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DNA nanotechnology defeats drug resistance in cancer cells

Researchers at The Ohio State University are working to develop DNA nanostructures that deliver medicine to drug-resistant cancer cells. These electron microscope images show the structures empty (left) and loaded with the cancer drug daunorubicin (right). The researchers have demonstrated for the first time that such “DNA origami” structures can be used to treat drug-resistant leukemia cells. Images by Randy Patton, courtesy of The Ohio State University.

Scaffolded DNA origami, added to the structural DNA nanotechnology toolkit 10 years ago, is a very powerful technique for folding DNA into complex nanostructures. We’ve cited its use to make make dynamic nanomachines (for example, here), and to make simple nanorobots for potential medical application (here). A recent news release from Ohio State University, written by Pam Frost Gorder, makes clear that even simple atomically precise DNA nanostructures hold great potential for solving a major problem, perhaps the major problem encountered during cancer chemotherapy: the evolution of drug resistance by the cancer. “DNA ‘Trojan horse’ smuggles drugs into resistant cancer cells

Cells mistake DNA casing for food, consume drugs and die

Researchers at The Ohio State University are working on a new way to treat drug-resistant cancer that the ancient Greeks would approve of—only it’s not a Trojan horse, but DNA that hides the invading force.

In this case, the invading force is a common cancer drug.

In laboratory tests, leukemia cells that had become resistant to the drug absorbed it and died when the drug was hidden in a capsule made of folded up DNA.

Previously, other research groups have used the same packaging technique, known as “DNA origami,” to foil drug resistance in solid tumors. This is the first time researchers have shown that the same technique works on drug-resistant leukemia cells.

The researchers have since begun testing the capsule in mice, and hope to move on to human cancer trials within a few years. Their early results appear in the journal Small [abstract].

The study involved a pre-clinical model of acute myeloid leukemia (AML) that has developed resistance against the drug daunorubicin. Specifically, when molecules of daunorubicin enter an AML cell, the cell recognizes them and pumps them back out through openings in the cell wall. It’s a mechanism of resistance that study co-author John Byrd of The Ohio State University Wexner Medical Center compared to sump pumps that draw water from a basement.

He and Carlos Castro, assistant professor of mechanical engineering, lead a collaboration focused on hiding daunorubicin inside a kind of molecular Trojan horse that can bypass the pumps so they can’t eject the drug from the cell.

“Cancer cells have novel ways of resisting drugs, like these pumps, and the exciting part of packaging the drug this way is that we can circumvent those defenses so that the drug accumulates in the cancer cell and causes it to die,” said Byrd, a professor of internal medicine and director of the Division of Hematology. “Potentially, we can also tailor these structures to make them deliver drugs selectively to cancer cells and not to other parts of the body where they can cause side effects.”

“DNA origami nanostructures have a lot of potential for drug delivery, not just for making effective drug delivery vehicles, but enabling new ways to study drug delivery. For instance, we can vary the shape or mechanical stiffness of a structure very precisely and see how that affects entry into cells,” said Castro, director of the Laboratory for Nanoengineering and Biodesign.

In tests, the researchers found that AML cells, which had previously shown resistance to daunorubicin, effectively absorbed drug molecules when they were hidden inside tiny rod-shaped capsules made of DNA. Under the microscope, the researchers tracked the capsules inside the cells with fluorescent tags.

Each capsule measures about 15 nanometers wide and 100 nanometers long—about 100 times smaller than the cancer cells it’s designed to infiltrate. With four hollow, open-ended interior compartments, it looks less like a pill a human would swallow and more like an elongated cinder block.

Postdoctoral researcher Christopher Lucas said that the design maximizes the surface area available to carry the drug. “The way daunorubicin works is it tucks into the cancer cell’s DNA and prevents it from replicating. So we designed a capsule structure that would have lots of accessible DNA base-pairs for it to tuck into. When the capsule breaks down, the drug molecules are freed to flood the cell.”

Castro’s team designed the capsules to be strong and stable, so that they wouldn’t fully disintegrate and release the bulk of the drugs until it was too late for the cell to spit them back out.

And that’s what they saw with a fluorescence microscope—the cells drew the capsules into the organelles that would normally digest them, if they were food. When the capsules broke down, the drugs flooded the cells and caused them to disintegrate. Most cells died within the first 15 hours after consuming the capsules.

This work is the first effort for the engineers in Castro’s lab to develop a medical application for the DNA origami structures they have been building.

Though DNA is stereotypically called the “building blocks of life,” engineers today use natural and synthetic DNA as literal building blocks for mechanical devices. Previously, the Ohio State engineers created tiny hinges [described here] and pistons [described here] of DNA.

As Castro pointed out, DNA is a polymer—albeit a naturally occurring one—and he and his colleagues shape it into tiny devices, tools or containers by exploiting the physical interactions of the bases that make up the polymer chain. They build chains from DNA sequences that will naturally attract and bind with one another in certain ways, so that long the long polymers automatically fold up, or “self-assemble,” into useful shapes.

In the case of this DNA Trojan horse, the researchers used the genome of a common bacteriophage, a virus that infects bacteria, and synthetic strands that were designed to fold up the bacteriophage DNA. Although the folded-up shape performs a function, the DNA itself does not, explained Patrick Halley, an engineering graduate student who is doing this work to earn his master’s degree.

“One of the hardest things to get across when you’re introducing this technology to people is that the DNA capsule doesn’t do anything except hold a shape. It’s just a static, rigid structure that carries things. It doesn’t encode any proteins or do anything else that we normally think of DNA as doing,” Halley said.

In keeping with the idea of DNA origami manufacturing, Castro said he hopes to create a streamlined and economically viable process for building the capsules—and other shapes as well—as part of a modular drug delivery system.

Byrd said the technique should potentially work on most any form of drug-resistant cancer if further work shows it can be effectively translated to animal models, though he stopped short of suggesting that it would work against pathogens such as bacteria, where the mechanisms for drug resistance may be different.

