Design and Evaluation of Artificial Hybrid Photoredox Biocatalysts
Not provided.
Biochemistry & Molecular Biology↗
SEARCH · Search NASA
Results for “biocatalysts”
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.
Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.
Cell-Free-Based Thermophilic Biocatalyst for the Synthesis of Amino Acids from One-Carbon Feedstocks
Bioproduction from one-carbon compounds, such as formate, is an attractive prospect due to reduced energy requirements and the possibility for using CO 2 as a sustainable feedstock. Formate-fixing pathways engineered using Escherichia coli lysate-based cell-free expression (CFE) biocatalysts have the potential to route 100% of feedstock carbon toward chemical synthesis but are undermined by siphoning of in-pathway metabolites and cofactors by the CFE background metabolism. To address this limitation, we engineer a CFE-based thermophilic multienzyme biocatalyst for the synthesis of serine and glycine from formate, bicarbonate, and ammonia. After expression of the thermophilic formate-to-serine pathway in a one-pot reaction, the mesophilic E. coli CFE background machinery is removed by simple heat denaturation, eliminating the siphoning of cofactors, inpathway metabolites, and products. After bioprocess optimization, including pathway gene expression duration and chemical synthesis temperature, we achieve near stoichiometric conversion of formate and bicarbonate to serine and glycine, reaching 97% of stoichiometric yield. The use of a moderately thermophilic biocatalyst allowed chemical synthesis to take place at mesophilic temperatures, enabling the balance of optimal enzyme activity with minimal metabolite/cofactor thermal degradation. In a fed-batch experiment, the biocatalyst shows sustained chemical synthesis rates for 8 h, paving the way toward a continuous bioprocess. Finally, a sensitivity analysis of cofactor usage revealed that the most expensive cofactors, THF and NADPH, can be reduced by 5-fold without significantly lowering product yields. To the best of our knowledge, this is the first instance of expressing a thermophilic pathway in an E. coli lysate-based CFE system to generate a thermophilic biocatalyst for use at mesophilic temperatures. The CFEbased thermophilic formate-to-serine biocatalyst triples the combined serine and glycine yield previously obtained by a CFE-based mesophilic formate-to-serine biocatalyst (30%), and quadruple the yield obtained by a purified enzyme system (22%). Ultimately, this work opens the door to using E. coli lysate-based CFE for thermophilic biocatalyst generation to achieve high chemical synthesis yields.
bacteria↗
Carbon Negative Synthesis of Amino Acids Using a Cell-Free-Based Biocatalyst
Biological systems can directly upgrade carbon dioxide (CO 2 ) into chemicals. The CO 2 fixation rate of autotrophic organisms, however, is too slow for industrial utility, and the breadth of engineered metabolic pathways for the synthesis of value-added chemicals is too limited. Biotechnology workhorse organisms with extensively engineered metabolic pathways have recently been engineered for CO 2 fixation. Yet, their low carbon fixation rate, compounded by the fact that living organisms split their carbon between cell growth and chemical synthesis, has led to only cell growth with no chemical synthesis achieved to date. Here, we engineer a lysate-based cell-free expression (CFE)-based multienzyme biocatalyst for the carbon negative synthesis of the industrially relevant amino acids glycine and serine from CO 2 equivalents–formate and bicarbonate–and ammonia. The formate-to-serine biocatalyst leverages tetrahydrofolate (THF)-dependent formate fixation, reductive glycine synthesis, serine synthesis, and phosphite dehydrogenase-dependent NAD(P)H regeneration to convert 30% of formate into serine and glycine, surpassing the previous 22% conversion using a purified enzyme system. We find that (1) the CFE-based biocatalyst is active even after 200-fold dilution, enabling higher substrate loading and product synthesis without incurring additional cell lysate cost, (2) NAD(P)H regeneration is pivotal to driving forward reactions close to thermodynamic equilibrium, (3) balancing the ratio of the formate-to-serine pathway genes added to the CFE is key to improving amino acid synthesis, and (4) efficient THF recycling enables lowering the loading of this cofactor, reducing the cost of the CFE-based biocatalyst. To our knowledge, this is the first synthesis of amino acids that can capture CO 2 equivalents for the carbon negative synthesis of amino acids using a CFE-based biocatalyst. Looking ahead, the CFE-based biocatalyst process could be extended beyond serine to pyruvate, a key intermediate, to access a variety of chemicals from aromatics and terpenes to alcohols and polymers.
37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
Smart Microbial Cell Technology: A high-throughput platform to optimize biocatalysts
Los Alamos National Laboratory developed an ultra-high-throughput screening platform to engineer custom biocatalysts that enhance the rate of chemical reactions critical in pharmaceuticals, renewable energy, and environmental cleanup. Current methods to find biocatalysts are slow. This platform scans genetic variations to optimize a single enzyme or microbial cell to generate a product efficiently. It selects rare mutations needed for biocatalyst optimization orders of magnitude faster than current screening methods. A custom sensor reporter gene circuit causes cells to fluoresce when they are making the target product. When coupled to flow cytometry, a million biocatalyst variants can be screened in hours.
