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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 19 records
All sourcesNASA NTRSDOE OSTI
DOE OSTI2021

Deep representation learning improves prediction of LacI-mediated transcriptional repression

Significance The understanding of protein function increases with new experimental and evolutionary datasets. A major challenge is to apply machine learning to these datasets to capture essential features of protein function. Here, we analyze the experimentally determined repression function for tens of thousands of mutants of the LacI protein. This study provides a continuous, noncategorical repression value across a majority of all single mutations and for thousands of higher-order mutations. To develop a top-performing model for the prediction of repression by LacI, we compare several leading variant effect prediction algorithms. A deep representation learning paradigm, first trained across millions of proteins from all known protein families and then fine-tuned using LacI experimental data, offers the highest predictive performance of repression function.

42 ENGINEERING↗
DOE OSTI2020

Materials Data on LaCI by Materials Project

LaCI crystallizes in the monoclinic C2/m space group. The structure is two-dimensional and consists of one LaCI sheet oriented in the (0, 0, 1) direction. La3+ is bonded in a 8-coordinate geometry to five equivalent C2- and three equivalent I1- atoms. There are a spread of La–C bond distances ranging from 2.53–2.89 Å. There are two shorter (3.28 Å) and one longer (3.38 Å) La–I bond lengths. C2- is bonded in a 6-coordinate geometry to five equivalent La3+ and one C2- atom. The C–C bond length is 1.33 Å. I1- is bonded in a 3-coordinate geometry to three equivalent La3+ atoms.

36 MATERIALS SCIENCE↗
DOE OSTI2021

The genotype-phenotype landscape of an allosteric protein

Allostery is a fundamental biophysical mechanism that underlies cellular sensing, signaling, and metabolism. Yet a quantitative understanding of allosteric genotype-phenotype relationships remains elusive. Here, we report the large-scale measurement of the genotype-phenotype landscape for an allosteric protein: the lac repressor from Escherichia coli , LacI. Using a method that combines long-read and short-read DNA sequencing, we quantitatively measure the dose-response curves for nearly 10 5 variants of the LacI genetic sensor. The resulting data provide a quantitative map of the effect of amino acid substitutions on LacI allostery and reveal systematic sequence-structure-function relationships. We find that in many cases, allosteric phenotypes can be quantitatively predicted with additive or neural-network models, but unpredictable changes also occur. For example, we were surprised to discover a new band-stop phenotype that challenges conventional models of allostery and that emerges from combinations of nearly silent amino acid substitutions.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Innovative Pathways: Transforming Impact Investments to Working Capital For Early Stage Cleantech Companies: Los Angeles Cleantech Incubator (January 1, 2020 to March 31, 2020)

Project Objectives: The focus for the entirety of the last quarter was on the completion of the pilot loan funding within our portfolio, and the closing out of our DOE-CAP Grant: “Innovative Pathways: Transforming Impact Investments to Working Capital for Early Stage Cleantech Companies.” The fourteenth quarter (Q14) of the program performance was also dedicated toward two main activities: the continued fundraising efforts for a formalized LACI debt fund, as well as finalization of the lending processes for pilot underwriting amongst the LACI Team and with Mission Driven Finance. This quarter marked key successes around the workflows of the DOE-CAP grant, specifically around fund capitalization efforts. Activities during the thirteenth quarter included: 1. Finalization of the DOE-CAP grant activities and work-flows; 2. Due diligence, underwriting and issuance of one (1) loan in Q1 2021 and three (3) in Q4 2020 for technology providers in grant-funded zero emissions mobility & community pilots, in partnership with LACI and local organizations, totaling ten (10) total loans; 3. Development of fundraising and marketing materials, and distribution to 30+ possible funding sources over the course of Q13 & Q14; and 4. Reinvigoration of conversations with Mission Driven Finance, the prospective underwriting partner for key activities and governance documents, and finalization of all loan processing activities once the fund is capitalized

29 ENERGY PLANNING, POLICY, AND ECONOMY↗
DOE OSTI2024

Development of inducible promoters for regulating gene expression in Clostridium tyrobutyricum for biobutanol production

