Engineering topics

Foster, Charles

Publications and source records attributed to Foster, Charles.

13 records

DOE OSTI2024

maranasgroup/yeastsMFA

The parameterization of kinetic models requires measurement of fluxes and/or metabolite levels for a base strain and a few genetic perturbations thereof. Unlike stoichiometric models that are mostly invariant to the specific strain, it remains unclear whether kinetic models constructed for different strains of the same species have similar or significantly different kinetic parameters. This important question underpins the applicability range and prediction limits of kinetic reconstructions. To this end, herein we parameterize two separate large-scale kinetic models using K-FIT with genome-wide coverage corresponding to two distinct strains of Saccharomyces cerevisiae: CEN.PK 113-7D strain (model k-sacce306-CENPK), and growth-deficient BY4741 (isogenic to S288c; model k-sacce306-BY4741). The metabolic network for each model contains 306 reactions, 230 metabolites, and 119 substrate-level regulatory interactions. The two models (for CEN.PK and BY4741) recapitulate, within one standard deviation, 77% and 75% of the fitted dataset fluxes, respectively, determined by 13C metabolic flux analysis for wild-type and eight single-gene knockout mutants of each strain. Strain-specific kinetic parameterization results indicate that key enzymes in the TCA cycle, glycolysis, and arginine and proline metabolism drive the metabolic differences between these two strains of S. cerevisiae. Our results suggest that although kinetic models cannot be readily used across strains as stoichiometric models, they can capture species-specific information through the kinetic parameterization process.

Dinh, Hoang↗

DOE OSTI2022

Toward low-cost biological and hybrid biological/catalytic conversion of cellulosic biomass to fuels

Hybrid processes, featuring biological conversion of lignocellulose to small molecules followed by chemo-catalytic conversion to larger molecules suitable for difficult-to-electrify transport modes, are a promising route to biomass-derived fuels in demand for climate stabilization.

09 BIOMASS FUELS↗

DOE OSTI2022

Functional Analysis of H + -Pumping Membrane-Bound Pyrophosphatase, ADP-Glucose Synthase, and Pyruvate Phosphate Dikinase as Pyrophosphate Sources in Clostridium thermocellum

Increased understanding of the central metabolism of C. thermocellum is important from a fundamental as well as from a sustainability and industrial perspective. In addition to showing that H + -pumping membrane-bound PPase, glycogen cycling, a Ppdk–malate shunt cycle, and acetate cycling are not significant sources of PP i supply, this study adds functional annotation of four genes and availability of an updated PP i stoichiometry from biosynthesis to the scientific domain.

Clostridium thermocellum↗

DOE OSTI2022

Assessing the impact of substrate-level enzyme regulations limiting ethanol titer in Clostridium thermocellum using a core kinetic model

Clostridium thermocellum is a promising candidate for consolidated bioprocessing because it can directly ferment cellulose to ethanol. Despite significant efforts, achieved yields and titers fall below industrially relevant targets. This implies that there still exist unknown enzymatic, regulatory, and/or possibly thermodynamic bottlenecks that can throttle back metabolic flow. By (i) elucidating internal metabolic fluxes in wild-type C. thermocellum grown on cellobiose via 13 C-metabolic flux analysis ( 13 C-MFA), (ii) parameterizing a core kinetic model, and (iii) subsequently deploying an ensemble-docking workflow for discovering substrate-level regulations, this paper aims to reveal some of these factors and expand our knowledgebase governing C. thermocellum metabolism. Generated 13 C labeling data were used with 13 C-MFA to generate a wild-type flux distribution for the metabolic network. Notably, flux elucidation through MFA alluded to serine generation via the mercaptopyruvate pathway. Using the elucidated flux distributions in conjunction with batch fermentation process yield data for various mutant strains, we constructed a kinetic model of C. thermocellum core metabolism (i.e. k-ctherm138). Subsequently, we used the parameterized kinetic model to explore the effect of removing substrate-level regulations on ethanol yield and titer. Upon exploring all possible simultaneous (up to four) regulation removals we identified combinations that lead to many-fold model predicted improvement in ethanol titer. In addition, by coupling a systematic method for identifying putative competitive inhibitory mechanisms using K-FIT kinetic parameterization with the ensemble-docking workflow, we flagged 67 putative substrate-level inhibition mechanisms across central carbon metabolism supported by both kinetic formalism and docking analysis.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2021

