Quantitative all-atom and coarse-grained molecular dynamics simulation studies of DNA

The remarkable molecule that encodes genetic information for all life on earth—DNA—is a polymer with unusual physical properties. The mechanical and electrostatic properties of DNA are utilized extensively by cells in the replication, regular maintenance, and expression of their genetic material. This can be illustrated by considering the journey of a typical gene regulating protein, the lac repressor, which recognizes a particular gene and prevents its expression. First, the large electrostatic charge density of DNA provides an energetic track that guides the repressor’s search for its target binding site. Next, as the protein moves along the DNA, it attempts to deform the DNA. The repressor is only able to form an active complex with DNA that has the right sequence-dependent flexibility. Finally, the repressor is believed to form a very small DNA loop that prevents the gene from being expressed. The stability of the loop can be expected to depend sensitively on the global flexibility of DNA. Thus, the key to understanding the some of the most important cellular processes lies in understanding the physical properties of DNA. Single-molecule experiments allow direct observation of the behavior of individual DNA molecules, but act on length and timescales that are often too large and fast to observe underlying DNA and DNA–protein dynamics. Acting on length and timescales that complement single-molecule experiments, molecular dynamics simulations provide a high-resolution glimpse into the mechanics of a biomolecular world. Here, several simulation studies are presented, each of which quantified one or more properties of DNA. Specifically, the repulsive forces between parallel duplex DNA molecules were measured; the short-ranged, attractive end-to-end stacking energy was obtained; a single-stranded DNA model was developed that reproduced experimental measurements of its extension upon applied force; and finally the nature of single-stranded DNA binding to a single-stranded DNA binding protein was investigated. These works represent important steps towards larger simulations of more biologically complete DNA–protein systems

End-to-end attraction of duplex DNA

Recent experiments [Nakata, M. et al., End-to-end stacking and liquid crystal condensation of 6 to 20 basepair DNA duplexes. Science 2007; 318:1276–1279] have demonstrated spontaneous end-to-end association of short duplex DNA fragments into long rod-like structures. By means of extensive all-atom molecular dynamic simulations, we characterized end-to-end interactions of duplex DNA, quantitatively describing the forces, free energy and kinetics of the end-to-end association process. We found short DNA duplexes to spontaneously aggregate end-to-end when axially aligned in a small volume of monovalent electrolyte. It was observed that electrostatic repulsion of 5′-phosphoryl groups promoted the formation of aggregates in a conformation similar to the B-form DNA double helix. Application of an external force revealed that rupture of the end-to-end assembly occurs by the shearing of the terminal base pairs. The standard binding free energy and the kinetic rates of end-to-end association and dissociation processes were estimated using two complementary methods: umbrella sampling simulations of two DNA fragments and direct observation of the aggregation process in a system containing 458 DNA fragments. We found the end-to-end force to be short range, attractive, hydrophobic and only weakly dependent on the ion concentration. The relation between the stacking free energy and end-to-end attraction is discussed as well as possible roles of the end-to-end interaction in biological and nanotechnological systems

A synthetic enzyme built from DNA flips 107 lipids per second in biological membranes.

Mimicking enzyme function and increasing performance of naturally evolved proteins is one of the most challenging and intriguing aims of nanoscience. Here, we employ DNA nanotechnology to design a synthetic enzyme that substantially outperforms its biological archetypes. Consisting of only eight strands, our DNA nanostructure spontaneously inserts into biological membranes by forming a toroidal pore that connects the membrane's inner and outer leaflets. The membrane insertion catalyzes spontaneous transport of lipid molecules between the bilayer leaflets, rapidly equilibrating the lipid composition. Through a combination of microscopic simulations and fluorescence microscopy we find the lipid transport rate catalyzed by the DNA nanostructure exceeds 107 molecules per second, which is three orders of magnitude higher than the rate of lipid transport catalyzed by biological enzymes. Furthermore, we show that our DNA-based enzyme can control the composition of human cell membranes, which opens new avenues for applications of membrane-interacting DNA systems in medicine

Optical Voltage Sensing Using DNA Origami.