Abstract Clostridium tyrobutyricumis an anaerobe known for its ability to produce short‐chain fatty acids, alcohols, and esters. We aimed to develop inducible promoters for fine‐tuning gene expression inC. tyrobutyricum. Synthetic inducible promoters were created by employing anEscherichia coli lacoperator to regulate the thiolase promoter (PCathl) fromClostridium acetobutylicum, with the best one (LacI‐Pto4s) showing a 5.86‐fold dynamic range with isopropyl β‐d‐thiogalactoside (IPTG) induction. A LT‐Pt7 system with a dynamic range of 11.6‐fold was then created by combining LacI‐Pto4s with a T7 expression system composing of RNA polymerase (T7RNAP) and Pt7lacpromoter. Furthermore, two inducible expression systems BgaR‐PbgaLA and BgaR‐PbgaLB with a dynamic range of ~40‐fold were developed by optimizing a lactose‐inducible expression system fromClostridium perfringenswith modified 5′ untranslated region (5′ UTR) and ribosome‐binding site (RBS). BgaR‐PbgaLB was then used to regulate the expressions of a bifunctional aldehyde/alcohol dehydrogenase encoded byadhE2 and butyryl‐CoA/acetate Co‐A transferase encoded bycat1 inC. tyrobutyricumwild type and Δcat1::adhE2, respectively, demonstrating its efficient inducible gene regulation. The regulatedcat1 expression also confirmed that the Cat1‐catalyzed reaction was responsible for acetate assimilation inC. tyrobutyricum. The inducible promoters offer new tools for tuning gene expression inC. tyrobutyricumfor industrial applications.

Biotechnology & Applied Microbiology↗
DOE OSTI2025

The Shape Factor for Pits and Its Impact on Pit Stability

This study employs the finite element method (FEM) to predict the impact of pit shape on pit stability via shape factors of various pit geometries relevant to localized corrosion. Lower values of the shape factor indicate an increased ease in maintaining pit stability. Analyzed geometries include undercut pits, bispherical pit-within-pit structures, and covered pits with perforated (lacy) covers. The effect of the water layer thickness, transport properties of the electrolyte inside and outside the pit, and cathode location on the pit shape factor were also explored. Results show that occluded pits exhibit lower shape factors than open ones, with disk-shaped pits decreasing further as c/r ratio and occlusion angle increase. The findings also suggest minimal influence from a secondary pit if the primary remains active. An equation is presented that quantifies the impact of lacy covers, revealing significant shape factor value reduction. Additionally, high salt concentrations inside pits have a limited stabilizing effect compared to geometry, while thin water layers and adjacent cathodic/sinks reduce pit stability.

Shehi, A. (ORCID:0009000271548041)↗
DOE OSTI2025

Leveraging a Southern California Energy Innovation Cluster to Pilot and Validate Emerging Energy Technologies

The EPIC Pilot Program, led by the Los Angeles Cleantech Incubator (LACI), was designed to help early-stage clean technology startups accelerate technical validation, investment readiness, and market entry. The project focused on supporting startups in designing and deploying small-scale pilots with the guidance of EPIC Partners, a network of regional stakeholders in the Southern California energy ecosystem, including but not limited to: the Los Angeles Department of Water and Power (LADWP), the California Energy Commission (CEC), Los Angeles County Metropolitan Transportation Authority (LA Metro), and Edison International, among others. Through mentorship, funding, and site access for technology deployments, the program provided a structured pathway for startups to test and refine their technologies in real-world settings.

