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At least 1,549 records · Page 86

Engineered reduction of S-adenosylmethionine alters lignin in sorghum

Abstract Background Lignin is an aromatic polymer deposited in secondary cell walls of higher plants to provide strength, rigidity, and hydrophobicity to vascular tissues. Due to its interconnections with cell wall polysaccharides, lignin plays important roles during plant growth and defense, but also has a negative impact on industrial processes aimed at obtaining monosaccharides from plant biomass. Engineering lignin offers a solution to this issue. For example, previous work showed that heterologous expression of a coliphage S -adenosylmethionine hydrolase (AdoMetase) was an effective approach to reduce lignin in the model plant Arabidopsis. The efficacy of this engineering strategy remains to be evaluated in bioenergy crops. Results We studied the impact of expressing AdoMetase on lignin synthesis in sorghum ( Sorghum bicolor L. Moench). Lignin content, monomer composition, and size, as well as biomass saccharification efficiency were determined in transgenic sorghum lines. The transcriptome and metabolome were analyzed in stems at three developmental stages. Plant growth and biomass composition was further evaluated under field conditions. Results evidenced that lignin was reduced by 18% in the best transgenic line, presumably due to reduced activity of the S -adenosylmethionine-dependent O -methyltransferases involved in lignin synthesis. The modified sorghum features altered lignin monomer composition and increased lignin molecular weights. The degree of methylation of glucuronic acid on xylan was reduced. These changes enabled a ~20% increase in glucose yield after biomass pretreatment and saccharification compared to wild type. RNA-seq and untargeted metabolomic analyses evidenced some pleiotropic effects associated with AdoMetase expression. The transgenic sorghum showed developmental delay and reduced biomass yields at harvest, especially under field growing conditions. Conclusions The expression of AdoMetase represents an effective lignin engineering approach in sorghum. However, considering that this strategy potentially impacts multiple S -adenosylmethionine-dependent methyltransferases, adequate promoters for fine-tuning AdoMetase expression will be needed to mitigate yield penalty.

Tian, Yang↗

Chapter 21: The Utility of Virtual Reality for Science and Engineering

In our daily usage of the large-scale immersive virtual environment at the National Renewable Energy Laboratory (NREL), we have observed how this VR system can be a useful tool to enhance scientific and engineering workflows. On multiple occasions, we have observed scientists and engineers discover features in their data using immersive environments that they had not seen in prior investigations of their data on traditional desktop displays. We have embedded more information into our analytics tools, allowing engineers to explore complex multivariate spaces. We have observed natural interactions with 3D objects and how those interactions seem to catalyze understanding. And we have seen improved collaboration with groups of stakeholders. In this chapter, we discuss these practical advantages of immersive visualization in the context of several real-world examples.

97 MATHEMATICS AND COMPUTING↗

Data from A Mass Spectrometry-Based High-Throughput Screening Method for Engineering Fatty Acid Synthases with Improved Production of Medium Chain Fatty Acids

Microbial cell factories have been extensively engineered to produce free fatty acids (FFAs) as key components of crucial nutrients, soaps, industrial chemicals, and fuels. However, our ability to control the composition of microbially synthesized FFAs is still limited, particularly, for producing medium‐chain fatty acids (MCFAs). This is mainly due to the lack of high‐throughput approaches for FFA analysis to engineer enzymes with desirable product specificity. Here we report a mass spectrometry (MS)‐based method for rapid profiling of MCFAs in Saccharomyces cerevisiae by using membrane lipids as a proxy. In particular, matrix‐assisted laser desorption/ionization time‐of‐flight (MALDI‐ToF) MS was used to detect shorter acyl chain phosphatidylcholines from membrane lipids and a higher m/z peak ratio at 730 and 758 was used as an indication for improved MCFA production. This colony‐based method can be performed at a rate of ~2 s per sample, representing a substantial improvement over gas chromatography‐MS (typically >30 min per sample) as the gold standard method for FFA detection. To demonstrate the power of this method, we performed site‐saturation mutagenesis of the yeast fatty acid synthase and identified nine missense mutations that resulted in improved MCFA production relative to the wild‐type strain. Colony‐based MALDI‐ToF MS screening provides an effective approach for engineering microbial fatty acid compositions in a high‐throughput manner.