This work points out that scaffolded DNA origami—because of the possibility to precisely control shape, size, and structure—offers important advantages compared to use of simpler, non-atomically precise nanoparticles for targeted drug delivery. Previous research using DNA nanostructures to deliver a similar drug to an adherent line of drug-resistant cancer cells had demonstrated that rod-shaped DNA nanostructures outperformed triangular ones. Further, nanoparticles 100 nm and smaller were known to be efficiently taken up by cancer cells. The authors’ choice of a drug that naturally intercalates with DNA allowed them to choose a simple design comprising four open-ended internal cavities, accessible to the environment and simultaneously providing a large cross section for mechanical stability. Other important features include rapid self-assembly in 5 minutes and stability in cell culture media. Because the DNA nanostructures (nicknamed “Horse” by the authors in analogy to the Trojan horse used by the Mycenaen Greeks to gain entry to Troy during the Trojan War) were endocytosed, thus bypassing the membrane-bound efflux pumps responsible for drug resistance, and digested in the lysosome to release the drug, no specific mechanisms to promote internalization and drug release were needed. But the many options for chemical functionalization, molecular recognition, and simple DNA-based mechanisms embodied in scaffold DNA origami nanostructures provide ample opportunities for developing even more efficacious delivery vehicles if needed for clinical applications.
—James Lewis, PhD

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Foresight Institute appoints Julia Bossmann as new president

Menlo Park, California – Foresight Institute, a leading think-tank for transformative future technologies, announced that Julia Bossmann has joined the organization as president.

“Julia’s breadth of vision for atomically-precise construction, artificial intelligence, and other transformative technologies will bring new energy to Foresight,” says Foresight co-founder Christine Peterson.

Bossmann holds a Masters degree with highest honors in psychology and neuroscience from the University of Dusseldorf and USC. Her professional experience includes scientific research in labs in Germany and in the USA, management consulting at McKinsey & Company, R&D at Bosch Research and Technology, and entrepreneurship at Anticip8 and Synthetic. Bossmann is a GSP alumna at Singularity University and a Global Shaper at the World Economic Forum.

Foresight Institute’s leadership change coincides with the organization’s 30-year anniversary. While keeping its esteemed cross-domain technical workshops going, Bossmann plans to expand engagement with the public and the scientific community. Interim president Steven Burgess continues to serve as COO.

“Foresight Institute has been leading thought on the technologies that transform our world,” says Ms. Bossmann. “With the current rise of world-changing technologies such as nanotechnology, synthetic biology, and artificial intelligence, never before has its mission been as critical and relevant as it is today. The key to our future is to make sure that the necessary beneficial innovations happen soon, and that they happen in a way that benefits society at large.”

About Foresight Institute

Foresight Institute is a leading think tank and public interest organization focused on transformative future technologies. Founded in 1986, its mission is to discover and promote the upsides, and help avoid the downsides, of nanotechnology, AI, biotech, and similar life-changing developments.

For further information, please contact

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Nanotechnologies to advance solar energy utilization

This cell consists of nanostructured arrays of anodes and cathodes, oxidation and reduction catalysts, and a central conductive membrane that allows for ion exchange. Credit: Lewis Research Group, Caltech

A few weeks ago Science published a review written by California Institute of Technology Chemistry Professor Nate Lewis titled “Research opportunities to advance solar energy utilization”. The link to the paper from Lewis’s publication page leads to the full text article on the journal’s web site. Prof. Lewis’s one-page summary of his own review concludes that “Considerable opportunities for cost reduction that can achieve both evolutionary and revolutionary performance improvements are present for all types of solar energy–conversion technologies.” The review enumerates those technologies, focusing on production of solar electricity through photovoltaics, solar thermal, and artificial photosynthesis to produce fuels from sunlight. Two fundamental constraints on solar energy systems are made explicit: (1) the low energy density of sunlight at Earth’s surface, necessitating large areas needed to capture and convert solar power; (2) the intermittent nature of sunlight, requiring affordable technologies for large-scale energy storage.

The first indication of the variety of opportunities available is the table listing a dozen types of photovoltaic materials available to absorb light and produce electricity, characterized according to the nature of the material, the maturity of the technology, production, and efficiency. The materials listed vary greatly in terms of chemical nature, physical form, and how they are currently manufactured. Increasing control over the structure of matter, ultimately approaching atomic precision, will give greater control over ease of manufacture, cost, durability, and efficiency of energy conversion. In addition to the efficiency of energy capture and conversion, factors like single crystal or thin film, or how well the photovoltaic material is encapsulated can affect, weight, ease and cost of installation, keeping toxic materials out of the environment, etc. Opportunities to improve current and near-term systems through research and innovation are accordingly more varied than immediately apparent. Another opportunity is cost reduction through replacement of expensive materials with earth-abundant materials.

As an isolated example of how opportunities for reduced cost and increased convenience lurk in non-obvious places, Lewis gives the example of heavy glass required to protect solar panels from hailstorms, causing solar panels to be inflexible and difficult to install and maintain, plus leading to increased costs for shipping, installation labor, and structural support. Atomically precise materials with much greater strength to weight ratios than conventional materials would likely find application here.

Lewis notes “The cost of persistent grid-scale storage currently far exceeds the levelized cost of solar electricity,” leading to a discussion of costs and research opportunities for improving battery storage.

The section on solar thermal power describes that different types of solar collecting systems working at temperatures ranging from 550 to 1200°C and located in areas of intense sunshine, such as the US southwest or Australia or north Africa, are inspiring research in areas like fabrication of materials that allow operation at higher temperatures.

A final section of the review describes research progress in the direct production of fuels using sunlight, such as artificial photosynthesis based on photoelectrochemical cells.

I will not try to describe the research opportunities involving various forms of nanotechnology pointed to in this review, but as an indication of the breadth of on-going work in this area, the following seven energy-related posts appeared over at Nanowerk during the past two days:

Refined protective layer for the ‘artificial leaf’
A process for providing sensitive semiconductors for solar water splitting (‘artificial leaves’) with an organic, transparent protective layer has been developed by researchers. The extremely thin protective layer made of carbon chains is stable, conductive, and covered with catalyzing nanoparticles of metal oxides. These accelerate the splitting of water when irradiated by light.

Solar water splitting using particulate photocatalyst sheets
Researchers at the University of Tokyo have achieved a solar-to-hydrogen energy conversion efficiency of 1.1% using particulate photocatalyst sheets developed by the research group. The photocatalyst sheet can potentially produce a large quantity of solar hydrogen inexpensively owing to its scalability and low fabrication cost.

Solar cells: Silicon profits from a dose of iron
By rapidly heating silicon wafers covered with thin iron silicide and aluminum films, A*STAR researchers have developed a way to eliminate many of the complicated, time-consuming steps needed to fabricate light harvesting solar cells.

Solid electrolytes open doors to solid-state batteries
Japanese scientists have synthesized two crystal materials that show great promise as solid electrolytes. All-solid-state batteries built using the solid electrolytes exhibit excellent properties, including high power and high energy densities, and could be used in long-distance electric vehicles.

Scientists extend the reach of single crystals
Materials scientists and physicists at Lehigh University have demonstrated a new method of making single crystals that could enable a wider range of materials to be used in microelectronics, solar energy devices and other high-technology applications.