14 SOLAR ENERGY↗
DOE OSTI2021

Multiplexed characterization of rationally designed promoter architectures deconstructs combinatorial logic for IPTG-inducible systems

Abstract A crucial step towards engineering biological systems is the ability to precisely tune the genetic response to environmental stimuli. In the case of Escherichia coli inducible promoters, our incomplete understanding of the relationship between sequence composition and gene expression hinders our ability to predictably control transcriptional responses. Here, we profile the expression dynamics of 8269 rationally designed, IPTG-inducible promoters that collectively explore the individual and combinatorial effects of RNA polymerase and LacI repressor binding site strengths. We then fit a statistical mechanics model to measured expression that accurately models gene expression and reveals properties of theoretically optimal inducible promoters. Furthermore, we characterize three alternative promoter architectures and show that repositioning binding sites within promoters influences the types of combinatorial effects observed between promoter elements. In total, this approach enables us to deconstruct relationships between inducible promoter elements and discover practical insights for engineering inducible promoters with desirable characteristics.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Xylanolytic metabolism is regulated by coordination of transcription factors XynR and XylR in extremely thermophilic Caldicellulosiruptorales

ABSTRACT Global transcription factors (TFs) control metabolic processes in bacteria to efficiently utilize available carbon. The orderCaldicellulosiruptoraleshas drawn interest due to the ability of its members to degrade components of lignocellulosic biomass. Regulatory reconstruction ofAnaerocellum (f. Caldicellulosiruptor) besciiidentified two major global transcription factors for xylan utilization, XynR and XylR, and the corresponding putative transcription factor binding sites. Recombinant versions of XynR (LacI family) and XylR (ROK family) were subjected to fluorescence polarization (FP) and biolayer interferometry (BLI) analysis to confirm the predicted binding sites. Four XynR sites and two XylR sites were validated, accounting for 20 of 26 genes regulated by XynR and six of seven genes regulated by XylR. Bioinformatic analysis of the individual genes controlled by the two regulators showed an inter-dependent scheme for xylan conversion; the transport of xylooligosaccharides (XOS) is dependent on XylR, while enzymes responsible for hydrolysis are controlled by both regulators. For xylose catabolism by the xylose isomerase-xylulose kinase pathway, regulation is also split, with XylR controlling xylose isomerase and XynR controlling xylokinase. The XynR/XylR regulator pair withinA. besciiis conserved in all sequenced species ofCaldicellulosiruptorales, suggesting similarities in regulating linear xylan conversion. In other xylanolytic thermophiles, XylR homologs control xylan degradation, compared to just 6 out of 26 genes forA. bescii. These results show that two separate regulatory schemes (dual repression) are coordinated byA. besciito effectively regulate the hemicellulose inventory and xylan catabolism. IMPORTANCE To take full advantage of extreme thermophiles as platform metabolic engineering microorganisms, the tools for genetic manipulation must be further developed, and strategies that exploit a better understanding of metabolic regulation need to be discerned.Anaerocellum bescii, the most studied of the extremely thermophilic fermentative anaerobic bacteria that can utilize microcrystalline cellulose, can degrade microcrystalline cellulose and hemicellulose and has been metabolically engineered to convert the resulting sugars to products such as ethanol and acetone. For xylan, in particular, two major global transcription factors (TFs), XynR and XylR, play a role in sugar metabolism, although their predicted regulatory interdependence from bioinformatics analysis has not been elucidated experimentally. Here, fluorescence polarization (FP) and biolayer interferometry (BLI) were used to explore this issue to support metabolic engineering efforts aimed at improving carbohydrate processing to industrial chemicals.

Biotechnology & Applied Microbiology↗
DOE OSTI2021

The ChiS-Family DNA-Binding Domain Contains a Cryptic Helix-Turn-Helix Variant

Sequence-specific DNA-binding domains (DBDs) are conserved in all domains of life. These proteins carry out a variety of cellular functions, and there are a number of distinct structural domains already described that allow for sequence-specific DNA binding, including the ubiquitous helix-turn-helix (HTH) domain. In the facultative pathogen Vibrio cholerae, the chitin sensor ChiS is a transcriptional regulator that is critical for the survival of this organism in its marine reservoir. We recently showed that ChiS contains a cryptic DBD in its C terminus. This domain is not homologous to any known DBD, but it is a conserved domain present in other bacterial proteins. Here, we present the crystal structure of the ChiS DBD at a resolution of 1.28 Å. We find that the ChiS DBD contains an HTH domain that is structurally similar to those found in other DNA-binding proteins, like the LacI repressor. However, one striking difference observed in the ChiS DBD is that the canonical tight turn of the HTH is replaced with an insertion containing a β-sheet, a variant which we term the helix-sheet-helix. Through systematic mutagenesis of all positively charged residues within the ChiS DBD, we show that residues within and proximal to the ChiS helix-sheet-helix are critical for DNA binding. Finally, through phylogenetic analyses we show that the ChiS DBD is found in diverse proteobacterial proteins that exhibit distinct domain architectures. Together, these results suggest that the structure described here represents the prototypical member of the ChiS-family of DBDs.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Infiltration metasomatism of the Allende coarse-grained calcium-aluminum-rich inclusions