Conversion↗

Digital risk analysis in nuclear engineering projects: Designing for safety, performance, reliability, and security

Cyber-informed engineering and security-by-design frameworks are important in promoting the need to identify cybersecurity concerns early in the systems engineering lifecycle so risks from adversarial cyber-attacks can be eliminated or reduced through engineering design practices. In addition to adversarial risk, risk in operational technology systems also includes non-adversarial and unintentional risk from other factors such as human performance errors, environmental conditions, design flaws, and device degradation or failure. This paper introduces a new concept for characterizing digital risk, both adversarial and non-adversarial, and provides the basis for initial research into a novel digital risk analysis approach focused on incorporating attack difficulty into a multi-attribute analysis technique using robust decision-making. This digital risk characterization is also used to frame a discussion on the challenges of competing objectives and competing stakeholder requirements in an integrated energy system project that incorporates a small modular reactor and industrial facility.

22 GENERAL STUDIES OF NUCLEAR REACTORS↗

Structure of Type IIL Restriction-Modification Enzyme MmeI in Complex with DNA Has Implications for Engineering New Specificities

The creation of restriction enzymes with programmable DNA-binding and -cleavage specificities has long been a goal of modern biology. The recently discovered Type IIL MmeI family of restriction-and-modification (RM) enzymes that possess a shared target recognition domain provides a framework for engineering such new specificities. However, a lack of structural information on Type IIL enzymes has limited the repertoire that can be rationally engineered. We report here a crystal structure of MmeI in complex with its DNA substrate and an S-adenosylmethionine analog (Sinefungin). The structure uncovers for the first time the interactions that underlie MmeI-DNA recognition and methylation (5’-TCCRAC-3’; R = purine) and provides a molecular basis for changing specificity at four of the six base pairs of the recognition sequence (5’-TCCRAC-3’). Surprisingly, the enzyme is resilient to specificity changes at the first position of the recognition sequence (5’-TCCRAC-3’). Collectively, the structure provides a basis for engineering further derivatives of MmeI and delineates which base pairs of the recognition sequence are more amenable to alterations than others.

59 BASIC BIOLOGICAL SCIENCES↗

How to engineer the unknown: Advancing a quantitative and predictive understanding of plant and soil biology to address climate change

Our basic understanding of carbon cycling in the biosphere remains qualitative and incomplete, precluding our ability to effectively engineer novel solutions to climate change. How can we attempt to engineer the unknown? This challenge has been faced before in plant biology, providing a roadmap to guide future efforts. We use examples from over a century of photosynthesis research to illustrate the key principles that will set future plant engineering on a solid footing, namely, an effort to identify the key control variables, quantify the effects of systematically tuning these variables, and use theory to account for these observations. The main contributions of plant synthetic biology will stem not from delivering desired genotypes but from enabling the kind of predictive understanding necessary to rationally design these genotypes in the first place. Only then will synthetic plant biology be able to live up to its promise.

59 BASIC BIOLOGICAL SCIENCES↗

Engineering and Two-Stage Evolution of a Lignocellulosic Hydrolysate-Tolerant Saccharomyces cerevisiae Strain for Anaerobic Fermentation of Xylose from AFEX Pretreated Corn Stover