Pumping up energy storage with metal oxides and graphene
Material scientists at Lawrence Livermore National Laboratory have found certain metal oxides increase capacity and improve cycling performance in lithium-ion batteries. The team synthesized and compared the electrochemical performance of three graphene metal oxide nanocomposites and found that two of them greatly improved reversible lithium storage capacity.

Carbon leads the way in clean energy
Groundbreaking research at Griffith University is leading the way in clean energy, with the use of carbon as a way to deliver energy using hydrogen. Professor Xiangdong Yao and his team from Griffith’s Queensland Micro- and Nanotechnology Centre have successfully managed to use the element to produce hydrogen from water as a replacement for the much more costly platinum.

The technologies that are relevant to energy applications range from nanomaterials and nanoscale processing to complex atomically precise arrangements of catalysts. It will be interesting to see how the mix evolves as technology improves and provides more effective solutions at lower costs.
—James Lewis, PhD

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Caltech celebrates ten years of Scaffolded DNA Origami

DNA origami smiley faces, each 1/1000 the width of a human hair, demonstrate that virtually any shape can be folded from DNA. (Scale bar: 100 nanometers) Credit: Paul W.K. Rothemund/Caltech

Since we frequently report progress in structural DNA nanotechnology made possible by the scaffolded DNA origami technique (most recently here), I cannot resist passing on these two news items that I stumbled upon at the Caltech web site, even though it is a day late for the first one, written by Lori Dajose: “Ten Years of DNA Origami“:

On March 16, 2006, Research Professor of Bioengineering, Computing and Mathematical Sciences, and Computation and Neural Systems Paul Rothemund (BS ’94) published a paper in Nature detailing his new method for folding DNA into shapes and patterns on the scale of a few nanometers. This marked a turning point in DNA nanotechnology, enabling precise control over designed molecular structures. Ten years later, the field has grown considerably. On March 14–16, 2016, the Division of Engineering and Applied Science will hold a symposium titled “Ten Years of DNA Origami” to honor Rothemund’s contribution to the field, to survey the spectrum of research it has inspired, and to take a look at what is to come.

“Think about DNA origami as a general-purpose pegboard for organizing nanometer-sized things,” Rothemund says. “Each DNA origami has 200 different attachment points, to which one can attach proteins, or tiny gold balls, or fluorescent molecules, or electrically conductive carbon nanotubes. There is no other way to juxtapose combinations of these elements into complex arrangements, and this is what researchers around the world, from biologists to physicists, are using DNA origami for. Biologists use DNA origami to position different protein enzymes next to each other, so that one enzyme can hand off its products to the next enzyme in a sort of nanoscale assembly line. Others are organizing electronic components in an attempt to make nanocircuits.”

The symposium was organized by Erik Winfree, professor of computer science, computation and neural systems, and bioengineering. “This amazing Caltech invention has had a remarkable impact in molecular nanotechnology research,” he says.

Talks will cover DNA nanotechnology, self-assembly and pattern formation, computational algorithms and software for origami design and analysis, applications in biology and biomedicine, applications in quantum physics, molecular motors and mechanical devices, biophysics and thermodynamics and kinetics, and more. The talks are open to the public, but attendees must first register online.

Continuing the celebration of Rothemund’s achievement “Paul Rothemund – DNA Origami: Folded DNA as a Building Material for Molecular Devices“:

Series:Earnest C. Watson Lecture Series

Wednesday, May 25, 2016, 8:00 pm, Beckman Auditorium

For 3.5 billion years, life has used DNA for information storage, to hold the blueprints of all living things. Over the last 35 years, humans have invented a new use for DNA—as a building material for molecular devices one hundred times smaller than the cell.

This talk will describe how complex DNA structures are designed via computer, synthesized using simple “kitchen chemistry,” and studied with atomic force microscopes a thousand times more powerful than standard light microscopes. We will explore how researchers around the world are applying DNA origami, from cancer-killing nanorobots, to exquisite light sources that will power quantum computers.

As an undergraduate at Caltech from 1990 to 1994, Paul W.K. Rothemund couldn’t decide whether to study biology, chemistry or computer science. Thus, he combined these interests and worked to build computers using DNA. After receiving a Ph.D. from USC in 2001, Rothemund returned to Caltech as a postdoctoral fellow, where he now remains as a research professor of Bioengineering, Computing and Mathematical Sciences, and Computation and Neural Systems. His primary interest is to transform biology into a discipline more like computer science or engineering, using techniques from programming and design. A complementary interest is to bring principles from biology, such as self-assembly, into the manufacture of technology, such as computers.

This is a free event; no tickets or reservations are required.

Paul Rothemund and Erik Winfree were the joint winners of the 2006 Foresight Institute Feynman Prize, in both the Theory and the Experimental categories, the only time that the same team has won both prizes the same year.

This month Nature Materials also commemorated the invention of DNA origami with an editorial titled “Returning to the fold“. After summarizing the enormous contributions this technology has made, the editorial also points out that (1) “a ‘killer application’ has yet to emerge from the many proof-of-concept studies that are currently underway”, and (2) that scale-up and cost considerations of DNA nanomaterials may be significant problems. But they conclude “continued efforts to understand the DNA folding process should lead to further improvements in structure control and product yield.” Certainly substantial problems remain, but from our view point the progress that we have cited provides reason to be optimistic that progress will continue.
—James Lewis, PhD

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Will medical 3D printing advance nanotechnology?

The world’s first 3D-printed vertebrae was implanted in a Chordoma cancer patient. Credit: Ralph Mobbs

About four years ago we speculated that the advent of personal 3D printing for computers might accelerate progress toward atomically precise manufacturing. A few months later we noted the extension of 3D printing from microscale into nanoscale resolution (about 100 nm—still three orders of magnitude from atomic precision, and still using only one material). Since then progress in the technology, often referred to as “additive manufacturing”, has been impressive, especially in medical applications, even to the point of progressing toward 3D printing of tissues and organs. One especially striking example of this progress is described by Steve Smith at Medical Daily “World’s First 3D-Printed Vertebrae Saves Man With Chordoma Cancer From Becoming A Quadriplegic“:

What a time it is for 3D printing in health care. Over the past year alone, doctors have successfully separated conjoined twins, given a cancer patient a titanium rib cage, and created muscle, bone, and ears from 3D-printing materials. This list continues to grow; in December 2015, a man in his 60s received the first 3D-printed vertebrae. Without it, he would have become fully paralyzed. …

Dr. Ralph Mobbs, an Australian neurosurgeon, took charge of treating the tumor, and relied on 3D printers and Australian medical device manufacturer Anatomics. In a combined effort, they were able to create exact replicas of the two vertebrae made out of titanium, according to Mashable. Anatomics even created entire replicas of Josevski’s spinal anatomy for doctors to conduct practice runs before the surgery.