We report on the mineralogy, petrography, and O and Al-Mg isotopic systematics of secondary mineralization in the metasomatically altered igneous Ca,Al-rich inclusions (CAIs) [compact type A (CTA), B1, B2, forsterite-bearing B (FoB), and C] from the CV3 carbonaceous chondrite Allende. This alteration affected mainly melilite, and to a lesser degree anorthite, and resulted in the formation of a variety of secondary minerals, including adrianite, Al-diopside, andradite, anorthite, calcite, celsian, clintonite, corundum, dmisteinbergite, ferroan olivine, ferroan monticellite, ferroan Al-diopside, forsterite, grossular, heazlewoodite, hedenbergite, hutcheonite, kushiroite, margarite, monticellite, Na-melilite, nepheline, pentlandite, pyrrhotite, sodalite, spinel, tilleyite, wadalite, and wollastonite. The secondary mineral assemblages are mainly defined by chemical compositions of the primary melilite replaced and elements introduced by an aqueous fluid. Gehlenitic melilite (Åk <35 ) in CTAs and mantles of B1s is mainly replaced by anorthite + grossular; clintonite, corundum, spinel, and Al-diopside are minor. Åkermanitic melilite (Åk 35-90 ) in type B2s, FoBs, and cores of B1s is replaced by the grossular + monticellite + wollastonite, grossular + monticellite, and grossular + Al-diopside assemblages; forsterite, spinel, clintonite, and Na-melilite are minor. In type Cs, lacy melilite (åkermanitic melilite with rounded inclusions of anorthite) is pseudomorphically replaced by the grossular + forsterite + monticellite and grossular + Al-diopside assemblages; Na-melilite is minor. Primary and secondary anorthites in the peripheral portions of CAIs are replaced by nepheline, sodalite, and ferromagnesian olivine. Some CAIs contain voids and cracks filled by andradite, hedenbergite, wollastonite, ±sodalite, ±grossular, ±monticellite, ±tilleyite, and ±calcite. All CAIs studied are surrounded by Wark-Lovering rims, fine-grained matrix-like rims composed of lath-shaped ferroan olivine and abundant nepheline grains, and a layer of salite-hedenbergite pyroxenes + andradite + wollastonite. Grossular associating with monticellite, Al-diopside, and forsterite and replacing åkermanitic melilite ( 27 Al/ 24 Mg ~ 2) has high 27 Al/ 24 Mg ratios (30–100) and shows no resolvable excess of radiogenic 26 Mg (26Mg*). The 27 Al/ 24 Mg ratios (7–10) and 26 Mg* (2–3‰) in the nearly monomineralic grossular veins crosscutting gehlenitic melilite are similar to those of the host melilite and plot along a regression line with 26 Al/ 27 Al ratio of ~5×10 –5 . Oxygen isotopic compositions of secondary minerals in the most Type Bs measured in situ with the UH Cameca ims-1280 and matrix-matched standards plot along mass-dependent fractionation line with Δ 17 O of ~ –3±2‰ with δ 18 O ranging from ~0 to ~10‰. Primary melilite and anorthite in the host CAIs are similarly 16 O-depleted, whereas spinel, forsterite, and most Al,Ti-diopside grains have 16 O-rich compositions (Δ 17 O ~ –25±2‰). Secondary grossular and forsterite in type Cs and type B1 CAI TS-34 show a range of Δ 17 O, from ~ –15 to ~ –1‰; the 16 O-enriched compositions of grossular and forsterite plot along the carbonaceous chondrite anhydrous mineral line. The similar ranges of Δ 17 O and positions on the three-isotope oxygen diagram are observed for primary anorthite; melilite is generally 16 O-depleted compared to anorthite (Δ 17 O ~ –5 to –1±2‰); spinel and fassaite are 16 O-rich (except very Ti-rich fassaite in TS-34 and CTA CAIs). We conclude that Allende CAIs experienced an open-system in situ metasomatic alteration at relatively high temperatures (200-250 °C) in the presence of CO 2 - and H 2 O-bearing fluid with Δ 17 O of ~ –3±2‰ followed by thermal metamorphism at ~ 500 °C on the CV chondrite parent asteroid. During the alteration, most elements were mobile: Si, Na, Cl, K, Fe, S, and Ni were introduced; Al, Ti, Mg, and Ba were locally mobilized; Ca and some Mg and Al were lost from the host inclusions. The alteration occurred after nearly complete decay of 26 Al, >3 Ma after crystallization of CAIs with the canonical ( 26 Al/ 27 Al) 0 of (5.25±0.02)×10 -5 ; 26 Mg* in grossular was inherited from the primary melilite and provide no chronological significance. Oxygen isotopic heterogeneity of primary minerals in the Allende CAIs at least partly is due to isotopic exchange with an aqueous fluid that largely affected melilite, anorthite, perovskite, Zr- and Sc-rich oxides and silicates, and possibly very Ti-rich fassaite.

