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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 181 records · Page 10
All sourcesNASA NTRSDOE OSTI
DOE OSTI2025

Microbial secondary metabolites: advancements to accelerate discovery towards application

Microbial secondary metabolites not only have key roles in microbial processes and relationships but are also valued in various sectors of today’s economy, especially in human health and agriculture. The advent of genome sequencing has revealed a previously untapped reservoir of biosynthetic capacity for secondary metabolites indicating that there are new biochemistries, roles and applications of these molecules to be discovered. New predictive tools for biosynthetic gene clusters (BGCs) and their associated pathways have provided insights into this new diversity. Advanced molecular and synthetic biology tools and workflows including cell-based and cell-free expression facilitate the study of previously uncharacterized BGCs, accelerating the discovery of new metabolites and broadening our understanding of biosynthetic enzymology and the regulation of BGCs. These are complemented by new developments in metabolite detection and identification technologies, all of which are important for unlocking new chemistries that are encoded by BGCs. This renaissance of secondary metabolite research and development is catalysing toolbox development to power the bioeconomy.

Dinglasan, Jaime Lorenzo N↗
DOE OSTI2022

Carbon-negative production of acetone and isopropanol by gas fermentation at industrial pilot scale

Many industrial chemicals that are produced from fossil resources could be manufactured more sustainably through fermentation. In this work, we describe the development of a carbon-negative fermentation route to producing the industrially important chemicals acetone and isopropanol from abundant, low-cost waste gas feedstocks, such as industrial emissions and syngas. Using a combinatorial pathway library approach, we first mined a historical industrial strain collection for superior enzymes that we used to engineer the autotrophic acetogen Clostridium autoethanogenum. Next, we used omics analysis, kinetic modeling and cell-free prototyping to optimize flux. Finally, we scaled-up our optimized strains for continuous production at rates of up to ~3 g/L/h and ~90% selectivity. Life cycle analysis confirmed a negative carbon footprint for the products. Unlike traditional production processes, which result in release of greenhouse gases, our process fixes carbon. These results show that engineered acetogens enable sustainable, high-efficiency, high-selectivity chemicals production. We expect that our approach can be readily adapted to a wide range of commodity chemicals.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Growth-uncoupled propanediol production in a Thermoanaerobacterium thermosaccharolyticum strain engineered for high ethanol yield

Abstract Cocultures of engineered thermophilic bacteria can ferment lignocellulose without costly pretreatment or added enzymes, an ability that can be exploited for low cost biofuel production from renewable feedstocks. The hemicellulose-fermenting species Thermoanaerobacterium thermosaccharolyticum was engineered for high ethanol yield, but we found that the strains switched from growth-coupled production of ethanol to growth uncoupled production of acetate and 1,2-propanediol upon growth cessation, producing up to 6.7 g/L 1,2-propanediol from 60 g/L cellobiose. The unique capability of this species to make 1,2-propanediol from sugars was described decades ago, but the genes responsible were not identified. Here we deleted genes encoding methylglyoxal reductase, methylglyoxal synthase and glycerol dehydrogenase. Deletion of the latter two genes eliminated propanediol production. To understand how carbon flux is redirected in this species, we hypothesized that high ATP levels during growth cessation downregulate the activity of alcohol and aldehyde dehydrogenase activities. Measurements with cell free extracts show approximately twofold and tenfold inhibition of these activities by 10 mM ATP, supporting the hypothesized mechanism of metabolic redirection. This result may have implications for efforts to direct and maximize flux through alcohol dehydrogenase in other species.

