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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 613 records · Page 34
All sourcesNASA NTRSDOE OSTI
DOE OSTI2021

Production of Biofuels from Biomass by Fungi

The use of abundantly available lignocellulosic biomass as a feedstock for biofuels has emerged as a sustainable alternative to fossil fuels. Industries across the world have shifted their focus to enhancing bioconversion through fungi, efficient and tractable organisms capable of producing valuable cost-effective enzymes. The many capabilities of fungal species offer opportunities for the tailored production of valuable compounds from a variety of substrates. This review details a high level examination of how fungal enzymes degrade plant polysaccharides to simple sugars through the sensitive coordination of transcription factors. Scientists have used a variety of tools like genetically engineering fungi, combining various species, and altering industrial/process conditions to enhance biomass deconstruction and fermentation while solving obstacles like inhibitory compounds. While ethanol and biodiesel are highlighted, additional biofuels are mentioned. These advances have uncovered themes like synergism and biocatalysis, led to the discovery of new fungal species, and revealed unique enzymatic mechanisms. Notably, this research highlights the complexity of metabolic systems within and between fungi. Fungi have been indispensable to the biofuel industry and future research will be crucial for energy sustainability.

Ottum, Eva MN↗
DOE OSTI2019

Cytosolic lipid droplets as engineered organelles for production and accumulation of terpenoid biomaterials in leaves

Cytosolic lipid droplets are endoplasmic reticulum-derived organelles typically found in seeds as reservoirs for physiological energy and carbon to fuel germination. In this work, we report synthetic biology approaches to co-produce high-value sesqui- or diterpenoids together with lipid droplets in plant leaves. The formation of cytosolic lipid droplets is enhanced in the transient Nicotiana benthamiana system through ectopic production of WRINKLED1, a key regulator of plastid fatty acid biosynthesis, and a microalgal lipid droplet surface protein. Engineering of the pathways providing the universal C5-building blocks for terpenoids and installation of terpenoid biosynthetic pathways through direction of the enzymes to native and non-native compartments boost the production of target terpenoids. We demonstrate that anchoring of distinct biosynthetic steps onto the surface of lipid droplets leads to efficient production of terpenoid scaffolds and functionalized terpenoids. The co-produced lipid droplets “trap” the terpenoids in the cells.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2012

Engineered microorganisms capable of producing target compounds under anaerobic conditions

The present invention is generally provides recombinant microorganisms comprising engineered metabolic pathways capable of producing C3-C5 alcohols under aerobic and anaerobic conditions. The invention further provides ketol-acid reductoisomerase enzymes which have been mutated or modified to increase their NADH-dependent activity or to switch the cofactor preference from NADPH to NADH and are expressed in the modified microorganisms. In addition, the invention provides isobutyraldehyde dehydrogenase enzymes expressed in modified microorganisms. Also provided are methods of producing beneficial metabolites under aerobic and anaerobic conditions by contacting a suitable substrate with the modified microorganisms of the present invention.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2016

Light-driven carbon dioxide reduction to methane by nitrogenase in a photosynthetic bacterium

Significance One of the most important life-sustaining metabolisms that results from a net reduction reaction is the conversion of nitrogen gas to ammonia by nitrogenase in a process known as nitrogen fixation. A nitrogenase variant has been described that converts carbon dioxide to methane in vitro. Here, we expressed this variant in an engineered strain of the photosynthetic bacterium Rhodopseudomonas palustris and showed biological light-driven methane production. Its ability to reduce carbon dioxide at ambient temperature and pressure with a single enzyme and energy provided by light makes the methane-producing strain of R. palustris an excellent starting point to understand how a biological system can marshal resources to produce an energy-rich hydrocarbon in one enzymatic step.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2022

Engineering of holocellulase in biomass-degrading fungi for sustainable biofuel production