“3D printing of body parts is the next phase of individualized health care,” Mobbs told The ABC Australia. “To restore bones, joints, [and] organs with this type of technology really is super exciting. [H]ere is our opportunity to really take it out there and to keep pushing the boundaries on the whole 3D-printed body part business.” …

This amazing medical accomplishment, and the others cited by Smith in the first paragraph quoted above, neither advances the resolution of 3D printing nor greatly increases the complexity of the product printed. (In the case of 3D-printed tissues the complexity is provided by seeding the structure with intact and functional human cells provided by biotechnology.) But one can hope that a rapidly growing base of important applications of 3D printing will eventually lead to improvements in resolution and complexity. Perhaps one could imagine an intermediate stage on the way to nanofactories for general purpose high throughput atomically precise manufacturing in which nanoscale atomically precise devices are used as inks in 3D printers to build macroscale products composed in large part of complex arrays of atomically precise machinery.
—James Lewis, PhD

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Crowd-sourced RNA structure design uncovers new insights

An Eterna interface used to enable players to select nucleotides when assessing the difficulty of RNA secondary structure design puzzles. Credit: Andersen-Lee et al., J. Mol. Biol. 2016.

Two years ago we commented on the success of “citizen scientists” playing an online game in outperforming the best available computerized design algorithms in designing RNA molecules to fold into predetermined structures. A news article appearing last month in Science, written by John Bohannon and discussing an Open Access paper (“Principles for Predicting RNA Secondary Structure Design Difficulty“) just published in the Journal of Molecular Biology, makes clear that the progress has continued “For RNA paper based on a computer game, authorship creates an identity crisis“:

… Today’s paper shows how far the effort has come. Among the game’s thousands of RNA design “puzzles,” there seem to be a small set that are particularly difficult. Among the most challenging structural features to figure out is symmetry, where an RNA strand folds into two or more identically shaped loops. The Eterna game includes an interface for players to propose hypotheses about how particular RNA structures will or will not fold into particular shapes. Those were distilled into a set of “designability” rules. The question was: Do only human designers struggle with thorny design problems, or do computer simulations tussle too?

The answer is that the computers struggled just as much as the people. Researchers report that three of the best existing computer algorithms, running on a supercomputer at Stanford, struggled to solve the very same RNA design problems as the humans. The result shows that the human “designability” rules do indeed correspond to problems that are hard not just for human brains but also for computers, the team reports today in the Journal of Molecular Biology. In fact, the hardest puzzles that could be solved by experienced Eterna players were unsolvable by the computer even after days of crunching. And to help improve the algorithms, computer scientists now have a set of benchmarks—the Eterna100—to gauge the design difficulty of RNA structures.

“This paper is a significant contribution,” says Jane Richardson, a biophysicist at Duke University in Durham, North Carolina. “The new design principles seem very understandable now that they’ve been described, and attention to them should indeed help automated design.” She notes that the game itself has become a useful tool for scientists. “One of our lab’s former students is an avid Eterna player who has also gotten insights from it for his own current research projects.”…

It is an interesting observation in the developing story of human vs. machine that despite computer dominance in games like Chess, and more recently Go, that in a domain in which the rules are still not all known, large groups of amateurs playing a game can outperform computers and teach new design rules to professional scientists. No doubt the computers will eventually triumph, but for now a long road with many opportunities for collaboration seems to lie ahead.
—James Lewis, PhD

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Tightly-fitted DNA parts form dynamic nanomachine

Three multilayer DNA components make up this rotary mechanism. The parts join together with a tight fit and leave just 2 nanometers of play around the axle, allowing the arm to swing but not wobble. Credit: Dietz Lab/TUM

Since its invention in 2006 by Paul Rothemund, scaffolded DNA origami has been used to build increasingly complex 2D and 3D structures, including organizing nanoscale functional objects in 3D space. The past few years have seen increasing progress in extending the usefulness of this very versatile method of constructing nanostructures from DNA. Three years ago we noted the report from Hendrik Dietz’s group of improvements in technique that resulted in faster folding, gave higher yields, and provided structures assembled with sub-nanometer positional precision. Higher yields of more precisely assembled DNA nanostructures opened the way to using DNA origami to build more dynamic structures evincing mechanical movements. A year ago we cited the accomplishments of a former postdoctoral fellow in Dietz’s group, Carlos Castro, in achieving well-defined programmed motions with DNA nanostructures, thus beginning to fabricate parts for machine designs based upon the way that macroscopic machines work. We reported additional progress from Castro and his colleagues here and here. A few months later we cited a report from Dietz’s lab on DNA components with complementary shapes that self assemble into nanoscale machinery. Finally, last December we noted the accomplishment of the Dietz group in using DNA nanotechnology to position molecules with atomic precision. A new result from the Dietz group recently published in the open access journal Science Advances and also available from Dietz’s publications page demonstrates (quoting from the abstract) “a nanoscale rotary mechanism that reproduces some of the dynamic properties of biological rotary motors in the absence of an energy source, such as random walks on a circle with dwells at docking sites.” Both this and the previous advance of positioning molecules with atomic precision are described in a press release from the Technical University of Munich “Nanoscale rotor and gripper push DNA origami to new limits“:

Dietz lab’s latest DNA nanomachines demonstrate dynamics and precision

Scientists at the Technical University of Munich (TUM) have built two new nanoscale machines with moving parts, using DNA as a programmable, self-assembling construction material. In the journal Science Advances, they describe a rotor mechanism formed from interlocking 3-D DNA components. Another recent paper, in Nature Nanotechnology, reported a hinged molecular manipulator, also made from DNA. These are just the latest steps in a campaign to transform so-called “DNA origami” into an industrially useful, commercially viable technology.

Inspired by nature’s nanomachines – such as the enzyme ATP synthase and the motor-driven flagella of bacteria – physicists in Prof. Hendrik Dietz’s lab at TUM keep expanding their own design and construction repertoire. They have systematically developed rules and procedures for creating self-assembled DNA origami structures with ever greater flexibility and control. Moving from DNA basepair matching to shape-complementary building techniques – with a variety of interlocking “bricks” – the researchers’ toolkit has advanced steadily in the direction of higher-level programming and modular assembly.

In step with this progress, they have honed methods needed to verify, for example, that a particular soup of nanoparticles really is packed with copies of whatever they designed: whether a switchable gear, an artificial membrane channel, an arbitrarily complicated test object, a “nanobook” that opens and closes, or a robot figure with movable arms.