09 BIOMASS FUELS↗
DOE OSTI2022

Structural features of Dnase1L3 responsible for serum antigen clearance

Autoimmunity develops when extracellular DNA released from dying cells is not cleared from serum. While serum DNA is primarily digested by Dnase1 and Dnase1L3, Dnase1 cannot rescue autoimmunity arising from Dnase1L3 deficiencies. Dnase1L3 uniquely degrades antigenic forms of cell-free DNA, including DNA complexed with lipids and proteins. The distinct activity of Dnase1L3 relies on its unique C-terminal Domain (CTD), but the mechanism is unknown. We used multiple biophysical techniques and functional assays to study the interplay between the core catalytic domain and the CTD. While the core domain resembles Dnase1, there are key structural differences between the two enzymes. First, Dnase1L3 is not inhibited by actin due to multiple differences in the actin recognition site. Second, the CTD augments the ability of the core to bind DNA, thereby facilitating the degradation of complexed DNA. Together, these structural insights will inform the development of Dnase1L3-based therapies for autoimmunity.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Monitoring the compaction of single DNA molecules in Xenopus egg extract in real time

DNA compaction is required for the condensation and resolution of chromosomes during mitosis, but the relative contribution of individual chromatin factors to this process is poorly understood. We developed a physiological, cell-free system using high-speed Xenopus egg extracts and optical tweezers to investigate real-time mitotic chromatin fiber formation and force-induced disassembly on single DNA molecules. Compared to interphase extract, which compacted DNA by ~60%, metaphase extract reduced DNA length by over 90%, reflecting differences in whole-chromosome morphology under these two conditions. Depletion of the core histone chaperone ASF1, which inhibits nucleosome assembly, decreased the final degree of metaphase fiber compaction by 29%, while depletion of linker histone H1 had a greater effect, reducing total compaction by 40%. Compared to controls, both depletions reduced the rate of compaction, led to more short periods of decompaction, and increased the speed of force-induced fiber disassembly. In contrast, depletion of condensin from metaphase extract strongly inhibited fiber assembly, resulting in transient compaction events that were rapidly reversed under high force. Altogether, these findings support a speculative model in which condensin plays the predominant role in mitotic DNA compaction, while core and linker histones act to reduce slippage during loop extrusion and modulate the degree of DNA compaction.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

The first DEP domain of the RhoGEF P-Rex1 autoinhibits activity and contributes to membrane binding

Phosphatidylinositol (3,4,5)-trisphosphate (PIP 3 )-dependent Rac exchanger 1 (P-Rex1) catalyzes the exchange of GDP for GTP on Rac GTPases, thereby triggering changes in the actin cytoskeleton and in transcription. Its overexpression is highly correlated with the metastasis of certain cancers. P-Rex1 recruitment to the plasma membrane and its activity are regulated via interactions with heterotrimeric Gβγ subunits, PIP 3 , and protein kinase A (PKA). Deletion analysis has further shown that domains C-terminal to its catalytic Dbl homology (DH) domain confer autoinhibition. Among these, the first dishevelled, Egl-10, and pleckstrin domain (DEP1) remains to be structurally characterized. DEP1 also harbors the primary PKA phosphorylation site, suggesting that an improved understanding of this region could substantially increase our knowledge of P-Rex1 signaling and open the door to new selective chemotherapeutics. Here we show that the DEP1 domain alone can autoinhibit activity in context of the DH/PH-DEP1 fragment of P-Rex1 and interacts with the DH/PH domains in solution. The 3.1 Å crystal structure of DEP1 features a domain swap, similar to that observed previously in the Dvl2 DEP domain, involving an exposed basic loop that contains the PKA site. Using purified proteins, we show that although DEP1 phosphorylation has no effect on the activity or solution conformation of the DH/PH-DEP1 fragment, it inhibits binding of the DEP1 domain to liposomes containing phosphatidic acid. Thus, we propose that PKA phosphorylation of the DEP1 domain hampers P-Rex1 binding to negatively charged membranes in cells, freeing the DEP1 domain to associate with and inhibit the DH/PH module.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2021