Biofuels, such as bioethanol, are a clean and sustainable form of energy and have emerged as a viable alternative to fossil fuels. Plant biomass is an important raw material for the production of clean and renewable energy. The holocellulose contained in the composition of plants may be broken down into simple sugars, such as glucose, which are fermented by yeast to produce bioethanol. The conversion of glucose polymers into fermentable sugars is accomplished by enzymes known as holocellulases, which are produced by lignocellulolytic fungi. These enzymes act synergistically for the efficient degradation of cellulose polymers, and the fine and coordinated regulation of this process is performed by transcription factors (TFs). TFs are regulatory proteins that bind to the promoter region of their target genes (CAZymes, sugar transporters, signaling proteins, other TFs, etc.) to induce or repress their transcription. This review aims to understand the main regulatory mechanisms involved in plant biomass degradation by the most studied lignocellulolytic fungi Trichoderma sp., Aspergillus sp., Penicillium sp., and Neurospora crassa. In this context, the most studied TFs related to holocellulose degradation and genetic modification of TFs or promoters as a valuable tool to improve enzyme production for biotechnological purposes have been discussed. Furthermore, this review enables the expansion of knowledge on the regulation of the cellulolytic system of filamentous fungi and the application of this knowledge to the improvement of numerous bioproducts. Engineering TFs and promoters may yield more efficient strains that may be active in plant biomass hydrolysis. In this way, the technological processes for obtaining ethanol from lignocellulose may become more commercially viable.

09 BIOMASS FUELS↗
DOE OSTI2021

Introduction of NADH-dependent nitrate assimilation in Synechococcus sp. PCC 7002 improves photosynthetic production of 2-methyl-1-butanol and isobutanol

Cyanobacteria hold promise for renewable chemical production due to their photosynthetic nature, but engineered strains frequently display poor production characteristics. These difficulties likely arise in part due to the distinctive photoautotrophic metabolism of cyanobacteria. Here, in this work, we apply a genome-scale metabolic model of the cyanobacteria Synechococus sp. PCC 7002 to identify strain designs accounting for this unique metabolism that are predicted to improve the production of various biofuel alcohols (e.g. 2-methyl-1-butanol, isobutanol, and 1-butanol) synthesized via an engineered biosynthesis pathway. Using the model, we identify that the introduction of a large, non-native NADH-demand into PCC 7002's metabolic network is predicted to enhance production of these alcohols by promoting NADH-generating reactions upstream of the production pathways. To test this, we construct strains of PCC 7002 that utilize a heterologous, NADH-dependent nitrite reductase in place of the native, ferredoxin-dependent enzyme to create an NADH-demand in the cells when grown on nitrate-containing media. We find that photosynthetic production of both isobutanol and 2-methyl-1-butanol is significantly improved in the engineered strain background relative to that in a wild-type background. We additionally identify that the use of high-nutrient media leads to a substantial prolongment of the production curve in our alcohol production strains. The metabolic engineering strategy identified and tested in this work presents a novel approach to engineer cyanobacterial production strains that takes advantage of a unique aspect of their metabolism and serves as a basis on which to further develop strains with improved production of these alcohols and related products.

2-methyl-1-butanol↗
DOE OSTI2014

Introduction of a Synthetic CO 2 -fixing Photorespiratory Bypass into a Cyanobacterium

Global photosynthetic productivity is limited by the enzymatic assimilation of CO 2 into organic carbon compounds. Ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO), the carboxylating enzyme of the Calvin-Benson cycle, poorly discriminates between CO 2 and O 2 , leading to photorespiration and the loss of fixed carbon and nitrogen. With the advent of synthetic biology, it is now feasible to design, synthesize, and introduce biochemical pathways in vivo. We engineered a synthetic photorespiratory bypass based on the 3-hydroxypropionate bi-cycle into the model cyanobacterium, Synechococcus elongatus sp. PCC 7942. The heterologously expressed cycle is designed to function as both a photorespiratory bypass and an additional CO 2 -fixing pathway, supplementing the Calvin-Benson cycle. We demonstrate the function of all six introduced enzymes and identify bottlenecks to be targeted in subsequent bioengineering. These results have implications for efforts to improve photosynthesis and for the “green” production of high value products of biotechnological interest.

3-Hydroxypropionate Bi-cycle↗
DOE OSTI2020

Engineering crassulacean acid metabolism (CAM) pathways in plants

Disclosed herein are method of altering CAM pathways in plants. In some examples, a disclosed method includes overexpressing one or more genes encoding one or more enzymes that carry out the basic biochemical sequence of nocturnal CO2 fixation (carboxylation) into C4 acids (malate), store C4 acids in the vacuole of the plant, and/or then decarboxylate and refix the released CO2 by C3 photosynthesis during the subsequent day in a plant cell, thereby altering CAM in the plant cell. Also disclosed herein are isolated polynucleotide sequences, transformation vectors, transgenic plant cells, plant part, and plants. The disclosed methods and compositions can be used to improve the water-use efficiency and drought tolerance and durability of plants, such as in plants in arid environments, and also enhance the ability of plants to perform.