Two new additions to the zoo

The latest additions to the lab’s zoo of DNA origami objects, two tiny 3-D machines with moving parts, demonstrate new capabilities. (See video at EurekAlert, 1’23″.)

Doctoral candidates Philip Ketterer and Elena Willner collaborated with Dietz in building a rotory [sic] mechanism from three multilayer DNA building blocks: a rotor unit, with a body roughly 32 nanometers long and a longer, lever-like extension; and two clamp elements that “click” together to form an axle bearing. The parts join with a tight fit and leave just 2 nanometers of play around the axle, allowing the rotor to swing but not to wobble. In one variant, the arm will rotate freely between random stopping points; in another, it will dwell in specified positions the researchers call docking sites. To date, this is the most complex rotary structure realized using DNA origami techniques.

To be clear, the rotor has no motor: It is propelled by Brownian motion, random movement of particles in solution. By demonstrating the feasibility of building such a machine, however, the researchers open the way for active devices under chemical or thermal power and control. “It’s like having built an engine,” Dietz says. “Now spark plugs and combustible fuel are the next items on the to-do list.”

In a separate project, Dietz and doctoral candidate Jonas Funke created a hinged machine on a scale suitable for manipulating individual molecules with atomic precision. The angle between the gripper’s two main structural elements can be controlled with DNA helices. Experiments with this DNA origami positioning device showed that it could be capable of precisely placing molecules, adjusting the distance between them in steps as small as the radius of a hydrogen atom. This work is significant in that it bucks a trend in the field toward building larger DNA origami devices without necessarily pushing the limits of precision. In addition, the results hint at one of the ways DNA nanomachines might someday be useful to control chemical reactions.

“On the one hand,” Dietz says, “we now really trust the placement precision within our structures – because we have actually placed two molecules and controlled their distance with atomic precision. On the other hand we have now a prototype rotary mechanism. Based on our measurements, its mobility is such that it could do up to 50,000 rpm if every rotary step it took would go only in one direction. In the next generation of devices, we will use the placement precision to couple chemical reactions to the movements of a rotor. This has the potential to result in a motor. This device, then, could be used for all kinds of purposes, such as actively propelling nanoscale drug-delivery vehicles, pumping and separating molecules across barriers, or packaging molecules into cargo compartments.”

Work with dynamic DNA nanostructures is clearly heading in a very interesting direction. Providing a motor to power a mechanism such as the one demonstrated in this paper, instead of relying on Brownian motion, would be a very important next step. Perhaps a few more generations of improved devices will produce nanomachines that will be able to make or break specific chemical bonds? Increasingly complex systems of such devices could perhaps provide a path to productive nanosystems and eventually to atomically precise manufacturing.
—James Lewis, PhD

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DNA nanotechnology provides new ways to arrange nanoparticles into crystal lattices

Several years ago we pointed to work from the collaboration of two Foresight Feynman Prize winners (Chad A. Mirkin, 2002, Experimental and George C. Schatz, 2008, Theoretical) that advanced the concept of using DNA to link together nanoparticles in specific 3D configurations: “Using DNA as bonds to build new materials from nanoparticles“. A news article written by Robert F. Service in a recent issue of ScienceDNA makes lifeless materials shapeshift” describes another major advance from Mirkin’s group, taking their 2011 advance to the next level:

Researchers have engineered tiny gold particles that can assemble into a variety of crystalline structures simply by adding a bit of DNA to the solution that surrounds them. Down the road, such reprogrammable particles could be used to make materials that reshape themselves in response to light, or to create novel catalysts that reshape themselves as reactions proceed.

“This paper is very exciting,” says Sharon Glotzer, a chemical engineer at the University of Michigan, Ann Arbor, who calls it “a step towards pluripotent matter.” David Ginger, a chemist at the University of Washington, Seattle, agrees: “This is a proof of concept of something that has been a nanoparticle dream.” Neither Glotzer nor Ginger has ties to the current research. …

The 2011 paper was titled “Nanoparticle Superlattice Engineering with DNA” (abstract, PDF courtesy of Mirkin lab). The current advance: “Transmutable nanoparticles with reconfigurable surface ligands” (abstract). Service explains the nature of this new advance as no longer needing to re-engineer the DNA linkers each time a material with a new crystal orientation is needed. He describes that it works by coating each nanoparticle, not with single DNA strands, but with a DNA hairpin that can be opened or closed by adding additional short DNA strands to the solution, thus changing the configuration of other nanoparticles with which it will bind.

The researchers also added more control over what crystals formed by changing the length of the DNA hairpins, the concentration at which they were assembled around the particles, and the concentration of different types of particles. They report in today’s issue of Science that they used these different knobs to create 10 different crystals. But according to Mirkin, his team already has the ability to cause particles to assemble into more than 500 different crystal forms. “This gives us the ability to make materials by design,” he says. …

The same issue of Science features another approach to engineering superlattices using DNA nanotechnology: “Diamond family of nanoparticle superlattices” [abstract].

Schematic illustration of the experimental strategy: Double stranded DNA bundles (gray) form tetrahedral cages. Single stranded DNA strands on the edges (green) and vertices (red) match up with complementary strands on gold nanoparticles. This results in a single gold particle being trapped inside each tetrahedral cage, and the cages binding together by tethered gold nanoparticles at each vertex. The result is a crystalline nanoparticle lattice that mimics the long-range order of crystalline diamond. The images below the schematic are (left to right): a reconstructed cryo-EM density map of the tetrahedron, a caged particle shown in a negative-staining TEM image, and a diamond superlattice shown at high magnification with cryo-STEM. Credit: Brookhaven National Laboratory

The research is described in this Brookhaven National Laboratory news release “Scientists Guide Gold Nanoparticles to Form ‘Diamond’ Superlattices“:

DNA scaffolds cage and coax nanoparticles into position to form crystalline arrangements that mimic the atomic structure of diamond

Using bundled strands of DNA to build Tinkertoy-like tetrahedral cages, scientists at the U.S. Department of Energy’s Brookhaven National Laboratory have devised a way to trap and arrange nanoparticles in a way that mimics the crystalline structure of diamond. The achievement of this complex yet elegant arrangement … may open a path to new materials that take advantage of the optical and mechanical properties of this crystalline structure for applications such as optical transistors, color-changing materials, and lightweight yet tough materials.

“We solved a 25-year challenge in building diamond lattices in a rational way via self-assembly,” said Oleg Gang, a physicist who led this research at the Center for Functional Nanomaterials (CFN) at Brookhaven Lab in collaboration with scientists from Stony Brook University, Wesleyan University, and Nagoya University in Japan.