Phage display and other peptide display technologies

ABSTRACT Phage display technology, which is based on the presentation of peptide sequences on the surface of bacteriophage virions, was developed over 30 years ago. Improvements in phage display systems have allowed us to employ this method in numerous fields of biotechnology, as diverse as immunological and biomedical applications, the formation of novel materials and many others. The importance of phage display platforms was recognized by awarding the Nobel Prize in 2018 ‘for the phage display of peptides and antibodies’. In contrast to many review articles concerning specific applications of phage display systems published in recent years, we present an overview of this technology, including a comparison of various display systems, their advantages and disadvantages, and examples of applications in various fields of science, medicine and the broad sense of biotechnology. Other peptide display technologies, which employ bacterial, yeast and mammalian cells, as well as eukaryotic viruses and cell-free systems, are also discussed. These powerful methods are still being developed and improved; thus, novel sophisticated tools based on phage display and other peptide display systems are constantly emerging, and new opportunities to solve various scientific, medical and technological problems can be expected to become available in the near future.

Jaroszewicz, Weronika (ORCID:0000000302948682)↗
DOE OSTI2022

A multi-plex protein expression system for production of complex enzyme formulations in Trichoderma reesei

Abstract Heterologous protein production has been challenging in the hyper-cellulolytic fungus, Trichoderma reesei as the species is known for poor transformation efficiency, low homologous recombination frequency, and marginal screening systems for the identification of successful transformants. We have applied the 2A-peptide multi-gene expression system to co-express four proteins, which include three cellulases: a cellobiohydrolase (CBH1), an endoglucanase (EG1), and a β-D-glucosidase (BGL1), as well as the enhanced green fluorescent protein (eGFP) marker protein. We designed a new chassis vector, pTrEno-4X-2A, for this work. Expression of these cellulase enzymes was confirmed by real-time quantitative reverse transcription PCR and immunoblot analysis. The activity of each cellulase was assessed using chromogenic substrates, which confirmed the functionality of the enzymes. Expression and activity of these enzymes were proportional to the level of eGFP fluorescence, thereby validating the reliability of this screening technique. An 18-fold differencein protein expression was observed between the first and third genes within the 2A-peptide construct. The availability of this new multi-gene expression and screening tool is expected to greatly impact multi-enzyme applications, such as the production of complex commercial enzyme formulations and metabolic pathway enzymes, especially those destined for cell-free applications.

09 BIOMASS FUELS↗
DOE OSTI2021

Engineering Forisome Scaffolds: Elucidating Spatial Self-Assembly Patterning of Bio-inorganic Complexes

Organisms can synthesize biomaterials incorporating an array of naturally occurring elements while overcoming challenges and insults. Although, it is known that most cellular biomaterials are synthesized in specialized cellular compartments, there are knowledge gaps about how organic/inorganic biomaterial synthesis is orchestrated inside cells. In addition, there is great potential in understanding how individual monomers can self-assembly into organized patterns to form responsive biomaterials. Forisomes are a natural responsive biomaterial found in legume plants that serve as a plug sieve element in the plant phloem that undergo anisotropic conformational changes by rapid (<1 s) ATP-independent from condensed spindle to plug-like form, triggered by the influx of Ca 2+ . Addressing principles of forisome synthesis and assembly will determine how biomaterials containing inorganic elements self-assemble and conduct chemical modification to produce biomaterials or undergo biomineralization. We employ transcription and translation (TXTL) using cell-free expression systems for forisome monomer expression, self-assembly, and pattern probing. We conducted experiments to precisely control forisome proteins synthesis of various monomers SEO1, SEO2, SEO3, and SEO4 to explore self- assembly. We demonstrate forisome self-assembly of the SEO monomers is possible and indicate unique monomer fluorescent labeling patterns that require additional analysis. We investigated locations and linkers for adding tetracysteine tag fluorophore probes to determine impacts of self-assembly and anisotropic conformational changes.

