Dynamic Approach to DNA Breathing

Metzler, Ralf; Ambjörnsson, Tobias

doi:10.1007/s10867-005-2410-y

Cited by 16 publications

(21 citation statements)

References 22 publications

(30 reference statements)

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“…Timescales derived from FCS data associated with the closing of individual basepairs range from τ f ∼ 10 −4 to τ f ∼ 10 −5 s [17]; our results are qualitiatively unchanged if we adopt as our timescale any value within this range. We find that within our model the fundamental timescale of Ref [18] (τ f = 28µs) gives good agreement with an experimentally observed rate-dependent unpeeling transition [14,15] (see Figure 5).…”

Section: Model Dynamicssupporting

confidence: 86%

“…Common origins of such slow dynamics include large energy barriers due to strong local interactions, large free energy barriers associated with collective reorganization, and the intrinsically slow evolution of long-wavelength motion. The local interactions disrupted by DNA overstretching are noncovalent in character and far too weak to explain the discrepancy between basic rates of basepair opening (∼ 10 −8 − 10 −4 s [8,17,18]) and apparent relaxation rates (seconds to minutes). The implicated importance of collective dynamics, however, is highly unusual for systems macroscopic in only one dimension, because in one dimension the interfaces between domains do not grow with increasing domain size.…”

Section: Entropy-driven Hysteresismentioning

confidence: 99%

“…Whether it reaches this equilibrium state depends on the ratio of rates for changes in molecular state and pulling: very rapid pulling gives the molecule little chance to equilibrate at a given tension. We assume that the fundamental timescale on which individual bases attempt to change state ('basepair closing time') is τ f = 28µs [18], chosen to agree with basepair fluctuation timescales measured using fluorescence correlation spectroscopy [17,18]. This is then the timescale for a single Monte Carlo sweep.…”

Section: Model Dynamicsmentioning

confidence: 99%

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There and (Slowly) Back Again: Entropy-Driven Hysteresis in a Model of DNA Overstretching

Whitelam

Pronk

Geissler

2008

Biophysical Journal

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When pulled along its axis, double-stranded DNA elongates abruptly at a force of approximately 65 pN. Two physical pictures have been developed to describe this overstretched state. The first proposes that strong forces induce a phase transition to a molten state consisting of unhybridized single strands. The second picture introduces an elongated hybridized phase called S-DNA. Little thermodynamic evidence exists to discriminate directly between these competing pictures. Here we show that within a microscopic model of DNA we can distinguish between the dynamics associated with each. In experiment, considerable hysteresis in a cycle of stretching and shortening develops as temperature is increased. Since there are few possible causes of hysteresis in a system whose extent is appreciable in only one dimension, such behavior offers a discriminating test of the two pictures of overstretching. Most experiments are performed upon nicked DNA, permitting the detachment (unpeeling) of strands. We show that the long-wavelength progression of the unpeeled front generates hysteresis, the character of which agrees with experiment only if we assume the existence of S-DNA. We also show that internal melting can generate hysteresis, the degree of which depends upon the nonextensive loop entropy of single-stranded DNA.

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Section: Model Dynamicssupporting

confidence: 86%

Section: Entropy-driven Hysteresismentioning

confidence: 99%

Section: Model Dynamicsmentioning

confidence: 99%

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There and (Slowly) Back Again: Entropy-Driven Hysteresis in a Model of DNA Overstretching

Whitelam

Pronk

Geissler

2008

Biophysical Journal

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“…Because the negatively charged phosphates of the DNA backbone repel each other, the double helix may spontaneously denature locally, opening up singlestranded zones even under physiological conditions. Salt shields the negative charges and stabilizes the double helix (24). Therefore, the lower salt concentrations required for TFAM-independent transcription could potentially affect the behavior of the DNA and cause breathing of the DNA template.…”

Section: Tfam-depleted Mitochondrial Extracts Cannot Initiate Transcrmentioning

confidence: 99%

Mammalian transcription factor A is a core component of the mitochondrial transcription machinery