The scientists employed a technique developed by Gang that uses fabricated DNA as a building material to organize nanoparticles into 3D spatial arrangements. They used ropelike bundles of double-helix DNA to create rigid, three-dimensional frames, and added dangling bits of single-stranded DNA to bind particles coated with complementary DNA strands.

“We’re using precisely shaped DNA constructs made as a scaffold and single-stranded DNA tethers as a programmable glue that matches up particles according to the pairing mechanism of the genetic code—A binds with T, G binds with C,” said Wenyan Liu of the CFN, the lead author on the paper. “These molecular constructs are building blocks for creating crystalline lattices made of nanoparticles.”

The difficulty of diamond

As Liu explained, “Building diamond superlattices from nano- and micro-scale particles by means of self-assembly has proven remarkably difficult. It challenges our ability to manipulate matter on small scales.”

The reasons for this difficulty include structural features such as a low packing fraction—meaning that in a diamond lattice, in contrast to many other crystalline structures, particles occupy only a small part of the lattice volume—and strong sensitivity to the way bonds between particles are oriented. “Everything must fit together in just such a way without any shift or rotation of the particles’ positions,” Gang said. “Since the diamond structure is very open, many things can go wrong, leading to disorder.”

“Even to build such structures one-by-one would be challenging,” Liu added, “and we needed to do so by self-assembly because there is no way to manipulate billions of nanoparticles one–by–one.”

Gang’s previous success using DNA to construct a wide range of nanoparticle arrays suggested that a DNA-based approach might work in this instance.

DNA guides assembly

The team first used the ropelike DNA bundles to build tetrahedral “cages”—a 3D object with four triangular faces. They added single-stranded DNA tethers pointing toward the interior of the cages using T,G,C,A sequences that matched up with complementary tethers attached to gold nanoparticles. When mixed in solution, the complementary tethers paired up to “trap” one gold nanoparticle inside each tetrahedron cage.

The arrangement of gold nanoparticles outside the cages was guided by a different set of DNA tethers attached at the vertices of the tetrahedrons. Each set of vertices bound with complementary DNA tethers attached to a second set of gold nanoparticles.

When mixed and annealed, the tetrahedral arrays formed superlattices with long-range order where the positions of the gold nanoparticles mimics the arrangement of carbon atoms in a lattice of diamond, but at a scale about 100 times larger.

“Although this assembly scenario might seem hopelessly unconstrained, we demonstrate experimentally that our approach leads to the desired diamond lattice, drastically streamlining the assembly of such a complex structure,” Gang said.

The proof is in the images. The scientists used cryogenic transmission electron microscopy (cryo-TEM) to verify the formation of tetrahedral frames by reconstructing their 3D shape from multiple images. Then they used in-situ small-angle x-ray scattering (SAXS) at the National Synchrotron Light Source (NSLS), and cryo scanning transmission electron microscopy (cryo-STEM) at the CFN, to image the arrays of nanoparticles in the fully constructed lattice.

“Our approach relies on the self-organization of the triangularly shaped blunt vertices of the tetrahedra (so called ‘footprints’) on isotropic spherical particles. Those triangular footprints bind to spherical particles coated with complementary DNA, which allows the particles to coordinate their arrangement in space relative to one another. However, the footprints can arrange themselves in a variety of patterns on a sphere. It turns that one particular placement is more favorable, and it corresponds to the unique 3D placement of particles that locks the diamond lattice,” Gang said.

The team supported their interpretation of the experimental results using theoretical modeling that provided insight about the main factors driving the successful formation of diamond lattices.

Sparkling implications

“This work brings to the nanoscale the crystallographic complexity seen in atomic systems,” said Gang, who noted that the method can readily be expanded to organize particles of different material compositions. The group has demonstrated previously that DNA-assembly methods can be applied to optical, magnetic, and catalytic nanoparticles as well, and will likely yield the long-sought novel optical and mechanical materials scientists have envisioned.

“We’ve demonstrated a new paradigm for creating complex 3D-ordered structures via self-assembly. If you can build this challenging lattice, the thinking is you can build potentially a variety of desired lattices,” he said.

Additional news coverage of these papers is provided by Matthew Gunther at Chemistry World “DNA-coated nanoparticles take crystal engineering into the diamond league“. With the technology for arranging nanoparticles into predetermined lattices continuing to improve, it will be interesting to see what kinds of materials and device arrays are produced.
—James Lewis, PhD

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Improving crystallographic resolution through using less perfect crystals

The analysis of the Bragg peaks alone (top) reveals far less detail than the analysis of the continuous diffraction pattern (bottom). Magnifying glasses show real data. Credit: DESY, Eberhard Reimann

Sometimes atomic resolution imaging can be a big help in understanding how molecular machinery works. A news release from the Biodesign Institute of Arizona State University suggests that this may become easier to do “New method opens crystal clear views of biomolecules – Fundamental discovery triggers paradigm shift in crystallography“:

A scientific breakthrough gives researchers access to the blueprint of thousands of molecules of great relevance to medicine, energy and biology. In a new study, researchers from Arizona State University (ASU), Deutsches Elektronen-Synchrotron (DESY) and Stanford Linear Accelerator Laboratory (SLAC) describe a simple way to determine the 3-dimensional structures of proteins and other molecules, many of which are inaccessible by existing methods.

The research findings appear in the current issue of the journal Nature [abstract].

Imaging the molecular building blocks of living things at the atomic scale is tricky. Often, the most difficult step is getting such molecules to form high-quality crystals needed for X-ray imaging. This international team describes a new method that can produce sharp images relying on crystals with very small imperfections, using the world’s brightest X-ray source at the Department of Energy’s SLAC National Accelerator Laboratory.

“Once the full potential of the new method is understood, it could turn out to be one of the biggest advances since the birth of crystallography,” said Mike Dunne, director of the Linac Coherent Light Source (LCLS) X-ray laser, a DOE Office of Science User Facility.

The structures of biomolecules reveal their modes of action and provide insights into the workings of the machinery of life. Unlocking the molecular structure of particular proteins, for example, can provide the basis for developing tailor-made drugs against numerous diseases or advancing clean energy technologies with the efficiency of nature and the stability of engineered systems.

The chosen target of the new study is a critical enzyme known as photosystem II. Petra Fromme, co-author of the new study and director of the Biodesign Institute’s Center for Applied Structural Discovery (CASD) at ASU highlights the importance of this molecule:

“Life on earth relies directly on photosynthesis—the natural process of converting light energy to chemical energy in plants and algae. The critical first step is carried out by the photosystem II protein complex, which uses sulight [sic] to split abundant sources of water (H2O) into hydrogen, electrons and all the oxygen that sustains our planet,” she says.