37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
Biocatalyst discovery and design for plastics deconstruction: A multi‐scale perspective
Plastic waste accumulation poses significant environmental challenges due to a lack of economical solutions for the molecular deconstruction of diverse synthetic polymers. Biological‐based degradation offers promise but is hindered by the crystallinity, hydrophobicity, and additive complexity of plastics, which restrict biocatalyst access and activity. To address these problems, we propose a multi‐scale framework that combines detailed materials characterization, optimization of plastic‐biomolecular interfacial interactions, and enhancement of biocatalytic kinetics to develop effective plastic‐deconstructing enzymes. This approach leverages principles from reaction kinetics, transport and interfacial phenomena, and enzyme engineering to systematically address barriers across diverse plastic types. Our framework aims to accelerate the discovery and optimization of biocatalysts capable of scalable, selective, and efficient deconstruction of plastic waste. These advances hold potential to enable sustainable biological recycling and upcycling pathways, contributing to global efforts in mitigating plastic pollution and promoting circular material economies.
37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
Principles and practice of designing microbial biocatalysts for fuel and chemical production
Abstract The finite nature of fossil fuels and the environmental impact of its use have raised interest in alternate renewable energy sources. Specifically, nonfood carbohydrates, such as lignocellulosic biomass, can be used to produce next generation biofuels, including cellulosic ethanol and other nonethanol fuels like butanol. However, currently there is no native microorganism that can ferment all lignocellulosic sugars to fuel molecules. Thus, research is focused on engineering improved microbial biocatalysts for production of liquid fuels at high productivity, titer, and yield. A clear understanding and application of the basic principles of microbial physiology and biochemistry are crucial to achieve this goal. In this review, we present and discuss the construction of microbial biocatalysts that integrate these principles with ethanol-producing Escherichia coli as an example of metabolic engineering. These principles also apply to fermentation of lignocellulosic sugars to other chemicals that are currently produced from petroleum.
09 BIOMASS FUELS↗
Modified biosensors and biocatalysts and methods of use
Biosensors including a nucleic acid encoding a PcaU protein, a PobR protein, a CatM protein, a PcaR protein, or a TphR protein are provided. In some examples, the biosensors include a promoter regulated by the sensed molecule operably linked to a reporter gene. The biosensors may be included in a vector or in cells including one or more of the biosensors or vectors. Modified chorismate pyruvate lyase (UbiC) and modified paraoxonase (PON1) proteins including one or more amino acid substitutions are provided. Finally, methods of selecting biocatalysts with increased activity including transforming a library of cells expressing a biosensor with one or more nucleic acids encoding one or more mutations in a gene involved in a biosynthesis pathway, determining activity of the reporter protein; and selecting a cell with increased reporter protein activity as expressing a biocatalyst with increased activity are provided.
Jha, Ramesh K.↗
Strategies for designing biocatalysts with new functions
Enzymes can be optimized to accelerate chemical transformations via a range of methods. In this review, we showcase how protein engineering and computational design techniques can be interfaced to develop highly efficient and selective biocatalysts.
09 BIOMASS FUELS↗
Advances in systems metabolic engineering of autotrophic carbon oxide-fixing biocatalysts towards a circular economy
High levels of anthropogenic CO 2 emissions are driving the warming of global climate. If this pattern of increasing emissions does not change, it will cause further climate change with severe consequences for the human population. On top of this, the increasing accumulation of solid waste within the linear economy model is threatening global biosustainability. The magnitude of these challenges requires several approaches to capture and utilize waste carbon and establish a circular economy. Microbial gas fermentation presents an exciting opportunity to capture carbon oxides from gaseous and solid waste streams with high feedstock flexibility and selectivity. Here we discuss available microbial systems and review in detail the metabolism of both anaerobic acetogens and aerobic hydrogenotrophs and their ability to utilize C1 waste feedstocks. More specifically, we provide an overview of the systems-level understanding of metabolism, key metabolic pathways, scale-up opportunities and commercial successes, and the most recent technological advances in strain and process engineering. Finally, we also discuss in detail the gaps and opportunities to advance the understanding of these autotrophic biocatalysts for the efficient and economically viable production of bioproducts from recycled carbon.
59 BASIC BIOLOGICAL SCIENCES↗
Methane Consuming Biocatalysts in Gas-Solid Reactor System
Carbon dioxide and methane are the two most abundant greenhouse gases with methane having a higher global warming potential (GWP) than carbon dioxide. Use of methanotrophs as a biocatalyst is a green alternative to chemical conversion of methane. Methanotrophs were immobilized in a 4-arm poly (-ethylene glycol)-tetra-acrylate, PEGTA, hydrogel that is supported by a 3D printed lattice containing micropores. The consumption rate of methane was 2.5 and 42 g CH 4 /L-hr in samples that were at 1 and 3 atm, respectively. A small increase in pressure of the system improves the consumption rate by almost 20x with minimal changes to the reactor system. This system relies on the mass transport of gas through a solid-like material rather than dissolving the gas in a liquid. Immobilization of cells in hydrogel allows for higher cell density and overall increased conversion rates compared to traditional stirred tank reactors. There are many other applications of the use of immobilized biological materials in polymers that require gas-phase reactions.
