Honey, I shrunk the DNA: DNA length as a probe for nucleic‐acid enzyme activity

Oijen, Antoine M. van

doi:10.1002/bip.20624

Cited by 25 publications

(23 citation statements)

References 83 publications

(106 reference statements)

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“…Given that the longitudinal axis of the eukaryotic MCM2-7 complex comprises ~115 Å (Remus et al, 2009), MCM2-7 should protect ~34 basepairs of dsDNA (115 Å ÷ 3.4 Å per basepair = 34 basepairs), which likely occurs in the context of pre-replication complexes. Importantly, ssDNA can be extended to 1.7 times the length of B-form DNA (Smith et al, 1996; van Oijen, 2007). Therefore, 20 nt (34 ÷ 1.7) of fully extended ssDNA would suffice to traverse the entire MCM2-7 channel.…”

Section: Discussionmentioning

confidence: 99%

Selective Bypass of a Lagging Strand Roadblock by the Eukaryotic Replicative DNA Helicase

Yardimci

Long

et al. 2011

Cell

317

355

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Summary The eukaryotic replicative DNA helicase, CMG, unwinds DNA by an unknown mechanism. In some models, CMG encircles and translocates along one strand of DNA while excluding the other strand. In others, CMG encircles and translocates along duplex DNA. To distinguish between these models, replisomes were confronted with strand-specific DNA roadblocks in Xenopus egg extracts. A ssDNA translocase should stall at an obstruction on the translocation strand but not the excluded strand, whereas a dsDNA translocase should stall at obstructions on either strand. We found that replisomes bypass large roadblocks on the lagging strand template much more readily than on the leading strand template. Our results indicate that CMG is a 3′ to 5′ ssDNA translocase, consistent with unwinding via “steric exclusion”. Given that MCM2-7 encircles dsDNA in G1, the data imply that formation of CMG in S phase involves remodeling of MCM2-7 from a dsDNA to a ssDNA binding mode.

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Section: Discussionmentioning

confidence: 99%

Selective Bypass of a Lagging Strand Roadblock by the Eukaryotic Replicative DNA Helicase

Yardimci

Long

et al. 2011

Cell

317

355

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“…[23][24][25] Such small fragments are involved in DNA packaging around histones, in genetic material loading in viruses (requiring compaction/bending), in gene transcription and regulation (requiring looping), in gene delivery via small volume carriers for gene therapy, and as engineering material components for nanofabrication. It is calculated 11 that in a mammalian-cell nucleus, the concentration of DNA would be $10 mg/ml.…”

Section: -22mentioning

confidence: 99%

The intrinsic viscosity of linear DNA

2011

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We measured the intrinsic viscosity of very small synthetic DNA molecules, of 20-395 base pairs, and incorporated them in a nearly complete picture for the whole span of molecular weights reported in the literature to date. A major transition is observed at M approximately 2 × 10(6) . It is found that in the range of approximately 7 × 10(3) ≤ M ≤ 2 × 10(6) , the intrinsic viscosity scales as [η] approximately M(1.05) , suggesting that short DNA chains are not as rigid as generally thought. The corresponding scaling for the range of 2 × 10(6) ≤ M ≤ 8 × 10(10) is [η] approximately M(0.69) . A comparison of our results with existing equations, for much narrower data distributions, is made, and the agreement is very satisfactory considering the huge range of data analyzed here. Experimental concerns such as the effect of ionic strength, polydispersity, temperature, and shear rate are discussed in detail. Some issues concerning the Huggins coefficient, polymer chain stiffness, and the relationship between the Mark-Houwink constants K, α are also presented; it is found that log K = 1.156 - 6.19α.

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“…However, studies by van Oijen and colleagues have used flow-stretched DNA molecules. 3,60 The force exerted on the DNA by the flowing buffer can be calculated based on both the end-to-end length of the DNA as well as the magnitude of the fluctuations exhibited by the beads. Because ssDNA and dsDNA exhibit distinct force-extension profiles, this flow-stretching approach to TPM is particularly well suited to studying systems that involve the conversion of dsDNA to ssDNA or vice versa, such as single molecules of λ exonuclease and single DNA replication forks.…”

Section: Tethered Particle Motion (Tpm)mentioning

confidence: 99%

“…[1][2][3] For example, groundbreaking studies in single-molecule visualization revealed many aspects of the dynamics of F1-ATPase rotary motor, including rotation rates and details of the mechanochemical reaction cycle. [4][5][6][7] Several studies also probed the behavior of mechanoenzymes such as myosin, kinesin and dynein; most prominent were those techniques that measured the step size and force exerted by these proteins as they travel along actin filaments or microtubules.…”

Section: Introductionmentioning

confidence: 99%

The importance of surfaces in single-molecule bioscience

2008

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The last ten years have witnessed an explosion of new techniques that can be used to probe the dynamic behavior of individual biological molecules, leading to discoveries that would not have been possible with more traditional biochemical methods. A common feature among these singlemolecule approaches is the need for the biological molecules to be anchored to a solid support surface. This must be done under conditions that minimize nonspecific adsorption without compromising the biological integrity of the sample. In this review we highlight why surface attachments are a critical aspect of many single-molecule studies and we discuss current methods for anchoring biomolecules. Finally, we provide a detailed description of a new method developed by our laboratory for anchoring and organizing hundreds of individual DNA molecules on a surface, allowing "high-throughput" studies of protein-DNA interactions at the single-molecule level.

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Honey, I shrunk the DNA: DNA length as a probe for nucleic‐acid enzyme activity

Cited by 25 publications

References 83 publications

Selective Bypass of a Lagging Strand Roadblock by the Eukaryotic Replicative DNA Helicase

Selective Bypass of a Lagging Strand Roadblock by the Eukaryotic Replicative DNA Helicase

The intrinsic viscosity of linear DNA

The importance of surfaces in single-molecule bioscience

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