In 2011,the team of researchers at ASU, (including Fromme and John Spence, professor of Physics, along with their collaborators), pioneered a technique called serial femtosecond crystallography. Here, structure determination is based on imaging of thousands of nanocrystals, so small that they can not even be seen under a microscope. The nanocrystals are hit “on-the-fly” by X-ray pulses that are so strong that they destroy any solid material but so short that a diffraction image is obtained before the destruction occurs.

The method was further developed in 2014, and was used to produce the first snapshot images of the water splitting process in photosysnthesis [sic]. It wasn’t clear however if the researchers would ever be able to see the dynamics of the process at atomic scale. Such fine detail coupled with dynamic imaging is essential for understanding the mechanisms of photosynthesis that split water and produce oxygen with visible light and earth abundant metals. Traditional X-ray crystallography methods could achive [sic] high resolution but suffer from damage from the X-rays and are not suitable to see dynamics of the process.

“This new method allows us to significantly improve the resolution of the images we took of photosystem II and gives us a new tool for capturing the dyanmics [sic] of some of life’s most important processes. When we understand how photosynthesis works at the chemical level we can use this as a blueprint for developing clean and renewable energy technologies,” Fromme adds.

“The best crystals are crystals with small imperfections”

More than 100 years ago, Australian-born British physicist William Lawrence Bragg found a way to use X-rays to probe the interior of crystals, which consist of regular arrays of atoms or molecules. This discovery launched the field of X-ray crystallography, one of the most important techniques for analyzing the structures of materials, chemical processes and biological molecules.

When X-rays pass through the crystal, they scatter off the protein molecules and form a pattern on a detector. This diffraction pattern is dominated by bright spots known as Bragg peaks, which researchers use to reconstruct the atomic structure of the molecules.

A perfectly ordered crystal would produce nothing but Bragg peaks. However, disorder limits the number of detectable peaks and therefore the resolution of the molecular image that can be obtained from the peaks alone.

The small displacements lead to “termination of the Bragg peaks”, but allow scientists to “see” the scattering from the sum of the individual molecules in the crystals. These tiny molecular displacements from a perfectly ordered crystal arrangement produce gently rippling patterns between and beyond the sharp Bragg peaks. While these patterns, known as “continuous diffraction,” have been actively studied, they had not been considered capable of producing high-resolution molecular images.

“We’ve now demonstrated that we can actually use the continuous diffraction of imperfect crystals to obtain better molecular images than with Bragg peaks alone,” said Kartik Ayyer, the study’s first author from the Center for Free-Electron Laser Science (CFEL) at the German research center DESY.

“Phased in”

Even when good diffraction is available the structure of a complex molecule is still extremely difficulty to determine. Without knowing the phase – the lag of the crests of one diffracted wave to another – it is not possible to compute an image of the molecule from the measured diffraction pattern.

To solve the tricky phase puzzle, more information must be known than just the intensity of the measured Bragg spots. Sometimes this information can be derived by X-[ray] analysis of crystals of chemically modified molecules, or inferred from the structure of a closely-related molecule, but these approaches are time consuming and can bias the results. Bragg peaks are strong, since they arise from constructive interference (like a crowd clapping in unison), but they are sparse and of limited content. The continuous diffraction used in the new method fills in the gaps in and beyond the Bragg peaks, giving vastly more information, which can be used to directly obtain the phase and improve the resolution.

ASU physicist and study co-author John Spence elaborates:

“In 1913, Paul Debye showed that Bragg diffraction from vibrating atoms in a crystal scatter between the Bragg spots. Henry Chapman (XFEL scientist with DESY and corresponding author) was able to show how, in a similar way, the non-Bragg scattering from LCLS X-ray snapshots from imperfect crystals can be used to provide a resolution-enhanced image of the protein molecule, and also solve the phase problem.

This simple concept leads to a paradigm shift in crystallography — the most ordered crystals are no longer the best to analyze. Imperfect crystals arise because some molecules do not align perfectly with majority of the molecules within the crystal [see Figure A in the original article], which creates a continuous diffraction pattern similar to those predicted for single molecules. Single molecule diffraction is the Holy Grail of modern X-ray science because it eliminates the crystallization process altogether, but has never been possible because the signals from individual molecules are far too weak.

“For the first time we have access to single molecule diffraction – we have never had this in crystallography before,” Chapman says. The technique elegantly weds X-ray diffraction of crystals and X-ray imaging of single particles, providing the best of both worlds.

The technique could provide a stepping-stone toward detailed imaging of single particles, including the large number of biological specimens that cannot easily be crystallized[.]

Fromme states that the new technique also opens the field for time resolved movies of biomolecules at work. “While perfect crystalline order restricts the movement of molecules, the new method allows us to study dynamic processes in single crystals of biomolecules.”

No doubt the earliest applications of this technology will focus on biomolecules and biomedical applications, as described above. As experience with the technology accumulates, however, it would be interesting to see it applied to dynamic artificial molecular machinery, such as this and these and these and this example of dynamic structural DNA nanotechnology.
—James Lewis, PhD

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DNA nanotechnology cages localize and optimize enzymatic reactions

Individual enzymes (orange and green) are first attached to half-cage structures. Half cages are then assembled into full cages, where reactants are brought into close proximity. Credit: Jason Drees for the Biodesign Institute, Arizona State University

About 7 years ago we pointed to two research reports as “evidence of the growing capability of DNA scaffolds to support complex and interactive functions” (Advancing nanotechnology by organizing functional components on addressable DNA scaffolds). One of the research groups featured in that post has just published another paper using DNA cages to hold enzymes and their substrates in the proper position to make reactions more efficient. A hat tip to nanowerk for reprinting this Arizona State University news release “Chemical Cages: new technique advances synthetic biology“:

Living systems rely on a dizzying variety of chemical reactions essential to development and survival. Most of these involve a specialized class of protein molecules—the enzymes.

In a new study, Hao Yan, director of the Center for Molecular Design and Biomimetics at Arizona State University’s Biodesign Institute presents a clever means of localizing and confining enzymes and the substrate molecules they bind with, speeding up reactions essential for life processes.

The research, which appears in the current issue of the journal Nature Communications [OPEN], could have far-reaching applications in fields ranging from improving industrial efficiencies to pioneering new medical diagnostics, guiding targeted drug delivery and producing smart materials. The work also promises to shed new light on particulars of cellular organization and metabolism.

