Quantifying DNA melting transitions using single-molecule force spectroscopy

Calderon, Christopher P.; Chen, Wei‐Hung; Lin, Kwei-Jay; Harris, Nolan C.; Kiang, Ching‐Hwa

doi:10.1088/0953-8984/21/3/034114

Cited by 53 publications

(135 citation statements)

References 44 publications

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“…These approaches are particularly sensitive to noise models, 61,86,94–96 and for this reason, it is often best to simultaneously infer the model for the system of interest together with the noise model in a self-consistent fashion. The hidden Markov model that we describe in greater depth in section 3.4.1 is one such example.…”

Section: Data-driven Modeling: Key Conceptsmentioning

confidence: 99%

Unraveling the Thousand Word Picture: An Introduction to Super-Resolution Data Analysis

Lee

Tsekouras

Calderon³

et al. 2017

Chem. Rev.

138

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Super-resolution microscopy provides direct insight into fundamental biological processes occurring at length scales smaller than light’s diffraction limit. The analysis of data at such scales has brought statistical and machine learning methods into the mainstream. Here we provide a survey of data analysis methods starting from an overview of basic statistical techniques underlying the analysis of super-resolution and, more broadly, imaging data. We subsequently break down the analysis of super-resolution data into four problems: the localization problem, the counting problem, the linking problem, and what we’ve termed the interpretation problem.

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Section: Data-driven Modeling: Key Conceptsmentioning

confidence: 99%

Unraveling the Thousand Word Picture: An Introduction to Super-Resolution Data Analysis

Lee

Tsekouras

Calderon³

et al. 2017

Chem. Rev.

138

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“…The DNA overstretching transition was originally observed on DNA with open ends (end-opened DNA), which is torsion-unconstrained (5,6). Since its discovery, three transitions have been debated to explain DNA overstretching (4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21) (Fig. 1): (i) "peeling" of one strand from the other to a peeled ssDNA strand under tension while the other ssDNA strand coils, (ii) "inside-strand separation" to two parallel ssDNA strands that share tension (melting bubbles), and (iii) "B-to-S" transition to a novel overstretched dsDNA, termed S-DNA.…”

mentioning

confidence: 99%

“…For end-opened DNA, two kinetically and thermodynamically distinct overstretching transitions have been reported (6)(7)(8)(9)(10)(11)(12), which can be selected by changing base-pair stability through temperature, salt concentration, and DNA sequence (8)(9)(10). One transition is the peeling transition to peeled ssDNA (8)(9)(10)18).…”

mentioning

confidence: 99%

Revealing the competition between peeled ssDNA, melting bubbles, and S-DNA during DNA overstretching by single-molecule calorimetry

Zhang

Chen

et al. 2013

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

127

163

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Double-stranded DNA (dsDNA) unconstrained by torsion undergoes an overstretching transition at about 65 pN, elongating the DNA to about 1.7-fold. Three possible structural transitions have been debated for the nature of DNA overstretching: (i) "peeling" apart of dsDNA to produce a peeled ssDNA strand under tension while the other strand coils, (ii) "inside-strand separation" of dsDNA to two parallel ssDNA strands that share tension (melting bubbles), and (iii) "B-to-S" transition to a novel dsDNA, termed S-DNA. Here we overstretched an end-opened DNA (with one open end to allow peeling) and an end-closed (i.e., both ends of the linear DNA are covalently closed to prohibit peeling) and torsion-unconstrained DNA. We report that all three structural transitions exist depending on experimental conditions. For the end-opened DNA, the peeling transition and the B-to-S transition were observed; for the end-closed DNA, the inside-strand separation and the B-to-S transition were observed. The peeling transition and the inside-strand separation are hysteretic and have an entropy change of approximately 17 cal/(K·mol), whereas the B-to-S transition is nonhysteretic and has an entropy change of approximately −2 cal/(K·mol). The force-extension curves of peeled ssDNA, melting bubbles, and S-DNA were characterized by experiments. Our results provide experimental evidence for the formation of DNA melting bubbles driven by high tension and prove the existence of nonmelted S-DNA. Our findings afford a full understanding of three possible force-driven structural transitions of torsion-unconstrained DNA and the resulting three overstretched DNA structures.DNA bubble | magnetic tweezers

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“…Pulling events capturing multiple tip interactions or oligomers were excluded from data analysis by selecting curves with only one detachment peak and of extension length similar to a single perlecan monomer. The force curves were fitted with either the extensible WLC (eWLC) model [61,62] (for native perlecan with or without GAG),

x = L_{c} true[1 - \frac{1}{{true(4 β L_{p} F true)}^{1 / 2}} + \frac{F}{K} true],

or inextensible WLC model [61,63–65] (for denatured DTT treated perlecan),

F false(x false) = \frac{1}{L_{p} β} true[\frac{1}{4 {false(1 - x / L_{c} false)}^{2}} + \frac{x}{L_{c}} - \frac{1}{4} true],

where x is the extension, F is the force, K is the elastic stretch modulus and β = 1/ k B T where k B is the Boltzmann constant, T is the temperature, L c is the contour length of the total end-to-end length of an extended molecule and L p is the persistence length. Histograms of the L c and L p distributions were fitted to a Gaussian curve to get statistical measures of the two parameters.…”

Section: Methodsmentioning

confidence: 99%

Single molecule force measurements of perlecan/HSPG2: A key component of the osteocyte pericellular matrix

et al. 2016

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Perlecan/HSPG2, a large, monomeric heparan sulfate proteoglycan (HSPG), is a key component of the lacunar canalicular system (LCS) of cortical bone, where it is part of the mechanosensing pericellular matrix (PCM) surrounding the osteocytic processes and serves as a tethering element that connects the osteocyte cell body to the bone matrix. Within the pericellular space surrounding the osteocyte cell body, perlecan can experience physiological fluid flow drag force and in that capacity function as a sensor to relay external stimuli to the osteocyte cell membrane. We previously showed a reduction in perlecan secretion alters the PCM fiber composition and interferes with bone’s response to mechanical loading in vivo. To test our hypothesis that perlecan core protein can sustain tensile forces without unfolding under physiological loading conditions, atomic force microscopy (AFM) was used to capture images of perlecan monomers at nanoscale resolution and to perform single molecule force measurement (SMFMs). We found that the core protein of purified full-length human perlecan is of suitable size to span the pericellular space of the LCS, with a measured end-to-end length of 170 ± 20 nm and a diameter of 2–4 nm. Force pulling revealed a strong protein core that can withstand over 100 pN of tension well over the drag forces that are estimated to be exerted on the individual osteocyte tethers. Data fitting with an extensible worm-like chain model showed that the perlecan protein core has a mean elastic constant of 890 pN and a corresponding Young’s modulus of 71 MPa. We conclude perlecan has physical properties that would allow it to act as a strong but elastic tether in the LCS.

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Quantifying DNA melting transitions using single-molecule force spectroscopy

Cited by 53 publications

References 44 publications

Unraveling the Thousand Word Picture: An Introduction to Super-Resolution Data Analysis

Unraveling the Thousand Word Picture: An Introduction to Super-Resolution Data Analysis

Revealing the competition between peeled ssDNA, melting bubbles, and S-DNA during DNA overstretching by single-molecule calorimetry

Single molecule force measurements of perlecan/HSPG2: A key component of the osteocyte pericellular matrix

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