The inability of the yeast Saccharomyces cerevisiae to ferment xylose effectively under anaerobic conditions is a major barrier to economical production of lignocellulosic biofuels. Although genetic approaches have enabled engineering of S. cerevisiae to convert xylose efficiently into ethanol in defined lab medium, few strains are able to ferment xylose from lignocellulosic hydrolysates in the absence of oxygen. This limited xylose conversion is believed to result from small molecules generated during biomass pretreatment and hydrolysis, which induce cellular stress and impair metabolism. Here, we describe the development of a xylose-fermenting S. cerevisiae strain with tolerance to a range of pretreated and hydrolyzed lignocellulose, including Ammonia Fiber Expansion (AFEX)-pretreated corn stover hydrolysate (ACSH). We genetically engineered a hydrolysate-resistant yeast strain with bacterial xylose isomerase and then applied two separate stages of aerobic and anaerobic directed evolution. The emergent S. cerevisiae strain rapidly converted xylose from lab medium and ACSH to ethanol under strict anaerobic conditions. Metabolomic, genetic and biochemical analyses suggested that a missense mutation in GRE3, which was acquired during the anaerobic evolution, contributed toward improved xylose conversion by reducing intracellular production of xylitol, an inhibitor of xylose isomerase. These results validate our combinatorial approach, which utilized phenotypic strain selection, rational engineering and directed evolution for the generation of a robust S. cerevisiae strain with the ability to ferment xylose anaerobically from ACSH.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

SAXS studies of X-ray induced disulfide bond damage: Engineering high-resolution insight from a low-resolution technique

A significant problem in biological X-ray crystallography is the radiation chemistry caused by the incident X-ray beam. This produces both global and site-specific damage. Site specific damage can misdirect the biological interpretation of the structural models produced. Cryo-cooling crystals has been successful in mitigating damage but not eliminating it altogether; however, cryo-cooling can be difficult in some cases and has also been shown to limit functionally relevant protein conformations. The doses used for X-ray crystallography are typically in the kilo-gray to mega-gray range. While disulfide bonds are among the most significantly affected species in proteins in the crystalline state at both cryogenic and higher temperatures, there is limited information on their response to low X-ray doses in solution, the details of which might inform biomedical applications of X-rays. In this work we engineered a protein that dimerizes through a susceptible disulfide bond to relate the radiation damage processes seen in cryo-cooled crystals to those closer to physiologic conditions. This approach enables a low-resolution technique, small angle X-ray scattering (SAXS), to detect and monitor a residue specific process. A dose dependent fragmentation of the engineered protein was seen that can be explained by a dimer to monomer transition through disulfide bond cleavage. This supports the crystallographically derived mechanism and demonstrates that results obtained crystallographically can be usefully extrapolated to physiologic conditions. Fragmentation was influenced by pH and the conformation of the dimer, providing information on mechanism and pointing to future routes for investigation and potential mitigation. The novel engineered protein approach to generate a large-scale change through a site-specific interaction represents a promising tool for advancing radiation damage studies under solution conditions.

59 BASIC BIOLOGICAL SCIENCES↗

Engineering-Scale Demonstration of DuraLith and Ceramicrete Waste Forms

To support the selection of a waste form for the liquid secondary wastes from the Hanford Waste Immobilization and Treatment Plant, Washington River Protection Solutions (WRPS) has initiated secondary waste form testing on four candidate waste forms. Two of the candidate waste forms have not been developed to scale as the more mature waste forms. This work describes engineering-scale demonstrations conducted on Ceramicrete and DuraLith candidate waste forms. Both candidate waste forms were successfully demonstrated at an engineering scale. A preliminary conceptual design could be prepared for full-scale production of the candidate waste forms. However, both waste forms are still too immature to support a detailed design. Formulations for each candidate waste form need to be developed so that the material has a longer working time after mixing the liquid and solid constituents together. Formulations optimized based on previous lab studies did not have sufficient working time to support large-scale testing. The engineering-scale testing was successfully completed using modified formulations. Further lab development and parametric studies are needed to optimize formulations with adequate working time and assess the effects of changes in raw materials and process parameters on the final product performance. Studies on effects of mixing intensity on the initial set time of the waste forms are also needed.