Modeling Growth Kinetics, Interspecies Cell Fusion, and Metabolism of a Clostridium acetobutylicum / Clostridium ljungdahlii Syntrophic Coculture

ABSTRACT Clostridium acetobutylicum and Clostridium ljungdahlii grown in a syntrophic culture were recently shown to fuse membranes and exchange cytosolic contents, yielding hybrid cells with significant shifts in gene expression and growth phenotypes. Here, we introduce a dynamic genome-scale metabolic modeling framework to explore how cell fusion alters the growth phenotype and panel of metabolites produced by this binary community. Computational results indicate C. ljungdahlii persists in the coculture through proteome exchange during fusing events, which endow C. ljungdahlii cells with expanded substrate utilization, and access to additional reducing equivalents from C. acetobutylicum -evolved H 2 and through acquisition of C. acetobutylicum -native cofactor-reducing enzymes. Simulations predict maximum theoretical ethanol and isopropanol yields that are increased by 0.64 mmol and 0.39 mmol per mmol hexose sugar consumed, respectively, during exponential growth when cell fusion is active. This modeling effort provides a mechanistic explanation for the metabolic outcome of cellular fusion and altered homeostasis achieved in this syntrophic clostridial community. IMPORTANCE Widespread cell fusion and protein exchange between microbial organisms as observed in synthetic C. acetobutylicum / C. ljungdahlii culture is a novel observation that has not been explored in silico . The mechanisms responsible for the observed cell fusion events in this culture are still unknown. In this work, we develop a modeling framework that captures the observed culture composition and metabolic phenotype, use it to offer a mechanistic explanation for how the culture achieves homeostasis, and identify C. ljungdahlii as primary beneficiary of fusion events. The implications for the events described in this study are far reaching, with potential to reshape our understanding of microbial community behavior synthetically and in nature.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2020

Metabolic flux analysis reaching genome wide coverage: lessons learned and future perspectives

13 C-MFA is currently the only technique capable of elucidating intracellular metabolic fluxes. Generally, in 13 C-MFA studies the reactions that can carry flux are mostly pre-specified by only considering canonical pathways and ignoring alternate ones. This may bias flux elucidation and cause labeling data to erroneously confirm implied assumptions. Here, by expanding the scope of the metabolic mapping models to match known genome-scale metabolism such estimation biases can be eliminated. However, this model expansion to genome-scale requires the construction of expanded atom mapping models, more efficient flux estimation algorithms, and formal estimation of confidence levels. Even though significant progress has been made in this direction, a number of challenges remain before widespread adoption by the community.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2020

In Vivo Thermodynamic Analysis of Glycolysis in Clostridium thermocellum and Thermoanaerobacterium saccharolyticum Using 13 C and 2 H Tracers

Clostridium thermocellum and Thermoanaerobacterium saccharolyticum are thermophilic anaerobic bacteria with complementary metabolic capabilities that utilize distinct glycolytic pathways for the conversion of cellulosic sugars to biofuels. We integrated quantitative metabolomics with 2 H and 13 C metabolic flux analysis to investigate the in vivo reversibility and thermodynamics of the central metabolic networks of these two microbes. We found that the glycolytic pathway in C. thermocellum operates remarkably close to thermodynamic equilibrium, with an overall drop in Gibbs free energy 5-fold lower than that of T. saccharolyticum or anaerobically grown Escherichia coli . The limited thermodynamic driving force of glycolysis in C. thermocellum could be attributed in large part to the small free energy of the phosphofructokinase reaction producing fructose bisphosphate. The ethanol fermentation pathway was also substantially more reversible in C. thermocellummthan in T. saccharolyticum. These observations help explain the comparatively low ethanol titers of C. thermocellum and suggest engineering interventions that can be used to increase its ethanol productivity and glycolytic rate. In addition to thermodynamic analysis, we used our isotope tracer data to reconstruct the T. saccharolyticum central metabolic network, revealing exclusive use of the Embden-Meyerhof-Parnas (EMP) pathway for glycolysis, a bifurcated tricarboxylic acid (TCA) cycle, and a sedoheptulose bisphosphate bypass active within the pentose phosphate pathway. Thermodynamics constitutes a key determinant of flux and enzyme efficiency in metabolic networks. Here, we provide new insights into the divergent thermodynamics of the glycolytic pathways of C. thermocellum and T. saccharolyticum, two industrially relevant thermophilic bacteria whose metabolism still is not well understood. We report that while the glycolytic pathway in T. saccharolyticum is as thermodynamically favorable as that found in model organisms, such as E. coli or Saccharomyces cerevisiae , the glycolytic pathway of C. thermocellum operates near equilibrium. The use of a near-equilibrium glycolytic pathway, with potentially increased ATP yield, by this cellulolytic microbe may represent an evolutionary adaptation to growth on cellulose, but it has the drawback of being highly susceptible to product feedback inhibition. The results of this study will facilitate future engineering of high-performance strains capable of transforming cellulosic biomass to biofuels at high yields and titers.