36 MATERIALS SCIENCE↗
Characterization of Coal Seam Produced Water from the Appalachian Basin: Geochemistry, Microbiology, and Biocatalyst Development for Biological CO 2 Conversion
To enhance domestic energy repositories, it is necessary to develop carbon utilization technologies to repurpose unused carbon sources, and/or enhance the economic viability and carbon conversion efficiency of current technologies. To this end we have developed a biocatalyst, enriched from coal seam produced water, to convert gaseous and liquid C1 compounds (e.g. CO 2 , bicarbonate, formate, and/or methanol) into acetate, a C2 organic acid with a projected global market size of $\$$32.4 billion USD by 2030.
01 COAL, LIGNITE, AND PEAT↗
The Arginine Kinase from the Tick Rhipicephalus sanguineus Is an Efficient Biocatalyst
Arginine kinase (AK) is a reversible enzyme that regulates invertebrates’ phosphagen arginine phosphate levels. AK also elicits an immune response in humans, and it is a major food allergen in crustacea and may be a target for novel antiparasitic drugs. Although AK has been primarily described in the shrimp, it is also present in other invertebrates, such as the brown tick Rhipicephalus sanguineus (Rs), the vector for Rocky Mountain Spotted Fever. Here we report the enzymatic activity and the crystal structure of AK from Rhipicephalus sanguineus (RsAK) in an open conformation without substrate or ligands and a theoretical structure of RsAK modeled bound with the substrate/product (Arg-ADP) in a closed conformation. The Michaelis-Menten kinetics confirmed that RsAK is an efficient biocatalyst due to its high kcat/Km parameter. The recombinant enzyme was expressed in bacteria and purified to a 20 mg/L culture yield. AK is an essential enzyme in invertebrates. Future work will be focused on the RsAK enzymatic inhibition that may lead to novel strategies to control this pest, a burden to animal and human health.
37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
Application of biocatalysts to Space Station ECLSS and PMMS water reclamation
Immobilized enzyme reactors have been developed and tested for potential water reclamation applications in the Space Station Freedom Environmental Control and Life Support System (ECLSS) and Process Materials Management System (PMMS). The reactors convert low molecular weight organic contaminants found in ECLSS and PMMS wastewaters to compounds that are more efficiently removed by existing technologies. Demonstration of the technology was successfully achieved with two model reactors. A packed bed reactor containing immobilized urease was found to catalyze the complete decomposition of urea to by-products that were subsequently removed using conventional ion exchange results. A second reactor containing immobilized alcohol oxidase showed promising results relative to its ability to convert methanol and ethanol to the corresponding aldehydes for subsequent removal. Preliminary assessments of the application of biocatalysts to ECLSS and PMMS water reclamation sytems are presented.
Jolly, Clifford D.↗
Thermophilic biocatalysts for one-step conversion of citrus waste into lactic acid
Agri-food residues offer significant potential as a raw material for the production of L-lactic acid through microbial fermentation. Weizmannia coagulans, previously known as Bacillus coagulans, is a spore-forming, lactic acid-producing, gram-positive, with known probiotic and prebiotic properties. This study aimed to evaluate the feasibility of utilizing untreated citrus waste as a sustainable feedstock for the production of L-lactic acid in a one-step process, by using the strain W. coagulans MA-13. By employing a thermophilic enzymatic cocktail (Cellic CTec2) in conjunction with the hydrolytic capabilities of MA-13, biomass degradation was enhanced by up to 62%. Moreover, batch and fed-batch fermentation experiments demonstrated the complete fermentation of glucose into L-lactic acid, achieving a concentration of up to 44.8 g/L. These results point to MA-13 as a microbial cell factory for one-step production of L-lactic acid, by combining cost-effective saccharification with MA-13 fermentative performance, on agri-food wastes. Moreover, the potential of this approach for sustainable valorization of agricultural waste streams is successfully proven.
37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
Evaluation of biocatalysts for artificial photosynthesis
Book chapter on methods for evaluation H2 and CO production in photocatalytic systems
Irfan, Hafsa [University of Rochester]↗
Structural and Biochemical Studies of a Biocatalyst for the Enzymatic Production of Wax Esters
Abstract not provided
Petronikolou, Nektaria↗
A flexible kinetic assay efficiently sorts prospective biocatalysts for PET plastic subunit hydrolysis
We have developed a colorimetric assay for the high throughput detection of enzymatic ester hydrolysis of PET plastic monomers, applied in a microplate format.
09 BIOMASS FUELS↗