12 MANAGEMENT OF RADIOACTIVE AND NON-RADIOACTIVE W↗

TriBITS Lifecycle Model: A Lean/Agile Software Lifecycle Model for Research-based Computational Science and Engineering and Applied Mathematical Software (V.1.0)

Software lifecycles are becoming an increasingly important issue for computational science and engineering (CSE) software. The process by which a piece of CSE software begins life as a set of research requirements and then matures into a trusted high-quality capability is both commonplace and extremely challenging. Although an implicit lifecycle is obviously being used in any effort, the challenges of this process - respecting the competing needs of research vs. production - cannot be overstated. Here we describe a proposal for a well-defined software lifecycle process based on modern Lean/Agile software engineering principles. What we propose is appropriate for many CSE software projects that are initially heavily focused on research but also are expected to eventually produce usable high-quality capabilities. The model is related to TriBITS, a build, integration and testing system, which serves as a strong foundation for this lifecycle model, and aspects of this lifecycle model are ingrained in the TriBITS system. Here, we advocate three to four phases or maturity levels that address the appropriate handling of many issues associated with the transition from research to production software. The goals of this lifecycle model are to better communicate maturity levels with customers and to help to identify and promote Software Engineering (SE) practices that will help to improve productivity and produce better software. An important collection of software in this domain is Trilinos, which is used as the motivation and the initial target for this lifecycle model. However, many other related and similar CSE (and non-CSE) software projects can also make good use of this lifecycle model, especially those that use the TriBITS system. Indeed this lifecycle process, if followed, will enable large-scale sustainable integration of many complex CSE software efforts across several institutions.

99 GENERAL AND MISCELLANEOUS↗

Brief 70 Nuclear Engineering Enrollments and Degrees, 2011: Summary Information

The survey includes degrees granted between September 1, 2010 and August 31, 2011. Enrollment information refers to the fall term 2011. The enrollment and degree data include students majoring in nuclear engineering or in an option program equivalent to a major. Thirty-two academic programs reported having nuclear engineering programs during 2011, and data was received from all thirty-two programs. The data for two nuclear engineering programs include enrollments and degrees in health physics options that are also reported in the health physics enrollments and degrees data.

96 KNOWLEDGE MANAGEMENT AND PRESERVATION↗

Genetically Engineered Magnetic Algae A Leap toward Affordable Biofuel from Algae

From the earliest days of internal-combustion engines, technological visionaries dreamed about engines running on fuel made from plants. In those early days, however, it was much easier to refine fuels from crude oil, which could be obtained cheaply in many parts of the world by simply drilling a hole in the ground. However, humanity now knows that the supply of fossil fuel is limited. One day, we will run out of fossil fuel, and we will need fuel from alternative sources. One possible alternative is biofuels, but these remain a dream away today, simply because producing such fuels costs as much as $\$$10 per gallon and requires petroleum products for their production. But this long-held dream may transcend the veil of sleep into the realm of reality, thanks to the development of a technology designed to harvest algae more easily and cost effectively—algae that are major candidates for producing tomorrow’s biofuels. At Los Alamos National Laboratory, we have developed the world’s first “magnetic algae.” We say magnetic, but more specifically our genetically engineered algae are paramagnetic. Paramagnetic materials are themselves not magnetic. Instead, such materials are pulled by magnetic fields. Hence, a magnetic needle made from soft iron is actually paramagnetic, because a magnet can pull it from a haystack. For the first time ever, we have genetically modified a model algae strain to make what is called intracellular magnetic nanoparticles.

09 BIOMASS FUELS↗

WTEC Panel Report on International Assessment of Research and Development in Simulation-Based Engineering and Science

