of specifically acting inhibitors (1, 2).Techniques including NMR and EPR relaxation have been developed to incisively probe motions in the ensemble on different time scales, ranging from pic...

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“…To characterize DNA conformational changes acting in MMR quantitatively, we herein build upon previous SAXS studies of naked DNA with gold nanocrystal labels (9)(10)(11). Our results show MutS bending of mispair-containing DNAs is greater than for complimentary sequence DNA (csDNA), that the ATP-mediated conversion of MutS to a sliding clamp involves loss of the DNA bend, and that MutL does not function to stabilize a MutSmediated bend, in contrast to some MMR models.…”

Section: Significancementioning

confidence: 97%

“…Gold-labeled DNA enables measurement over length scales sufficient to accommodate several proteins to identify cooperative effects on DNA. Because X-rays scatter predominantly from electrons, using heavy atom labels (7)(8)(9)(10)(11) provides high contrast relative to organic molecules. By using labels of moderate size (∼5 nm), the scattering from gold nanocrystals dominates all other scattering signals by three orders of magnitude, thereby reducing analysis complexity while minimizing nanocrystal influence on biological macromolecules.…”

mentioning

confidence: 99%

DNA conformations in mismatch repair probed in solution by X-ray scattering from gold nanocrystals

Hura

Tsai

Claridge

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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DNA metabolism and processing frequently require transient or metastable DNA conformations that are biologically important but challenging to characterize. We use gold nanocrystal labels combined with small angle X-ray scattering to develop, test, and apply a method to follow DNA conformations acting in the Escherichia coli mismatch repair (MMR) system in solution. We developed a neutral PEG linker that allowed gold-labeled DNAs to be flashcooled and stored without degradation in sample quality. The 1,000-fold increased gold nanocrystal scattering vs. DNA enabled investigations at much lower concentrations than otherwise possible to avoid concentration-dependent tetramerization of the MMR initiation enzyme MutS. We analyzed the correlation scattering functions for the nanocrystals to provide higher resolution interparticle distributions not convoluted by the intraparticle distribution. We determined that mispair-containing DNAs were bent more by MutS than complementary sequence DNA (csDNA), did not promote tetramer formation, and allowed MutS conversion to a sliding clamp conformation that eliminated the DNA bends. Addition of second protein responder MutL did not stabilize the MutS-bent forms of DNA. Thus, DNA distortion is only involved at the earliest mispair recognition steps of MMR: MutL does not trap bent DNA conformations, suggesting migrating MutL or MutS/MutL complexes as a conserved feature of MMR. The results promote a mechanism of mismatch DNA bending followed by straightening in initial MutS and MutL responses in MMR. We demonstrate that small angle X-ray scattering with gold labels is an enabling method to examine protein-induced DNA distortions key to the DNA repair, replication, transcription, and packaging. D NA is frequently considered a passive component in interactions with proteins involved in DNA metabolism. Despite this view, many proteins use DNA structural features to mediate catalysis and identify damaged DNA through the effects of damage on DNA rigidity and conformation (1-6). The view of DNA as a passive element is therefore at least in part due to a paucity of robust tools to examine dynamic DNA conformational states during multistep reactions. Gold-labeled DNA enables measurement over length scales sufficient to accommodate several proteins to identify cooperative effects on DNA. Because X-rays scatter predominantly from electrons, using heavy atom labels (7-11) provides high contrast relative to organic molecules. By using labels of moderate size (∼5 nm), the scattering from gold nanocrystals dominates all other scattering signals by three orders of magnitude, thereby reducing analysis complexity while minimizing nanocrystal influence on biological macromolecules. Importantly, small angle X-ray scattering (SAXS) provides global information on conformations adopted by a population of macromolecules in almost any solution condition (12)(13)(14).Mismatch repair (MMR) is an evolutionarily conserved process that corrects mismatches generated during DNA replication (15,16). Despite the i...

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A Molecular Ruler for Measuring Quantitative Distance Distributions

Cited by 60 publications

References 30 publications

An effective model of DNA like helicoidal structure: with length fluctuation nonlinearity

An effective model of DNA like helicoidal structure: with length fluctuation nonlinearity

From a structural average to the conformational ensemble of a DNA bulge

DNA conformations in mismatch repair probed in solution by X-ray scattering from gold nanocrystals

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