42 ENGINEERING↗
DOE OSTI2024

Disruptive Technology for Carbon Negative Commodity Chemicals

This project was designed to develop an economically attractive process for the carbon negative production of the commodity biochemical succinic acid. To compete against petroleum derived biochemicals, the process must deliver high raw material conversion efficiency and high volumetric productivity to reduce both recurring and capital-related costs, respectively. The proposed process introduces additional electrons sourced from hydrogen into the biosynthetic pathway so that the carbon from two CO2 molecules can be combined with each glucose to significantly increase succinic acid yields. The process also uses ultrafiltration to continuously remove the product from the bioreactor to avoid product inhibition and extend the productive life of the cell extract. Genetic deletions and specific enzyme removal during extract preparation more efficiently direct both atomic and electronic resources toward product formation. A preliminary technoeconomic analysis indicates that these process innovations made possible by cell-free production will enable large scale production with attractive ROI and profitability. This two-year project provided some 29 distinct insights, analytical advances, and process improvements that resulted in a demonstration of process feasibility. However, an estimated two to three years of additional development will be required to achieve convincing pilot scale demonstrations that will motivate large scale investments.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2023

Perspectives on biorefineries in microbial production of fuels and chemicals

Microbes drive our complex biosphere by regulating the global ecosystem through cycling elements and energy. Humankind has barely begun leveraging this biotransformation capacity to impact global economies and ecologies. Advances in genetic engineering, molecular analysis, metabolic flux modeling, microbial consortia/biome mapping and engineering, cell-free bioproduction, artificial intelligence/machine learning and the ever expanding -omics frontiers have set the stage for paradigm changes to how humankind produces, uses, transforms, and recycles carbon and energy through microbes. Harnessing this enormous potential could drive a global bioeconomy and manage carbon at a planetary level but requires understanding and application at a grand scale across a broad range of science and engineering disciplines. The penultimate manifestation of these advances is the “bio-refinery”, which is often referenced, but is a long way from being fully developed as a global carbon management platform. Broadening the feed stocks, processing operations, and product portfolio to a sequential cascade optimizing the conversion as a whole instead of limited outputs could greatly advance deployment and stability of a bioeconomy.

09 BIOMASS FUELS↗
DOE OSTI2025

Purification and expression of a novel bacteriocin, JUQZ-1, against Pseudomonas syringae pv. Actinidiae (PSA), secreted by Brevibacillus laterosporus Wq-1, isolated from the rhizosphere soil of healthy kiwifruit

Kiwifruit canker, caused by Pseudomonas syringae pv. actinidiae (PSA), has led to significant losses in the kiwifruit industry each year. Due to the drug resistance feature of PSA, biological control is currently the most promising method. Developing biocontrol bacteria against PSA could help solve the issue of drug resistance generated during the chemical control of PSA to a certain extent. In this research, a Wq-1 strain that demonstrated excellent inhibitory activity against PSA was isolated from the rhizosphere soil of healthy kiwifruit. Based on the morphological characteristics and phylogenetic analysis of the 16S rRNA gene sequence, the isolated strain was identified as Brevibacillus laterosporus Wq-1. Bacteriostatic proteins were isolated from the cell-free culture filtrate of strain Wq-1 and were found to have a molecular weight of approximately 12 kDa, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Liquid chromatography–tandem mass spectrometry (LC–MS/MS) detection revealed that there were several peptides in the target band that were consistent with protein 01021 in the genome. The gene of the 01021 protein was cloned into the plasmid pPICZa, and the recombinant bacteriocin was successfully expressed using the Pichia pastoris X33 expression system. The recombinant protein 01021 effectively inhibited the growth of PSA. This is the first report of the protein’s antimicrobial activity, distinguishing it from previously identified bacteriocins. Therefore, we named this bacteriocin JUQZ-1. In addition, our results showed that the protein JUQZ-1 not only exhibited a broad bacteriostatic spectrum but also high thermal and pH stability suitable for harsh environmental conditions., JUQZ-1, a protein with antimicrobial properties and strong environmental tolerance, may serve as a promising alternative to antibiotics.