The technique involves the design of specialized, nanometer-scale cages, which self-assemble from lengths of DNA. The cages hold enzyme and substrate in close proximity, considerably accelerating the rate of reactions and shielding them from degradation.

“We have been designing programmable DNA nanostructures with increasing complexity for many years, and it is now time to ask what can we do with these structures,” Yan says. “There are numerous other applications from this emerging technology. Through our interdisciplinary collaborative effort, we here describe the use of designer DNA nanocages to compartmentalize enzymatic reactions in a confined environment. Drawing inspiration from Nature, we have uncovered interesting properties, some unexpected.”

Zhao Zhao, a researcher in the Center for Molecular Design and Biomimetics was the lead author of the paper, which was co-authored with researchers from ASU as well as the Department of Chemistry, Rutgers and the Department of Chemistry, Single Molecule Analysis Group, University of Michigan.

Enzyme world

As chemical activators for virtually every reaction in the body, enzymes are key participants in the normal activity of cells, tissues, fluids, and organs. Hundreds of thousands of metabolic enzymes are present in the human body, involved in diverse activities including DNA copying and repair and the transformation of glucose into useable energy. Elsewhere, some 22 digestive enzymes break down carbohydrates (amylases), fats (lipases ) and sugars (disaccharides), while so-called protease enzymes digest proteins.

Enzymes tend to be highly specific, not only in the useful functions they perform, but the precise substrates with which they will work. Substrate molecules of exactly the right size and shape bind with their appropriate enzymes as the correct key fits into the ridges and grooves of a lock.

Substrates latch onto enzyme molecules at a particular region known as the active site. Once enzyme and substrate have combined, a chemical product is formed and then released, returning the enzyme to its original configuration where it is ready to operate on a new molecule of substrate.

In order for such reactions to take place in an efficient manner, Nature has devised methods of compartmentalization, forming natural reactor sites where enzyme-substrate reactions unfold. The cell itself is such a compartment, as are various membrane-bound organelles found in eukaryotes, (cells containing a nucleus), including mitochondria, lysosomes and peroxisomes.

Compartmentalization of reactants helps to overcome a variety of challenges, bringing binding chemicals into cozy proximity, isolating enzyme-substrate complexes from competing reaction chemicals, improving the yield of product molecules produced and reducing the toxicity various intermediary chemicals can sometimes cause.

In order to induce or catalyze chemical reactions for a variety of purposes, synthetic biologists have copied a page from Nature’s recipe book, designing artificial compartments fabricated from proteins, lipids or the nucleic acids found in DNA, (as in the current study).

Close encounters

Yan and his colleagues designed their synthetic reactors to house enzymes and their substrates, allowing chemical conversions to take place in a controlled environment. Each minute structure, measuring just 54 nanometers across, is something like a Faberge egg whose separate halves fit together to encapsulate their chemical contents. (A nanometer is one billionth of a meter or roughly 80,000 times smaller than the width of a human hair.)

Using the base pairing properties of DNA’s four nucleotides, labeled A, T, C and G allows nanoscale architects like Yan to construct myriad forms in two- and three-dimensions. In the new study, DNA nanocages were used to encapsulate metabolic enzymes with high assembly yield and fine-tuned control over reactants and products.

The construction of the nanocages takes place in two steps. First, individual enzymes are attached into open half-cage structures. Then, the half-cages are fitted together into a full, closed nanocage. To create the half-cages, a technique known as DNA origami is used. Lengths of viral DNA are prepared to self-assemble into a honeycomb lattice, with A nucleotides pairing with C and T with G [ sic ].

The open-sided half cages of the DNA nanocages allow the access of large protein molecules into the nanocage’s internal cavity. The two half-cages are fitted together with the aid of short bridge DNA strands that bind with complementary DNA sequences extending from the edges of either half-cage, (see accompanying animation). The small gaps on each of the top and bottom surfaces of the DNA nanocage allow the diffusion of small molecules across the DNA walls.

Probing the nanoscale

To examine the resulting structures, Transmission Electron Microscopy was used, along with gel electrophoresis and single molecule fluorescence experiments which demonstrated that close to 100 percent of the DNA segments properly formed half-cage structures and more than 90 percent formed full cages.

The study examined six different enzymes, ranging in size from the smallest, which measured ~44kD (kilodaltons) to the largest, ~ 450 kD. All six enzymes were successfully encapsulated in nanocages, though the yields varied according to enzyme size. The largest enzyme examined, known as β-galactosidase, showed the lowest yield of 64 percent.

Next, the activity of enzyme-substrate pairs was evaluated. In addition to bringing the enzyme-substrate pair into closer binding proximity, encapsulation in the nanocage is also believed to facilitate activity through the unique electrical charge density conditions within the nanocage.

Subsequent experiments demonstrated that most of the effect on enzyme-substrate activity in nanocages is due to the unique charge environment within nanocages, rather than enzyme-substrate proximity. The authors suggest that encapsulated enzymes exhibit higher activity within densely packed DNA cages as a result of the highly ordered, hydrogen-bonded water environment surrounding them.

An evaluation of enzyme activity showed a 4- to 10-fold increase for enzymes encapsulated in nanocages, compared with the activity of free enzymes. Enzyme turnover rate—defined as the maximum number of chemical conversions of substrate molecules per second—was inversely correlated with the size of encapsulated enzymes, with the smallest enzyme yielding the highest turnover.

Future cages

The DNA cages demonstrated their resiliency during the experiments, retaining their structural form throughout the enzymatic reactions. They also protected encapsulated enzymes from deactivation due to digestive chemicals, while permitting the uninterrupted diffusion of small-molecule substrates and reaction products through the nanopores of the DNA cage.

Encapsulation in nanocages was shown to increase the fraction of active enzyme molecules and their individual turnover numbers. The method thus provides a new molecular tool to modify the local environment surrounding enzymes and their substrates, opening the door to new applications in smart materials and biomedical applications. Among the latter are futuristic, programmable cages that could be used as nanoscale delivery mechanisms for a wide range of therapeutic agents.

This application of DNA nanotechnology focused on optimizing the confinement of enzymes, ultimately for use in biomedical applications, or for optimizing biotechnology, or producing smart materials. A step further along the track to higher complexity would be to combine a DNA scaffold with a DNA arm to mimic an enzyme cascade. Two years ago we pointed to work, also led by Prof. Yan, in which a DNA arm was added to an open DNA scaffold to pass a substrate from one enzyme to the next in a cascade. It will be interesting to watch their progress in this area, using increasingly complex DNA nanostructures to optimize reaction and sequences of reactions.
—James Lewis, PhD

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