Cushman, John C.↗
DOE OSTI2024

Constructing the Nitrogen Flux Maps (NFMs) of Plants

The main objectives of this project are to construct plant N flux maps (NFMs) from plant genomes and to determine functionality of AT enzymes and plant N metabolic network. To address this grand challenge, this project made use of rapidly growing numbers of plant genomes, high-throughput functional characterization platforms, and computational modeling to deduce both biochemical and systems level functionality of ATs and NFMs. The obtained NFMs will provide a novel framework to advance basic understanding of plant N metabolism and facilitate rational engineering of plants with high productivity even under limited N input.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2016

Synthetic and systems biology for microbial production of commodity chemicals

The combination of synthetic and systems biology is a powerful framework to study fundamental questions in biology and produce chemicals of immediate practical application such as biofuels, polymers, or therapeutics. However, we cannot yet engineer biological systems as easily and precisely as we engineer physical systems. In this review, we describe the path from the choice of target molecule to scaling production up to commercial volumes. We present and explain some of the current challenges and gaps in our knowledge that must be overcome in order to bring our bioengineering capabilities to the level of other engineering disciplines. Challenges start at molecule selection, where a difficult balance between economic potential and biological feasibility must be struck. Pathway design and construction have recently been revolutionized by next-generation sequencing and exponentially improving DNA synthesis capabilities. Although pathway optimization can be significantly aided by enzyme expression characterization through proteomics, choosing optimal relative protein expression levels for maximum production is still the subject of heuristic, non-systematic approaches. Toxic metabolic intermediates and proteins can significantly affect production, and dynamic pathway regulation emerges as a powerful but yet immature tool to prevent it. Host engineering arises as a much needed complement to pathway engineering for high bioproduct yields; and systems biology approaches such as stoichiometric modeling or growth coupling strategies are required. A final, and often underestimated, challenge is the successful scale up of processes to commercial volumes. Sustained efforts in improving reproducibility and predictability are needed for further development of bioengineering.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2026

Elucidating biodegradation of dimethyl terephthalate by two Rhodococcus strains for its valorization applications

Dimethyl terephthalate (DMT) serves as the precursor in the production of polyethylene terephthalate, polytrimethylene terephthalate, and polybutylene terephthalate. The widespread use of DMT in the polymer industry and its ubiquitous existence in end products raise alarms about its potential harm to humans and animals. DMT can enter the environment through the degradation of polymers and their end products, and cause endocrine disruption, oxidative stress, and an elevated risk of cancer. In recent years, DMT has also gained renewed interest in its potential for plastic recycling and upcycling. In this study, we identified two strains of Rhodococcus that possess DMT-degrading capabilities and utilized transcriptomic analysis and gene knockout to elucidate the mechanisms of DMT degradation. R. opacus PD630 and R. jostii RPET were found to convert up to 1 g/L DMT into mono-methyl terephthalate (MMT). A putative DMTase (RS34275) was identified for this conversion. R. jostii RPET also demonstrates the ability to convert DMT into MMT and to utilize MMT for its cellular growth via the terephthalate pathway. A putative MMTase (RS21885) as the sole enzyme was identified for the conversion of MMT into terephthalate in the RPET strain. In addition, we successfully produced lycopene and lipids from an engineered RPET strain using DMT as a substrate. Our findings will facilitate future DMT bioremediation and bio-upcycling of DMT-associated plastics, enabling the production of value-added products.

Biodegradation↗
DOE OSTI2025

A highly active Burkholderia polyketoacyl-CoA thiolase for production of triacetic acid lactone

Triacetic acid lactone (TAL) is a versatile platform chemical traditionally biosynthesized via decarboxylative Claisen condensation by 2-pyrone synthase. However, this route is limited by poor efficiency and dependence on malonyl-CoA. Here, we show that non-decarboxylative Claisen condensation by polyketoacyl-CoA thiolases offers a more efficient alternative. Through mining homologs of a previously reported enzyme from Cupriavidus necator, we identify five thiolases with TAL production activity. One candidate, BktBbr from Burkholderia sp. RF2-non_BP3, exhibits approximately 30-fold higher activity in vitro and supports 30-fold higher TAL titers in Escherichia coli compared to the original enzyme. Fed-batch fermentation achieves titers up to 2.8 g L⁻¹. Structural analysis of BktBbr co-crystallized with CoA esters guides rational engineering to further enhance performance. Our discovery of a highly active thiolase establishes an alternative enzymatic route to produce TAL efficiently, providing a scalable foundation for sustainable biomanufacturing.

