Ab initio phasing macromolecular structures using electron-counted MicroED data

Structures of two globular proteins were determined ab initio using microcrystal electron diffraction (MicroED) data that were collected on a direct electron detector in counting mode. Microcrystals were identified using a scanning electron microscope (SEM) and thinned with a focused ion beam (FIB) to produce crystalline lamellae of ideal thickness. Continuous-rotation data were collected using an ultra-low exposure rate to enable electron counting in diffraction. For the first sample, triclinic lysozyme extending to a resolution of 0.87 Å, an ideal helical fragment of only three alanine residues provided initial phases. These phases were improved using density modification, allowing the entire atomic structure to be built automatically. A similar approach was successful on a second macromolecular sample, proteinase K, which is much larger and diffracted to a resolution of 1.5 Å. These results demonstrate that macromolecules can be determined to sub-ångström resolution by MicroED and that ab initio phasing can be successfully applied to counting data

Paper Session II-A - ISOBUS A Faster, Better, Cheaper Tool for Space Flight Experiments

Space exploration and related investigations have been suffering from programmatic inefficiencies inherent to customized projects. One-of-a-kind space investigations such as experiments, installations, platforms, and missions all lack the profit-driven architectures and money-making methodologies that characterize commercial enterprise. The foundation of long-tenm commercial success is in the smart and efficient utilization of capital investment. An enterprise that throws away its tools, its infrastructure, its expertise, and its capital, every time it completes a project is not likely to be able to afford to do so again and again. When resources are scarce, one must utilize them efficiently. Proven commercial methodologies such as standardization, mass production, miniaturization, modular interchangeability, and reusability . of tools, facilities, and resources are the principal techniques by which products can be created faster-better-cheaper. Commercial investigators in intensely competitive fields, such as biotechnology, have successfully applied these principles to their experimental setups, tools, and support systems. We must similarly employ commercial principles if we are to survive the expensive challenge of future space exploration. This paper introduces a faster-bettercheaper\u27\u27 approach for space investigators. The approach employs a tool called ISOBUS

Collection of continuous rotation MicroED Data from Ion Beam Milled Crystals of Any Size

Microcrystal electron diffraction (MicroED) allows for macromolecular structure solution from nanocrystals. To create crystals of suitable size for MicroED data collection, sample preparation typically involves sonication or pipetting a slurry of crystals from a crystallization drop. The resultant crystal fragments are fragile and the quality of the data that can be obtained from them is sensitive to subsequent sample preparation for cryoelectron microscopy as interactions in the water-air interface can damage crystals during blotting. Here, we demonstrate the use of a focused ion beam to generate lamellae of macromolecular protein crystals for continuous rotation MicroED that are of ideal thickness, easy to locate, and require no blotting optimization. In this manner, crystals of nearly any size may be scooped and milled to desired dimensions prior to data collection, thus streamlining the methodology for sample preparation for MicroED

Excitation of guided waves in layered structures with negative refraction

We study the electromagnetic beam reflection from layered structures that include the so-called double-negative materials, also called left-handed metamaterials. We predict that such structures can demonstrate a giant lateral Goos-Hanchen shift of the scattered beam accompanied by splitting of the reflected and transmitted beams due to the resonant excitation of surface waves at the interfaces between the conventional and double-negative materials as well as due to excitation of leaky modes in the layered structures. The beam shift can be either positive or negative, depending on the type of the guided waves excited by the incoming beam. We also perform finite-difference time-domain simulations and confirm the major effects predicted analytically.Comment: 13 pqages, 10 figures. Also available at http://www.opticsexpress.org/abstract.cfm?URI=OPEX-13-2-48