Shuai, Yang↗
DOE OSTI2023

Simple Bioparticle Filtration Device Based on an Ultralow-Fouling Zwitterionic Polyurethane Membrane for Rapid Large-Volume Separation of Plasma and Viruses from Whole Blood

Plasma separation from whole blood is oftent required as an essential first step when performing blood tests with a viral assay. However, developing a point-of-care plasma extraction device with a large output and high virus recovery remains a significant obstacle to the success of on-site viral load tests. Here, we report a portable, easy-to-use, cost-efficient, membrane-filtration-based plasma separation device that enables rapid large-volume plasma extraction from whole blood, designed for point-of-care virus assays. The plasma separation is realized by a low-fouling zwitterionic polyurethane-modified cellulose acetate (PCBU-CA) membrane. The zwitterionic coating on the cellulose acetate membrane can decrease surface protein adsorption by 60% and increase plasma permeation by 46% compared with a pristine membrane. The PCBU-CA membrane, with its ultralow-fouling properties, enables rapid plasma separation. The device can yield a total of 1.33 mL plasma from 10 mL whole blood in 10 min. The extracted plasma is cell-free and exhibits a low hemoglobin level. In addition, our device demonstrated a 57.8% T7 phage recovery in the separated plasma. The results of real-time polymerase chain reaction analysis confirmed that the nucleic acid amplification curve of the plasma extracted by our device is comparable to that obtained by centrifugation. With its high plasma yield and good phage recovery, our plasma separation device provides an excellent replacement for traditional plasma separation protocols for point-of-care virus assays and a broad spectrum of clinical tests.

36 MATERIALS SCIENCE↗
NASA NTRS1972

Thermolabile triose phosphate isomerase in a psychrophilic Clostridium.

It was found that a psychrophilic Clostridium contains a triose phosphate isomerase which is very labile at moderate temperatures. An investigation showed that the optimal growth temperature of the psychrophile was between 15 and 20 deg C. No growth occurred at 25 deg C. The thermostability of the glycolytic enzymes in the cell-free extracts of Clostridium sp. strain 69 was studied. The data obtained show that the triose phosphate isomerase is quite labile at moderate temperatures. The instability of the enzyme is sufficient to explain the low maximum growth temperature of the psychrophile.

Shing, Y. W.↗
NASA NTRS1979

Seasat scanning multichannel microwave radiometer - Results of the Gulf of Alaska workshop

Scanning multichannel microwave radiometer results obtained by the Gulf of Alaska Seasat Experiment Workshop are reported. The Seasat SMMR provided data from five channels operating at 6.6, 10.7, 18, 21, and 37 GHz at vertical and horizontal polarizations. Two preliminary algorithms were used to retrieve geophysical parameters from the data: the Wentz algorithm (Bierman et al., 1978) based on a theoretically derived function for computing brightness temperatures and the Wilheit algorithm, based on statistical relationships between brightness temperatures and the geophysical parameters obtained from an ensemble of realistic sea-surface temperature values, wind speeds, atmospheric temperature profiles, water vapor profiles and cloud models. In spite of the immaturity of the data-processing algorithms, results are encouraging. For open ocean, rain-free cells of high-quality surface truth wind determinations display standard deviations of 3 m/sec about a bias of 1.5 m/sec. The sea-surface temperature exhibits a standard deviation of about 1.5 deg C about a bias of 3 to 5 deg C under a variety of meteorological conditions.

Lipes, R. G.↗
NASA NTRS1982

Atrophy of rat skeletal muscles in simulated weightlessness

A hypokinetic rat model was used for elucidation of the mechanism of skeletal muscle wasting which occurs in weightlessness. Rats were suspended from a back-harness with the head tilted downward and the hind limbs totally unloaded. A progressive decrease in the size of the soleus muscle from suspended rats was observed as a function of time. The rate of protein degradation of the homogenates from the soleus muscles of suspended and control animals was not significantly different. The rate of cell-free protein synthesis was severely repressed in the atrophied muscle. An initial rise in the levels of plasma glucose and corticosterone was observed on the second day of suspension, but they subsequently returned to normal values.

Feller, D. D.↗