New software aids design of 3-D DNA structures

Recent demonstrations of the ability to build complex 3-D shapes from DNA (this post and previous work by William Shih and collaborators published in Science August 2009 abstract) create demand for an easier way to design complex shapes from folded DNA strands. Now new software facilitates designing three dimensional shapes using scaffolded DNA origami. Physorg.com points to this written by Anne Trafton, MIT News Office “Origami: Not just for paper anymore“:

… A major hurdle to [designing complex curved and bent structures from a folded DNA strand] has been automation of the design process. Now a team at MIT, led by biological engineer Mark Bathe, has developed software that makes it easier to predict the three-dimensional shape that will result from a given DNA template. While the software doesn’t fully automate the design process, it makes it considerably easier for designers to create complex 3-D structures, controlling their flexibility and potentially their folding stability.

“We ultimately seek a design tool where you can start with a picture of the complex three-dimensional shape of interest, and the algorithm searches for optimal sequence combinations,” says Bathe, the Samuel A. Goldblith Assistant Professor of Applied Biology. “In order to make this technology for nanoassembly available to the broader community — including biologists, chemists, and materials scientists without expertise in the DNA origami technique — the computational tool needs to be fully automated, with a minimum of human input or intervention.”

Bathe and his colleagues described their new software in the Feb. 25 issue of Nature Methods. In that paper [abstract], they also provide a primer on creating DNA origami with collaborator Hendrik Dietz at the Technische Universitaet Muenchen. “One bottleneck for making the technology more broadly useful is that only a small group of specialized researchers are trained in scaffolded DNA origami design,” Bathe says.

… “DNA is in many ways better suited to self-assembly than proteins, whose physical properties are both difficult to control and sensitive to their environment,” Bathe says.

Bathe’s new software program interfaces with a software program from Shih’s lab called caDNAno, which allows users to manually create scaffolded DNA origami from a two-dimensional layout. The new program, dubbed CanDo, takes caDNAno’s 2-D blueprint and predicts the ultimate 3-D shape of the design. This resulting shape is often unintuitive, Bathe says, because DNA is a flexible object that twists, bends and stretches as it folds to form a complex 3-D shape.

A PDF of the paper has been made available by the Dietz Lab at TU Munich here. An accompanying editorial “Into the fold” [abstract, full text requires free registration] gives a one-page overview of DNA origami and its potential applications. It points to the above paper and a paper [abstract] from Shih and his colleagues on purifying DNA nanostructures with improved yield of intact structures, and it discusses what is yet needed for DNA origami to reach its full potential: other and longer DNA single strands to use as scaffolds, improved methods to chemically conjugate functional groups to specific DNA sites, and a “top-down design solution” to fully automate design. Now we know what developments to watch for!

DNA nanotechnology builds 3D forms with complex curves (includes video)

Miniature architectural forms – some no larger than viruses – have been constructed through a revolutionary technique known as DNA origami. Now, Hao Yan, Yan Liu and their colleagues at ASU’s Biodesign Institute have expanded the capability of this method to construct arbitrary, two- and three-dimensional shapes, mimicking those commonly found in nature.

Such diminutive forms may ultimately find their way into a wide array of devices, from ultra-tiny computing components to nanomedical sentries used to target and destroy aberrant cells or deliver therapeutics at the cellular or even molecular level.

In today’s issue of Science [abstract], the Yan group describes an approach that capitalizes on (and extends) the architectural potential of DNA. The new method is an important step in the direction of building nanoscale structures with complex curvature – a feat that has eluded conventional DNA origami methods. …

“Our goal is to develop design principles that will allow researchers to model arbitrary 3-D shapes with control over the degree of surface curvature. In an escape from a rigid lattice model, our versatile strategy begins by defining the desired surface features of a target object with the scaffold, followed by manipulation of DNA conformation and shaping of crossover networks to achieve the design,” Liu said.