Wang, Zilong [Joint BioEnergy Institute (JBEI), Em↗
DOE OSTI2022

Comparative and pangenomic analysis of the genus Streptomyces

Abstract Streptomycetes are highly metabolically gifted bacteria with the abilities to produce bioproducts that have profound economic and societal importance. These bioproducts are produced by metabolic pathways including those for the biosynthesis of secondary metabolites and catabolism of plant biomass constituents. Advancements in genome sequencing technologies have revealed a wealth of untapped metabolic potential from Streptomyces genomes. Here, we report the largest Streptomyces pangenome generated by using 205 complete genomes. Metabolic potentials of the pangenome and individual genomes were analyzed, revealing degrees of conservation of individual metabolic pathways and strains potentially suitable for metabolic engineering. Of them, Streptomyces bingchenggensis was identified as a potent degrader of plant biomass. Polyketide, non-ribosomal peptide, and gamma-butyrolactone biosynthetic enzymes are primarily strain specific while ectoine and some terpene biosynthetic pathways are highly conserved. A large number of transcription factors associated with secondary metabolism are strain-specific while those controlling basic biological processes are highly conserved. Although the majority of genes involved in morphological development are highly conserved, there are strain-specific varieties which may contribute to fine tuning the timing of cellular differentiation. Overall, these results provide insights into the metabolic potential, regulation and physiology of streptomycetes, which will facilitate further exploitation of these important bacteria.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2012

In planta production of the highly potent resveratrol analogue pterostilbene via stilbene synthase and O-methyltransferase co-expression

Resveratrol and related stilbenes are thought to play important roles in defence responses in several plant species and have also generated considerable interest as nutraceuticals owing to their diverse health-promoting properties. Pterostilbene, a 3,5-dimethylether derivative of resveratrol, possesses properties similar to its parent compound and, additionally, exhibits significantly higher fungicidal activity in vitro and superior pharmacokinetic properties in vivo. Recombinant enzyme studies carried out using a previously characterized O-methyltransferase sequence from Sorghum bicolor (SbOMT3) demonstrated its ability to catalyse the A ring-specific 3,5-bis-O-methylation of resveratrol, yielding pterostilbene. A binary vector was constructed for the constitutive co-expression of SbOMT3 with a stilbene synthase sequence from peanut (AhSTS3) and used for the generation of stably transformed tobacco and Arabidopsis plants, resulting in the accumulation of pterostilbene in both species. A reduced floral pigmentation phenotype observed in multiple tobacco transformants was further investigated by reversed-phase HPLC analysis, revealing substantial decreases in both dihydroquercetin-derived flavonoids and phenylpropanoid-conjugated polyamines in pterostilbene-producing SbOMT3/AhSTS3 events. These results demonstrate the potential utility of this strategy for the generation of pterostilbene-producing crops and also underscore the need for the development of additional approaches for minimizing concomitant reductions in key phenylpropanoid-derived metabolites.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Experimentally Validated Reconstruction and Analysis of a Genome-Scale Metabolic Model of an Anaerobic Neocallimastigomycota Fungus

Recent genomic analyses have revealed that anaerobic gut fungi possess both the largest number and highest diversity of lignocellulolytic enzymes of all sequenced fungi, explaining their ability to decompose lignocellulosic substrates, e.g., agricultural waste, into fermentable sugars. Despite their potential, the development of engineering methods for these organisms has been slow due to their complex life cycle, understudied metabolism, and challenging anaerobic culture requirements.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2016

Applications of Carboxylic Acid Reductases in Oleaginous Microbes

Carboxylic acid reductases (CARs) are recently emerging reductive enzymes for the direct production of aldehydes from biologically-produced carboxylic acids. Recent work has demonstrated that these powerful enzymes are able to reduce a very broad range of volatile- to long-chain fatty acids as well as aromatic acids. Here, we express four CAR enzymes from different fungal origins to test their activity against fatty acids commonly produced in oleaginous microbes. These in vitro results will inform metabolic engineering strategies to conduct mild biological reduction of carboxylic acids in situ, which is conventionally done via hydrotreating catalysis at high temperatures and hydrogen pressures.

09 BIOMASS FUELS↗