58 GEOSCIENCES↗
DOE OSTI2021

Genome-Wide Transcription Factor DNA Binding Sites and Gene Regulatory Networks in Clostridium thermocellum

Clostridium thermocellum is a thermophilic bacterium recognized for its natural ability to effectively deconstruct cellulosic biomass. While there is a large body of studies on the genetic engineering of this bacterium and its physiology to-date, there is limited knowledge in the transcriptional regulation in this organism and thermophilic bacteria in general. The study herein is the first report of a large-scale application of DNA-affinity purification sequencing (DAP-seq) to transcription factors (TFs) from a bacterium. We applied DAP-seq to > 90 TFs in C. thermocellum and detected genome-wide binding sites for 11 of them. We then compiled and aligned DNA binding sequences from these TFs to deduce the primary DNA-binding sequence motifs for each TF. These binding motifs are further validated with electrophoretic mobility shift assay (EMSA) and are used to identify individual TFs’ regulatory targets in C. thermocellum . Our results led to the discovery of novel, uncharacterized TFs as well as homologues of previously studied TFs including RexA-, LexA-, and LacI-type TFs. We then used these data to reconstruct gene regulatory networks for the 11 TFs individually, which resulted in a global network encompassing the TFs with some interconnections. As gene regulation governs and constrains how bacteria behave, our findings shed light on the roles of TFs delineated by their regulons, and potentially provides a means to enable rational, advanced genetic engineering of C. thermocellum and other organisms alike toward a desired phenotype.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Light-inducible protein degradation in E. coli with the LOVdeg tag

Molecular tools for optogenetic control allow for spatial and temporal regulation of cell behavior. In particular, light-controlled protein degradation is a valuable mechanism of regulation because it can be highly modular, used in tandem with other control mechanisms, and maintain functionality throughout growth phases. Here, we engineered LOVdeg, a tag that can be appended to a protein of interest for inducible degradation in Escherichia coli using blue light. We demonstrate the modularity of LOVdeg by using it to tag a range of proteins, including the LacI repressor, CRISPRa activator, and the AcrB efflux pump. Additionally, we demonstrate the utility of pairing the LOVdeg tag with existing optogenetic tools to enhance performance by developing a combined EL222 and LOVdeg system. Finally, we use the LOVdeg tag in a metabolic engineering application to demonstrate post-translational control of metabolism. Together, our results highlight the modularity and functionality of the LOVdeg tag system and introduce a powerful new tool for bacterial optogenetics.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Light-inducible protein degradation in E. coli with the LOVdeg tag