To achive this idea, Yan’s graduate student Dongran Han began by making simple 2-D concentric ring structures, each ring formed from a DNA double helix. The concentric rings are bound together by means of strategically placed crossover points. These are regions where one of the strands in a given double helix switches to an adjacent ring, bridging the gap between concentric helices. Such crossovers help maintain the structure of concentric rings, preventing the DNA from extending. …

Varying the number of nucleotides between crossover points and the placement of crossovers allows the designer to combine sharp and rounded elements in a single 2-D form …

The network of crossover points also can be designed in such a way as to produce combinations of in-plane and out-of-plane curvature, allowing for the design of curved 3D nanostructures. While this method shows considerable versatility, the range of curvature is still limited for standard B form DNA, which will not tolerate large deviations from its preferred configuration – 10.5 base pairs/turn. However, as Jeanette Nangreave, one of the paper’s co-authors, explains, “Hao recognized that if you could slightly over twist or under twist these helices, you could produce different bending angles.”

Combining the method of concentric helices with such non-B-form DNA (with 9-12 base pairs/turn), enabled the group to produce sophisticated forms, including spheres, hemispheres, ellipsoid shells and finally—as a tour de force of nanodesign – a round-bottomed nanoflask, which appears unmistakably in a series of startling transmission electron microscopy images …

Yan hopes to further expand the range of nanoforms possible through the new technique. Eventually, this will require longer lengths of single-stranded DNA able to provide necessary scaffolding for larger, more elaborate structures. He credits his brilliant student (and the paper’s first author) Dongran Han with a remarkable ability to conceptualize 2-D and 3-D nanoforms and to navigate the often-perplexing details of their design. Ultimately however, more sophisticated nanoarchitectures will require computer-aided design programs – an area the team is actively pursuing.

The successful construction of closed, 3-D nanoforms, such as the sphere, has opened the door to many exciting possibilities for the technology, particularly in the biomedical realm. Nanospheres could be introduced into living cells for example, releasing their contents under the influence of endonucleases or other digestive components. Another strategy might use such spheres as nanoreactors – sites where chemicals or functional groups could be brought together to accelerate reactions or carry out other chemical manipulations.

These 3D structures are large and complex compared with the 2.0 nm diameter of the DNA double helix. PowerPoint slides of several figures from the Science paper can be downloaded without a subscription. One of these shows the sphere to be 42 nm in diameter, the prolate ellipsoid to be 35 nm by 66 nm, and the round-bottomed nanoflask to be 40 nm wide and 70 nm tall.

Real-time monitoring of atomic force microscope probes

The use of atomic force microscopes to manipulate atoms on surfaces (particularly on silicon surfaces—see for example this post and this post) is one promising alternative path for advancing nanotechnology toward productive nanosystems and molecular manufacturing. As was apparent in a recent interview of Professor Philip Moriarty of the University of Nottingham, many of the major technical obstacles encountered with atomic force microscopes involve the quality and reproducibility of the tips. Physorg.com points to a National Institute of Standards and Technology (NIST) news release of a way to improve AFM tips. “Getting the Point: Real-Time Monitoring of Atomic-Microscope Probes Adjusts for Wear“

Scientists at the National Institute of Standards and Technology (NIST) have developed a way to measure the wear and degradation of the microscopic probes used to study nanoscale structures in situ and as it’s happening. Their technique can both dramatically speed up and improve the accuracy of the most precise and delicate nanoscale measurements done with atomic force microscopy (AFM).

If you’re trying to measure the contours of a surface with a ruler that’s crumbling away as you work, then you at least need to know how fast and to what extent it is being worn away during the measurement.

This has been the challenge for researchers and manufacturers trying to create images of the surfaces of nanomaterials and nanostructures. Taking a photo is impossible at such small scales, so researchers use atomic force microscopes. Think of a device like a phonograph needle being used, on a nanoscale, to measure the peaks and valleys as it’s dragged back and forth across a surface. These devices are used extensively in nanoscale imaging to measure the contours of nanostructures, but the AFM tips are so small that they tend to wear down as they traverse the surface being measured.

Today, most researchers stop the measurement to “take a picture” of the tip with an electron microscope, a time-consuming method prone to inaccuracies.

NIST materials engineer Jason Killgore has developed a method for measuring in real time the extent to which AFM tips wear down. Killgore measures the resonant frequency of the AFM sensor tip, a natural vibration rate like that of a tuning fork, while the instrument is in use. Because changes to the size and shape of the tip affect its resonant frequency, he is able to measure the size of the AFM’s tip as it works—in increments of a tenth of a nanometer, essentially atomic scale resolution. …

The potential impact of this development is considerable. Thousands of AFMs are in use at universities, manufacturing plants and research and development facilities around the world. Improving their ability to measure and image nanosized devices will improve the quality and effectiveness of those devices. Another benefit is that developing new measurement tips—and studying the properties of new materials used in those tips—will be much easier and faster, given the immediate feedback about wear rates.