This WTEC panel report assesses the international research and development activities in the field of Simulation- Based Engineering and Science (SBE&S). SBE&S involves the use of computer modeling and simulation to solve mathematical formulations of physical models of engineered and natural systems. SBE&S today has reached a level of predictive capability that it now firmly complements the traditional pillars of theory and experimentation/observation. As a result, computer simulation is more pervasive today – and having more impact – than at any other time in human history. Many critical technologies, including those to develop new energy sources and to shift the cost-benefit factors in healthcare, are on the horizon that cannot be understood, developed, or utilized without simulation. A panel of experts reviewed and assessed the state of the art in SBE&S as well as levels of activity overseas in the broad thematic areas of life sciences and medicine, materials, and energy and sustainability; and in the crosscutting issues of next generation hardware and algorithms; software development; engineering simulations; validation, verification, and uncertainty quantification; multiscale modeling and simulation; and SBE&S education. The panel hosted a U.S. baseline workshop, conducted a bibliometric analysis, consulted numerous experts and reports, and visited 59 institutions and companies throughout East Asia and Western Europe to explore the active research projects in those institutions, the computational infrastructure used for the projects, the funding schemes that enable the research, the collaborative interactions among universities, national laboratories, and corporate research centers, and workforce needs and development for SBE&S.

97 MATHEMATICS AND COMPUTING↗

Trilinos Developers SQE Guide: ASC Software Quality Engineering Practices Version 3.0

The Trilinos Project is an effort to develop algorithms and enabling technologies within an object-oriented software framework for the solution of large-scale, complex multi-physics engineering and scientific problems. A new software capability is introduced into Trilinos as a package. A Trilinos package is an integral unit and, although there are exceptions such as utility packages, each package is typically developed by a small team of experts in a particular algorithms area such as algebraic preconditioners, nonlinear solvers, etc. The Trilinos Developers SQE Guide is a resource for Trilinos package developers who are working under Advanced Simulation and Computing (ASC) and are therefore subject to the ASC Software Quality Engineering Practices as described in the Sandia National Laboratories Advanced Simulation and Computing (ASC) Software Quality Plan: ASC Software Quality Engineering Practices Version 3.0 document. The Trilinos Developer Policies webpage contains a lot of detailed information that is essential for all Trilinos developers. The Trilinos Software Lifecycle Model defines the default lifecycle model for Trilinos packages and provides a context for many of the practices listed in this document.

Willenbring, James Michael↗

Integrating the human element into the systems engineering process and MBSE methodology

In response to the challenges related to the increasing size and complexity of systems, organizations have recognized the need to integrate human considerations in the beginning stages of systems development. Human Systems Integration (HSI) seeks to accomplish this objective by incorporating human factors within systems engineering (SE) processes and methodologies, which is the focus of this paper. A representative set of HSI methods from multiple sources are organized, analyzed, and mapped to the systems engineering Vee-model. These methods are then consolidated and evaluated against the SE process and Models-Based Systems Engineering (MBSE) methodology to determine where and how they could integrate within systems development activities in the form of specific enhancements. Overall conclusions based on these evaluations are presented and future research areas are proposed.

60 APPLIED LIFE SCIENCES↗

Evaluation of American Indian Science and Engineering Society Intertribal Middle School Science and Math Bowl Project

The American Indian Science and Engineering Society (AISES) has been funded under a U.S. Department of Energy (DOE) grant (Grant Award No. DE-SC0004058) to host an Intertribal Middle-School Science and Math Bowl (IMSSMB) comprised of teams made up of a majority of American Indian students from Bureau of Indian Education-funded schools and public schools. The intent of the AISES middle school science and math bowl is to increase participation of American Indian students at the DOE-sponsored National Science Bowl. Although national in its recruitment scope, the AISES Intertribal Science and Math Bowl is considered a “regional” science bowl, equivalent to the other 50 regional science bowls which are geographically limited to states. Most regional bowls do not have American Indian student teams competing, hence the AISES bowl is meant to encourage American Indian student teams to increase their science knowledge in order to participate at the national level. The AISES competition brings together teams from various American Indian communities across the nation. Each team is provided with funds for travel to and from the event, as well as for lodging and meals. In 2011 and 2012, there were 10 teams participating; in 2013, the number of teams participating doubled to 20. Each Science and Math Bowl team is comprised of four middle school — grades 6 through 8 — students, one alternate, and a teacher who serves as advisor and coach — although in at least two cases, the coach was not a teacher, but was the Indian Education Coordinator. Each team member must have at least a 3.0 GPA. Furthermore, the majority of students in each team must be comprised of American Indian, Alaska Native or Native Hawaiian students. Under the current DOE grant, AISES sponsored three annual middle school science bowl competitions over the years 2011, 2012 and 2013. The science and math bowls have been held in late March concurrently with the National American Indian Science and Engineering Fair (NAISEF) and EXPO at the Albuquerque, NM Convention Center. Albuquerque is also the home of the AISES national office. The AISES staff also recruits volunteers to assist with implementation of the science and math bowl event. In 2011, there were 7 volunteers; in 2012, 15 volunteers, and in 2013, 19 volunteers. Volunteers are recruited from a variety of local sources, including Sandia Laboratories, Southwest Indian Polytechnic Institute students, Department of Defense, as well as family members of AISES staff. For AISES, the goals of the Intertribal Middle School Science and Math Bowl project are to have more Native students learn science, for them to gain confidence in competing, and to reward their effort in order to motivate them to pursue studies in the sciences and engineering. For DOE, the goals of the project are to get more Native students to compete at the National Science Bowl, held in Washington, DC.