Molecular tools for optogenetic control allow for spatial and temporal regulation of cell behavior. In particular, light-controlled protein degradation is a valuable mechanism of regulation because it can be highly modular, used in tandem with other control mechanisms, and maintain functionality throughout growth phases. Here, we engineered LOVdeg, a tag that can be appended to a protein of interest for inducible degradation in Escherichia coli using blue light. We demonstrate the modularity of LOVdeg by using it to tag a range of proteins, including the LacI repressor, CRISPRa activator, and the AcrB efflux pump. Additionally, we demonstrate the utility of pairing the LOVdeg tag with existing optogenetic tools to enhance performance by developing a combined EL222 and LOVdeg system. Finally, we use the LOVdeg tag in a metabolic engineering application to demonstrate post-translational control of metabolism. Together, our results highlight the modularity and functionality of the LOVdeg tag system and introduce a powerful new tool for bacterial optogenetics.

Tague, Nathan (ORCID:0000000281146700)↗
DOE OSTI2021

Genome-wide Transcription Factor DNA Binding Sites and Gene Regulatory Networks in Clostridium thermocellum

Clostridium thermocellum is a thermophilic bacterium recognized for its natural ability to effectively deconstruct cellulosic biomass. While there is a large body of studies on the genetic engineering of this bacterium and its physiology to-date, there is limited knowledge in the transcriptional regulation in this organism and thermophilic bacteria in general. The study herein is the first report of a high-throughput application of DNA-affinity purification sequencing (DAP-seq) to transcription factors (TFs) from a thermophile. We applied DAP-seq to >90 TFs in C. thermocellum and detected genome-wide binding sites for 11 of them. We then compiled and aligned DNA binding sequences from these TFs to deduce the primary DNA-binding sequence motifs for each TF. These binding motifs are further validated with electrophoretic mobility shift assay (EMSA) and are used to identify individual TFs’ regulatory targets in C. thermocellum. Our results led to the discovery of novel, uncharacterized TFs as well as homologues of previously studied TFs including RexA-, LexA- and LacI-type TFs. We then used these data to reconstruct gene regulatory networks for the 11 TFs individually, which resulted in a global network encompassing the TFs with some interconnections. As gene regulation governs and constrains how bacteria behave, our findings shed light on the roles of TFs delineated by their regulons, and potentially provides a means to enable rational, advanced genetic engineering of C. thermocellum and other organisms alike towards a desired phenotype.

09 BIOMASS FUELS↗
DOE OSTI2022

EV Charging and the Impacts of Electricity Demand Charges

These slides were presented at the Los Angeles Cleantech Incubator's (LACI's) April 2022 combined Light-Duty and Goods Movement Working Group meetings. The talk summarized recent and ongoing research at NREL to model EV charging demands for passenger and commercial M/HD vehicles and estimate the impacts of utility demand charges on charging costs.

ADVANCED PROPULSION SYSTEMS,POWER TRANSMISSION AND↗
DOE OSTI2023

Boron coated straw-based neutron multiplicity counter for neutron interrogation of TRISO fueled pebbles

Pebble bed reactors (PBRs) can improve the safety and economics of the nuclear energy production. PBRs rely on TRIstructural-ISOtropic (TRISO) fuel pebbles for enhanced fission product retention. Accurate characterization of individual fuel pebbles would enable the validation of computational models, efficient use of TRISO fuel, and improve fuel accountability. Here, we have developed and tested a new neutron multiplicity counter (NMC) based on 192 boron coated straw (BCS) detectors optimized for 235 U assay in TRISO fuel. The new design yielded a singles and doubles neutron detection efficiency of 4.71% and 0.174%, respectively, and a die-away time of 16.7 μs. The NMC has a low intrinsic gamma-ray detection efficiency of 8.71 x 10 –8 at an exposure rate of 80.3 mR/h. In simulation, a high-efficiency version of the NMC encompassing 396 straws was able to estimate the 235 U in a pebble with a relative uncertainty and error both below 2% in 100 s.

11 NUCLEAR FUEL CYCLE AND FUEL MATERIALS↗