The response of single DNA molecules to externally applied forces and torques was directly measured using an angular optical trap. Upon overwinding, DNA buckled abruptly as revealed by a sharp extension drop followed by a torque plateau. When the DNA was held at the buckling transition, its extension hopped rapidly between two distinct states. Furthermore, the initial plectonemic loop absorbed approximately twice as much extension as was absorbed into the plectoneme upon each additional turn. The observed extension change after buckling and the postbuckling torque support a recent DNA elasticity model.The bending and torsional properties of DNA influence numerous cellular processes, notably DNA compaction, replication, transcription, and protein-DNA binding. DNA elasticity regulates how proteins bend and twist DNA upon binding and how translocating molecular motors exert torque and force on their DNA substrates. Single molecule techniques have proven to be powerful approaches for the investigation of the response of DNA to mechanical stress; individual DNA molecules can be stretched and twisted under physiologically relevant conditions. To date the stretching and bending elasticities of DNA have been well characterized through measurements of the force-extension relation of DNA [1,2]. However, somewhat less is known regarding the torsional elasticity of DNA, at least in part due to difficulties in making direct torque measurements. The most prevalent method to twist DNA is to use magnetic tweezers to rotate a magnetic bead via rotation of a magnetic field [3,4]. Twisting DNA can also be achieved by rotation of a micropipette cantilever [5]. These approaches have provided many important insights into DNA torsional properties even without torque detection. A recent and novel technique directly measured torque in DNA via viscous drag force on a small bead attached to the side of a DNA molecule [6]. This approach requires taut DNA to minimize writhe and thus is more suited for measurements under high force ( > 15 pN). More recently, an angular optical trap that we developed has permitted simultaneous and direct measurements of force and torque for concurrent observation of the tensile and torsional behaviors of DNA over broad ranges of forces and torques [7,8]. In addition, its wider bandwidth is well suited for detection of highly kinetic processes. Previously we showed that nanofabricated quartz cylinders are ideally suited as handles for angular trapping. During DNA supercoiling, the torque, angle, force, and extension of a DNA molecule can be simultaneously monitored at kHz rates. In this work, we have directly measured the torsional modulus of DNA in the intermediate force regime, determined basic relations regarding the dependence of torque on applied force, made the first observation of the abrupt formation of the initial plectoneme † Corresponding author. mwang@physics.cornell.edu. * Present address: Mayo Clinic, Rochester, Minnesota 55902, USA. (interwound loop) in positively supercoiled DNA, and mo...
DNA melting under torsion plays an important role in a wide variety of cellular processes. In the present Letter, we have investigated DNA melting at the single-molecule level using an angular optical trap. By directly measuring force, extension, torque, and angle of DNA, we determined the structural and elastic parameters of torsionally melted DNA. Our data reveal that under moderate forces, the melted DNA assumes a left-handed structure as opposed to an open bubble conformation and is highly torsionally compliant. We have also discovered that at low forces melted DNA properties are highly dependent on DNA sequence. These results provide a more comprehensive picture of the global DNA force-torque phase diagram.
The nucleosome, the fundamental packing unit of chromatin, has a distinct chirality: 147 bp of DNA are wrapped around the core histones in a left-handed, negative superhelix. It has been suggested that this chirality has functional significance, particularly in the context of the cellular processes that generate DNA supercoiling, such as transcription and replication. However, the impact of torsion on nucleosome structure and stability is largely unknown. Here we perform a detailed investigation of single nucleosome behavior on the high affinity 601 positioning sequence under tension and torque using the angular optical trapping technique. We find that torque has only a moderate effect on nucleosome unwrapping. In contrast, we observe a dramatic loss of H2A/H2B dimers upon nucleosome disruption under positive torque, while (H3/H4)2 tetramers are efficiently retained irrespective of torsion. These data indicate that torque could regulate histone exchange during transcription and replication.
Methods for exerting and measuring forces on single molecules have revolutionized the study of the physics of biology. However, it is often the case that biological processes involve rotation or torque generation, and these parameters have been more difficult to access experimentally. Recent advances in the single molecule field have led to the development of techniques which add the capability of torque measurement. By combining force, displacement, torque, and rotational data, a more comprehensive description of the mechanics of a biomolecule can be achieved. In this review, we highlight a number of biological processes for which torque plays a key mechanical role. We describe the various techniques that have been developed to directly probe the torque experienced by a single molecule, and detail a variety of measurements made to date using these new technologies. We conclude by discussing a number of open questions and propose systems of study which would be well suited for analysis with torsional measurement techniques.
As a single DNA molecule is positively supercoiled under constant tension, its extension initially increases due to a negative twist-stretch coupling1, 2. The subsequent attainment of an extension maximum has previously been assumed to be indicative of the onset of a phase transition from B- to scP-DNA2. Here we show that an extension maximum in fact does not coincide with the onset of a phase transition. This transition is evidenced by a direct observation of a torque plateau using an angular optical trap. Instead we find that the shape of the extension curve can be well explained with a theory by John Marko3 that incorporates both DNA twist-stretch coupling and bending fluctuations. This theory also provides a more accurate method of determining the value of the twist-stretch coupling modulus, which has possibly been underestimated in previous studies that did not take into consideration the bending fluctuations2. Our study demonstrates the importance of torque detection in the correct identification of phase transitions as well as the contribution of the twist-stretch coupling and bending fluctuations to DNA extension.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.