59 BASIC BIOLOGICAL SCIENCES↗

NASA NTRS2016

Team Update on North American Proton Facilities for Radiation Testing

In the wake of the closure of the Indiana University Cyclotron Facility (IUCF), this presentation provides an overview of the options for North American proton facilities. This includes those in use by the aerospace community as well as new additions from the cancer therapy regime. In addition, proton single event testing background is provided for understanding the criteria needed for these facilities for electronics testing.

Cancer therapy↗

NASA NTRS2016

Team Update on North American Proton Facilities for Radiation Testing

Single event effects testing↗

NASA NTRS2016

Team Update on North American Proton Facilities for Radiation Testing

Proton cyclotrons↗

NASA NTRS2016

Team Update on North American Proton Facilities for Radiation Testing

Single event effects testing↗

NASA NTRS1995

Progress in HTS trapped field magnets: J(sub c), area, and applications

Progress in trapped field magnets is reported. Single YBCO grains with diameters of 2 cm are made in production quantities, while 3 cm, 4 1/2 cm and 6 cm diameters are being explored. For single grain tiles: J(sub c) is approximately 10,000 A/cm(exp 2) for melt textured grains; J(sub c) is approximately 40,000 A/cm2 for light ion irradiation; and J(sub c) is approximately 85,000 A/cm(exp 2) for heavy ion irradiation. Using 2 cm diameter tiles bombarded by light ions, we have fabricated a mini-magnet which trapped 2.25 Tesla at 77K, and 5.3 Tesla at 65K. A previous generation of tiles, 1 cm x 1 cm, was used to trap 7.0 Tesla at 55K. Unirradiated 2.0 cm tiles were used to provide 8 magnets for an axial gap generator, in a collaborative experiment with Emerson Electric Co. This generator delivered 100 Watts to a resistive load, at 2265 rpm. In this experiment activation of the TFMs was accomplished by a current pulse of 15 ms duration. Tiles have also been studied for application as a bumper-tether system for the soft docking of spacecraft. A method for optimizing tether forces, and mechanisms of energy dissipation are discussed. A bus bar was constructed by welding three crystals while melt-texturing, such that their a,b planes were parallel and interleaved. The bus bar, an area of approximately 2 cm(exp 2), carried a transport current of 1000 amps, the limit of the testing equipment available.

Weinstein, Roy↗

NASA NTRS1995

Progress in HTS Trapped Field Magnets: J(sub c), Area, and Applications

Progress in trapped field magnets is reported. Single YBCO grains with diameters of 2 cm are made in production quantities, while 3 cm, 4 1/2 cm and 6 cm diameters are being explored. For single grain tiles: J(sub c) - 10,000 A/sq cm for melt textured grains; J(sub c) - 40,000 A/sq cm for light ion irradiation; and J(sub c) - 85,000 A/J(sub c) for heavy ion irradiation. Using 2 cm diameter tiles bombarded by light ions, we have fabricated a mini-magnet which trapped 2.25 Tesla at 77K, and 5.3 Tesla at 65K. A previous generation of tiles, 1 cm x 1 cm, was used to trap 7.0 Tesla at 55K. Unirradiated 2.0 cm tiles were used to provide 8 magnets for an axial gap generator, in a collaborative experiment with Emerson Electric Co. This generator delivered 100 Watts to a resistive load, at 2265 rpm. In this experiment, activation of the TFMs was accomplished by a current pulse of 15 ms duration. Tiles have also been studied for application as a bumper-tether system for the soft docking of spacecraft. A method for optimizing tether forces, and mechanisms of energy dissipation are discussed. A bus bar was constructed by welding three crystals while melt-texturing, such that their a,b planes were parallel and interleaved. The bus bar, of area approx. 2 sq cm, carried a transport current of 1000 amps, the limit of the testing equipment available.

Weinstein, Roy↗