We explore the potential of DNA nanotechnology for developing novel optical voltage sensing nanodevices that convert a local change of electric potential into optical signals. As a proof-of-concept of the sensing mechanism, we assembled voltage responsive DNA origami structures labeled with a single pair of FRET dyes. The DNA structures were reversibly immobilized on a nanocapillary tip and underwent controlled structural changes upon application of an electric field. The applied field was monitored through a change in FRET efficiency. By exchanging the position of a single dye, we could tune the voltage sensitivity of our DNA origami structure, demonstrating the flexibility and versatility of our approach. The experimental studies were complemented by coarse-grained simulations that characterized voltage-dependent elastic deformation of the DNA nanostructures and the associated change in the distance between the FRET pair. Our work opens a novel pathway for determining the mechanical properties of DNA origami structures and highlights potential applications of dynamic DNA nanostructures as voltage sensors

A nanoscopic glimpse of the cells drafting table

A blueprint for life is contained within the cells of every living organism in the form of DNA molecules. Most of us think about DNA in terms of phenotypes such as brown or blue eyes. However, on the cellular level, specialized molecular machinery operate on DNA to create proteins, repair deadly errors in the genetic code, and replicate the genome. The familiar DNA double-helix is split into its constituent strands during these processes so that the genetic code may be read. Single-stranded DNA binding protein(SSB; depicted in orange) is omnipresent during the processes of DNA replication and repair, where it binds excess single-stranded DNA very tightly to prevent double-stranded DNA from forming. We constructed a coarse-grained model of DNA (bottom right; large beads) and SSB that captures the behavior of all-atom DNA (top left; small spheres). The new model extends the timescale amenable to simulation thousands of times. Existing evidence demonstrated that SSB could wrap DNA in two or three ways,but our simulations revealed at least six distinct yet dynamic wrapping patterns, one of which is depicted. This work represents a step towards our larger goal of constructing a complete model of the DNA replication machinery

SI2-SSE: Collaborative Research: Integrated Tools for DNA Nanostructure Design and Simulation

One slide description of the collaborative project aimed at building a community of tools for structural DNA nanotechnolog

Atomic View of Photosynthetic Metabolite Permeability Pathways and Confinement in Cyanobacterial Carboxysomes

Carboxysomes are protein microcompartments found in cyanobac- teria, whose shell encapsulates rubisco at the heart of carbon fixa- tion in the Calvin-Benson-Bassham (CBB) cycle. Carboxysomes are thought to locally concentrate CO2 and exclude O2 from the shell interior to improve rubisco efficiency through selective metabolite permeability, creating a concentrated catalytic center. However, per- meability coefficients have not previously been determined for these gases, or for CBB cycle intermediates such as bicarbonate (HCO3 – ), 3-phosphoglyceric acid (3-PGA), or ribulose-1,5-bisphosphate (RuBP). Starting from a high resolution cryo-EM structure of a synthetic β-carboxysome shell, we perform unbiased all-atom molecular dynamics (MD) to track metabolite permeability across the shell. The synthetic carboxysome shell is found to have similar permeability coefficients for multiple metabolites, and is not selectively permeable to HCO3– relative to CO2. To resolve how these comparable permeabilities can be reconciled with the clear role of the carboxysome in the carbon-concentrating mechanism in cyanobacteria, complementary atomic-resolution Brownian Dynamics (ARBD) simulations estimate the mean first passage time for CO2 assimilation in a crowded model carboxysome. Despite a relatively high CO2 permeability of 10−2 cm/s across the carboxysome shell, the shell proteins reflect enough CO2 back towards rubisco that over 2600 CO2 molecules can be fixed by rubisco for every 1 CO2 molecule that escapes under typical conditions. The permeabilities determined from all-atom molecular simulation are key inputs into flux modeling, and the insight gained into carbon fixation can facilitate the engineering of carboxysomes and other bacterial microcompartments for multiple applications

Stretching and Controlled Motion of Single-Stranded DNA in Locally Heated Solid-State Nanopores

Practical applications of solid-state nanopores for DNA detection and sequencing require the electrophoretic motion of DNA through the nanopores to be precisely controlled. Controlling the motion of single-stranded DNA presents a particular challenge, in part because of the multitude of conformations that a DNA strand can adopt in a nanopore. Through continuum, coarse-grained and atomistic modeling, we demonstrate that local heating of the nanopore volume can be used to alter the electrophoretic mobility and conformation of single-stranded DNA. In the nanopore systems considered, the temperature near the nanopore is modulated <i>via</i> a nanometer-size heater element that can be radiatively switched on and off. The local enhancement of temperature produces considerable stretching of the DNA fragment confined within the nanopore. Such stretching is reversible, so that the conformation of DNA can be toggled between compact (local heating is off) and extended (local heating is on) states. The effective thermophoretic force acting on single-stranded DNA in the vicinity of the nanopore is found to be sufficiently large (4–8 pN) to affect such changes in the DNA conformation. The local heating of the nanopore volume is observed to promote single-file translocation of DNA strands at transmembrane biases as low as 10 mV, which opens new avenues for using solid-state nanopores for detection and sequencing of DNA