We can also hope that this method will facilitate the manipulation of atoms for mechanosynthesis being done by pioneers like Zyvex Labs, Prof. Moriarty, and the team that won the 2009 Feynman Prize for Experimental work.

Nanotechnology boosts anticancer drug cocktail many times over

Using nanoparticles for drug delivery, particularly to treat cancer, has been under development for several years. Liposomes were one of the earliest and simplest types of nanoparticles used for cancer drug delivery, and were often not much more complex than vesicles of lipid bilayer, typically less than 200 nm in diameter, encapsulating an anticancer drug. Now more complex and sophisticated nanoparticles promise to be much more effective in treating cancer. We thank KurzweilAI for pointing to this news release from Sandia National Laboratories and the University of New Mexico “Sandia and UNM lead effort to destroy cancers: Boosting medicine with nanotechnology strengthens drug cocktail many times over“

Melding nanotechnology and medical research, Sandia National Laboratories, the University of New Mexico, and the UNM Cancer Research and Treatment Center have produced an effective strategy that uses nanoparticles to blast cancerous cells with a mélange of killer drugs.

In the cover article [abstract] of the May issue of Nature Materials, available online April 17 , the researchers describe silica nanoparticles about 150 nanometers in diameter as honeycombed with cavities that can store large amounts and varieties of drugs.

“The enormous capacity of the nanoporous core, with its high surface area, combined with the improved targeting of an encapsulating lipid bilayer [called a liposome], permit a single ‘protocell’ loaded with a drug cocktail to kill a drug-resistant cancer cell,” says Sandia researcher and UNM professor Jeff Brinker, the principal investigator. “That’s a millionfold increase in efficiency over comparable methods employing liposomes alone — without nanoparticles — as drug carriers.”

The nanoparticles and the surrounding cell-like membranes formed from liposomes together become the combination referred to as a protocell: the membrane seals in the deadly cargo and is modified with molecules (peptides) that bind specifically to receptors overexpressed on the cancer cell’s surface. (Too many receptors is one signal the cell is cancererous.) The nanoparticles provide stability to the supported membrane and contain and release the therapeutic cargo within the cell.

A current Food and Drug Administration-approved nanoparticle delivery strategy is to use liposomes themselves to contain and deliver the cargo. In a head-to-head comparison of targeted liposomes and protocells with identical membrane and peptide compositions, Brinker and colleagues report that the greater cargo capacity, stability and targeting efficacy of protocells leads to many times greater cytotoxicity [destruction] directed specifically toward human liver cancer cells.

Another advantage to protocells over lipsomes alone, says lead author Carlee Ashley, a Harry S. Truman post-doctoral fellow at Sandia’s California site in Livermore, is that liposomes used as carriers need specialized loading strategies that make the process more difficult. “We’ve demonstrated we can just soak nanoparticles to load them with unique drug combinations needed for personalized medicine. They effectively encapsulate toxins as well as siRNA [ribonucleic acid] that silence expressions of proteins.”

RNA, the biological messenger that tells cells which proteins to manufacture, in this case is used to silence the cellular factory, a way of causing apoptosis or cell death. “Si” is short for “silence.”

The lipids also serve as a shield that restricts toxic chemotherapy drugs from leaking from the nanoparticle until the protocell binds to and takes hold within the cancer cell. This means that few poisons leak into the system of the human host, if the protocells find no cancer cells. This cloaking mitigates toxic side effects expected from conventional chemotherapy.

Instead, the particles — crafted small enough to float under the radar of the liver and other cleansing organs — can circulate harmlessly for days or weeks, depending on their engineered size, seeking their prey. …

“Proteins modified with a targeting peptide that binds to a particular carcinoma exhibit a 10,000-fold greater affinity for that cancer than for other unrelated cells,” Ashley said. …

The method may be commercially available in five years, researchers estimate.