96 KNOWLEDGE MANAGEMENT AND PRESERVATION↗

Acquisition of Kerr Dam and Establishment of a Wholesale Power Generation Corporation to Operate It: Hiring of Power market Coordinator and Staff Engineer (Final Report)

This project aimed at supporting one key component of a major multi-step undertaking on the part of the CSKT: the acquisition of the Kerr Hydroelectric project and its subsequent operation as a wholesale power generation facility. This project provided support to kick-start the development of the organizational structure to acquire and operate the facility by acquiring critical expertise necessary for the acquisition by funding in part two key personnel for the first two years of the four-year organizational development process. These individuals provided the Tribes with expert knowledge in the highly specialized areas of resource balancing, power marketing, and hydro-engineering; essential prerequisites to the Tribes' ability to build an organization for the operation of the Kerr Project and to securing financial backing for the acquisition. Goals achieved: • Establishing an efficient and economic conveyance process, and transition plans • Establishing an efficient and effective Tribal wholesale power generation corporation to manage the plant, balance the resources, and market the power from the Kerr Project. The success of this project, which is essential to the Tribes' acquisition of the Kerr Hydroelectric facility, helps to address poverty and unemployment among Tribal members by generating a number of highly skilled and specialized, high-paying Tribal member jobs and providing a stream of income from power sales that will be used for Tribal economic development. Objectives achieved: The project supported the position of Power Plant Operations and Maintenance engineer and power marketing coordinator positions. These are key, in part, to the Tribes' successful acquisition and operation of the facility because they will enable to the Tribes to gain the very specialized expertise required to operate a large wholesale power generation facility. Specific objectives include: Objective 1: Hire a power marketing coordinator to develop and coordinate the appropriate power marketing strategy for the sale of power generated by the operation of Kerr Dam. Objective 2: Hire a staff engineer.

13 HYDRO ENERGY↗

Annual Report FY2014 Alternative Fuels DISI Engine Research

Due to concerns about future petroleum supply and accelerating climate change, increased engine efficiency and alternative fuels are of interest. This project contributes to the science-base needed by industry to develop highly efficient DISI engines that also beneficially exploit the different properties of alternative fuels. Lean operation is studied since it can provide higher efficiencies than traditional non-dilute stoichiometric operation. Since lean operation can lead to issues with ignition stability, slow flame propagation and low combustion efficiency, focus is on techniques that can overcome these challenges. Specifically, fuel stratification can be used to ensure ignition and completeness of combustion, but may lead to soot and NO x emissions challenges. Advanced ignition system and intake air preheating both promote ignition stability. Controlled end-gas autoignition can be used maintain high combustion efficiency for ultra-lean well-mixed conditions. However, the response of both combustion and exhaust emission to these techniques depends on the fuel properties. Therefore, to achieve optimal fuel-economy gains, the combustion-control strategies of the engine must adopt to the fuel being utilized.

32 ENERGY CONSERVATION, CONSUMPTION, AND UTILIZATI↗