Shi

Dierckx

Wanrooij

et al. 2012

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

163

150

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Transcription factor A (TFAM) functions as a DNA packaging factor in mammalian mitochondria. TFAM also binds sequence-specifically to sites immediately upstream of mitochondrial promoters, but there are conflicting data regarding its role as a core component of the mitochondrial transcription machinery. We here demonstrate that TFAM is required for transcription in mitochondrial extracts as well as in a reconstituted in vitro transcription system. The absolute requirement of TFAM can be relaxed by conditions that allow DNA breathing, i.e., low salt concentrations or negatively supercoiled DNA templates. The situation is thus very similar to that described in nuclear RNA polymerase II-dependent transcription, in which the free energy of supercoiling can circumvent the need for a subset of basal transcription factors at specific promoters. In agreement with these observations, we demonstrate that TFAM has the capacity to induce negative supercoils in DNA, and, using the recently developed nucleobase analog FRET-pair tC O -tC nitro , we find that TFAM distorts significantly the DNA structure. Our findings differ from recent observations reporting that TFAM is not a core component of the mitochondrial transcription machinery. Instead, our findings support a model in which TFAM is absolutely required to recruit the transcription machinery during initiation of transcription.T he mtDNA is a double-stranded circular molecule that encodes 22 tRNAs, 2 rRNAs, and 13 subunits of the respiratory chain. Transcription is initiated from two sites, the light-and heavy-strand promoters (LSP and HSP1, respectively), and proceeds to produce near genome-length polycistronic transcripts, which are subsequently processed to generate the individual RNA molecules (1). Transcription from LSP also produces the RNA primers required for initiation of mtDNA replication at the origin of the heavy strand (2-4). In vivo experiments have identified a second transcription initiation site (HSP2) downstream of HSP1, but the sequence requirements of this promoter remain to be defined (5, 6).In budding yeast, the basic machinery for mtDNA transcription consists only of two factors: the mitochondrial RNA polymerase (Rpo41) and its accessory factor Mtf1, also denoted sc-mtTFB (1). In mammalian cells, it has been reported that mitochondrial transcription also requires the transcription factor A (TFAM), a high-mobility group-box (HMG) protein (7,8). TFAM plays a role as an mtDNA packaging factor that can bind, wrap, and bend DNA in a non-sequence-specific manner (9-12). TFAM also binds sequence-specifically to sites upstream (−15 to −35) of the HSP1 and LSP transcription start sites (13,14). The human mitochondrial RNA polymerase (POLRMT) is distantly related to the bacteriophage T7 RNA polymerase and has the capacity to recognize promoter elements (14). In combination, POLRMT, TFAM, and the mammalian Mtf1 homolog TFB2M can initiate transcription from a promoter-containing DNA fragment in vitro (15). TFB2M forms a heterodimeric complex with POLRMT and in...

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“…We calculate the resulting tension subject to the constraint of mechanical equilibrium. Basepair fluctuations are assumed to occur on a timescale derived from fluorescence correlation spectroscopy [36,37,38]; we discuss this choice in Appendix B.…”

Section: Modelmentioning

confidence: 99%

Stretching chimeric DNA: A test for the putative S-form

Whitelam

Pronk

Geissler

2008

The Journal of Chemical Physics

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Double-stranded DNA 'overstretches' at a pulling force of about 65 pN, increasing in length by a factor of 1.7. The nature of the overstretched state is unknown, despite its considerable importance for DNA's biological function and technological application. Overstretching is thought by some to be a force-induced denaturation, and by others to consist of a transition to an elongated, hybridized state called S-DNA. Within a statistical mechanical model we consider the effect upon overstretching of extreme sequence heterogeneity. 'Chimeric' sequences possessing halves of markedly different AT composition elongate under fixed external conditions via distinct, spatially segregated transitions. The corresponding force-extension data display two plateaux at forces whose difference varies with pulling rate in a manner that depends qualitatively upon whether the hybridized S-form is accessible. This observation implies a test for S-DNA that could be performed in experiment. Our results suggest that qualitatively different, spatially segregated conformational transitions can occur at a single thermodynamic state within single molecules of DNA.

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Dynamic Approach to DNA Breathing

Cited by 16 publications

References 22 publications

There and (Slowly) Back Again: Entropy-Driven Hysteresis in a Model of DNA Overstretching

There and (Slowly) Back Again: Entropy-Driven Hysteresis in a Model of DNA Overstretching

Mammalian transcription factor A is a core component of the mitochondrial transcription machinery

Stretching chimeric DNA: A test for the putative S-form

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