During the past few weeks several very different types of nanoparticles have been reported as showing great promise, and these will be the subject of additional posts soon. Nanomedicine, and especially targeted drug delivery, is looking like an area where accelerating progress is demonstrating the value of investing in building increasingly complex nanostructures. Will this trend continue until we reach the point of atomically precise manufacture of medical nanorobots?

Much faster directed evolution of proteins could speed development of molecular machine systems

One of the principal recommendations of the Technology Roadmap for Productive Nanosystems was to “Support the development of modular molecular composite nanosystems (MMCNs)” (see Productive Nanosystems: A Technology Roadmap, page 12 of 198-page PDF) in which large DNA frameworks (of the sort that we cited last week) are to be used to support relatively rigid functional objects of engineered proteins. Such proteins could be engineered through either rational design or directed evolution (for an excellent update on the latter see this post by Eric Drexler on Metamodern from October 2010). Laboratory-directed evolution can, however, be laborious, so anything to make it faster and easier might substantially advance this approach. Now ScienceDaily points to this Harvard Gazette article by Steve Bradt that announces a major improvement “Speeding up biomolecular evolution: New approach proves 100 times faster than before“:

Scientists at Harvard University have harnessed the prowess of fast-replicating bacterial viruses, also known as phages, to accelerate the evolution of biomolecules in the laboratory. The work, reported in the journal Nature [abstract], could ultimately allow the tailoring of custom pharmaceuticals and research tools from lab-grown proteins, nucleic acids, and other such compounds.

The researchers, led by Professor David R. Liu, say that their approach — dubbed “phage-assisted continuous evolution,” or PACE — is roughly 100 times faster than conventional laboratory evolution, and far less labor-intensive for scientists.

“Most modern drugs are based on small organic molecules, but biological macromolecules may be better suited as pharmaceuticals in some cases,” said Liu, a professor of chemistry and chemical biology at Harvard and an investigator with the Howard Hughes Medical Institute. “Our work provides a new solution to one of the key challenges in the use of macromolecules as research tools or human therapeutics: how to rapidly generate proteins or nucleic acids with desired properties.”

Liu and Harvard co-authors Kevin M. Esvelt and Jacob C. Carlson achieved up to 60 rounds of protein evolution every 24 hours by linking laboratory evolution to the life cycle of a virus that infects bacteria. This phage’s life cycle of just 10 minutes is among the fastest known. Because this generation time is so brief, the phage makes a perfect vehicle for accelerated protein evolution. The PACE system uses E. coli host cells to produce the resulting proteins, to serve as factories for phage production, and to perform the key selection step that allows phage-carrying genes encoding desired molecules to flourish.

In three protein evolution experiments, PACE was able to generate an enzyme with a new target activity within a week, achieving up to 200 rounds of protein evolution during that time. Conventional laboratory evolution methods, Liu said, would require years to complete this many rounds of evolution. …

It remains to be seen just how general this method will be since it depends on linking the activity to be evolved to protein production in bacteria. Nevertheless, it looks like the individual components of MMCN development are progressing nicely. Perhaps the next challenge is to see if the pieces can be put together to make effective molecular machine systems, leading eventually to atomically precise productive nanosystems.

Nanotechnology in Civil Infrastructure

Nanotechnology in Civil Infrastructure is a state-of-the art reference source describing the latest developments in nano-engineering and nano-modification of construction materials to improve the bulk properties, development of sustainable, intelligent, and smart concrete materials through the integration of nanotechnology based self-sensing and self-powered materials and cyber infrastructure technologies, review of nanotechnology applications in pavement engineering, development of novel, cost-effective, high-performance and long-lasting concrete products and processes through nanotechnology-based innovative processing of cement and cement paste, and advanced nanoscience modeling, visualization, and measurement systems for characterizing and testing civil infrastructure materials at the nano-scale.

Nanotechnology application In Energy Sector

Nanotechnology applications could provide decisive technological breakthroughs in the energy sector and have a considerable impact on creating the sustainable energy supply that is required to make the transition from fossil fuels. Possibilities range from gradual short- and medium-term improvements for a more efficient use of conventional and renewable energy sources all the way to completely new long-term approaches for energy recovery and utilization. With enough political will – and funding – nanotechnology could make essential contributions to sustainable energy supply and global climate protection policies. The technological foundation is there, all it takes is political leadership to create the right research and investment conditions to make it happen.