Qualitative Analyses of Polishing and Precoating FIB Milled Crystals for MicroED

Microcrystal electron diffraction (MicroED) leverages the strong interaction between matter and electrons to determine protein structures from vanishingly small crystals. This strong interaction limits the thickness of crystals that can be investigated by MicroED, mainly due to absorption. Recent studies have demonstrated that focused ion-beam (FIB) milling can thin crystals into ideal-sized lamellae; however, it is not clear how to best apply FIB milling for MicroED. Here, the effects of polishing the lamellae, whereby the last few nanometers are milled away using a low-current gallium beam, are explored in both the platinum-precoated and uncoated samples. Our results suggest that precoating samples with a thin layer of platinum followed by polishing the crystal surfaces prior to data collection consistently led to superior results as indicated by higher signal-to-noise ratio, higher resolution, and better refinement statistics. This study lays the foundation for routine and reproducible methodology for sample preparation in MicroED

Solving a "Hard" Problem to Approximate an "Easy" One: Heuristics for Maximum Matchings and Maximum Traveling Salesman Problems

We consider geometric instances of the Maximum Weighted Matching Problem (MWMP) and the Maximum Traveling Salesman Problem (MTSP) with up to 3,000,000 vertices. Making use of a geometric duality relationship between MWMP, MTSP, and the Fermat-Weber-Problem (FWP), we develop a heuristic approach that yields in near-linear time solutions as well as upper bounds. Using various computational tools, we get solutions within considerably less than 1% of the optimum. An interesting feature of our approach is that, even though an FWP is hard to compute in theory and Edmonds' algorithm for maximum weighted matching yields a polynomial solution for the MWMP, the practical behavior is just the opposite, and we can solve the FWP with high accuracy in order to find a good heuristic solution for the MWMP.Comment: 20 pages, 14 figures, Latex, to appear in Journal of Experimental Algorithms, 200

Paper Session I-A - In-Space Welding Visions & Realities

This paper establishes the value of having an in-space welding capability and identifies its applications, both near-term for Shuttle-Spacelab missions and Space Station Freedom, and longer-term for the First Lunar Outpost and Manned Mission to Mars. The leading candidate technologies, consisting of Electron Beam, Gas Tungsten Arc, Plasma Arc, and Laser Beam, are examined against the criteria for an in-space welding system. Research and development work to date, striving to achieve an in-space welding capability, is reviewed. Finally, a series of strategic NASA flight experiments is discussed as the remaining development required for achieving a complete in-space welding capability, which can fully serve the Space Exploration Initiative. This paper summarizes the visions and realities associated with in-space welding

Harmonic scalpel versus flexible CO2 laser for tongue resection: A histopathological analysis of thermal damage in human cadavers

<p>Abstract</p> <p>Background</p> <p>Monopolar cautery is the most commonly used surgical cutting and hemostatic tool for head and neck surgery. There are newer technologies that are being utilized with the goal of precise cutting, decreasing blood loss, reducing thermal damage, and allowing faster wound healing. Our study compares thermal damage caused by Harmonic scalpel and CO2 laser to cadaveric tongue.</p> <p>Methods</p> <p>Two fresh human cadaver heads were enrolled for the study. Oral tongue was exposed and incisions were made in the tongue akin to a tongue tumor resection using the harmonic scalpel and flexible C02 laser fiber at various settings recommended for surgery. The margins of resection were sampled, labeled, and sent for pathological analysis to assess depth of thermal damage calculated in millimeters. The pathologist was blinded to the surgical tool used. Control tongue tissue was also sent for comparison as a baseline for comparison.</p> <p>Results</p> <p>Three tongue samples were studied to assess depth of thermal damage by harmonic scalpel. The mean depth of thermal damage was 0.69 (range, 0.51 - 0.82). Five tongue samples were studied to assess depth of thermal damage by CO2 laser. The mean depth of thermal damage was 0.3 (range, 0.22 to 0.43). As expected, control samples showed 0 mm of thermal damage. There was a statistically significant difference between the depth of thermal injury to tongue resection margins by harmonic scalpel as compared to CO2 laser, (p = 0.003).</p> <p>Conclusion</p> <p>In a cadaveric model, flexible CO2 laser fiber causes less depth of thermal damage when compared with harmonic scalpel at settings utilized in our study. However, the relevance of this information in terms of wound healing, hemostasis, safety, cost-effectiveness, and surgical outcomes needs to be further studied in clinical settings.</p