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All sourcesNASA NTRSDOE OSTI
DOE OSTI2023

"A" is for Ammonia: Why and How Fertilizer May Power the Planet

Ammonia (chemical formula NH3) is so pervasive in our history that human beings simply wouldn't exist without it: 4 billion people on this planet are fed by food grown using ammonia fertilizer, but this powerful molecule consumes nearly 2% of the world's energy supply to be produced at an industrial scale. This talk will first explore the energy challenges of ammonia's current means of industrial production, followed by a new vision for its synthesis using only renewable electricity, air and water, and opportunities to use ammonia as a hydrogen source. Fertilizer may one day fuel more than just our mouths.

ammonia↗
DOE OSTI2025

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↗
DOE OSTI2024

Toward sustainable production of N-containing products via nonthermal plasma-enhanced conversion of natural gas resources

Nonthermal plasmas can directly activate and cleave the strong chemical bonds in molecular nitrogen and methane to facilitate the transformation of these inherently stable molecules through the application of electrical energy. Here, we report a low temperature, atmospheric pressure, nonthermal plasma for the “one-pot” synthesis of olefins, alkynes, higher molecular weight hydrocarbons, ammonia, and nitrogen-containing liquids from a representative shale gas feed enriched with nitrogen. Furthermore, we reproducibly observe a wide range of valuable and synthetically challenging gas-phase and liquid-phase products containing C-N, C-C, and N-H bonding by controlling the N 2 concentration in the inlet feed stream. In nitrogen-lean regimes, hydrocarbon products dominate (e.g., ethylene, acetylene, etc.), while nitrogen-rich regimes promote incorporation of nitrogen into the products, leading to the formation of ammonia and liquid products containing a variety of functionalities (e.g., nitriles, amines, heterocycles). High resolution electrospray ionization mass spectrometry was used to measure molecular weights and identify the chemical formulas of the liquid products. Van Krevelen diagrams were created and showed many products with compositions around H/C = 2 and N/C = 0.5, indicating the potential importance of intermediate species with these ratios for liquid formation (e.g., CH 3 CN + H).

03 NATURAL GAS↗
DOE OSTI2021

Metal nitride materials for solar-thermal ammonia production [Slides]

Solar Thermal Ammonia Production has the potential to synthesize ammonia in a green, renewable process that can greatly reduce the carbon footprint left by the conventional Haber-Bosch reaction. Co 3 Mo 3 N has been identified as a potential candidate for ammonia production. It is synthesized via oxide precursor synthesis followed by nitridation under 10% H 2 /N 2 . The synthesis method can be extended to other candidate nitrides. The Co 3 Mo 3 N → Co 6 Mo 6 N reduction is demonstrated on TGA with rapid kinetics. The formation of NH 3 is qualitatively observed, but not quantitatively determined. The material retains crystal structure, but no secondary phases are observed in XRD. Partial re-nitridation back to CMN331 of ~35% of max nitridation is observed. Reaction parameters in TGA differ from experimental conditions in the literature. Experiments at Georgia Tech better mimic re-nitridation conditions with more sensitive, quantitative analytical techniques (GC-MS). The ASU NH 3 synthesis/re-nitridation reactor is under development and will permit experiments (reduction/re-nitridation) under precisely controlled T, pH 2 .

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2024

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↗
DOE OSTI2023

Solar Thermochemical Ammonia Production (STAP) (Final Report)

Ammonia (NH 3 ) is an energy-dense chemical and a vital component of fertilizer. In addition, it is a carbon-neutral liquid fuel and a potential candidate for thermochemical energy storage for high-temperature concentrating solar power (CSP). Currently, NH 3 synthesis occurs via the Haber-Bosch process, which requires high pressures (15-25 MPa) and medium to high temperatures (400-500 °C). N 2 and H 2 are essential feedstocks for this NH 3 production process. H 2 is generally derived from methane via steam reforming; N 2 is sourced from air, after oxygen removal via combustion of hydrocarbons. Both processes consume hydrocarbons, resulting in the release of CO 2 . In addition, hydrocarbon fuels are burned to produce the heat and mechanical energy required to perform the NH 3 reaction, further increasing CO 2 emissions. Overall, the production of ammonia via the Haber-Bosch (H-B) process is responsible for up to 1.4% of the world’s carbon emissions. The development of a renewable pathway to NH 3 synthesis, which utilizes concentrated solar irradiation as a process heat instead of fossil fuels and operates under low or ambient pressure, will result in a decrease (or elimination) of greenhouse gas emissions as well as avoid the cost, complexity, and safety issues inherent in high-pressure processes. Most current efforts to “green” ammonia production involve either electrolysis or simply replacing the energy source for H-B with renewable electricity, but otherwise leaving the process intact. The effort proposed here would create a new paradigm for the synthesis of NH 3 utilizing solar-thermal heat, water, and air as feedstocks, providing a truly green method of production. The overall objective of the STAP (Solar Thermal Ammonia Production) project was to develop a solar thermochemical looping technology to produce and store nitrogen (N 2 ) from air for the subsequent production of ammonia (NH 3 ) via an advanced two-stage process. The goal is a cost-effective and energy efficient technology for the renewable N 2 production and synthesis of NH 3 from H 2 (produced from H 2 O) and air using solar-thermal energy from concentrating sunlight, under pressures an order of magnitude lower than H-B NH 3 production. Our process involves two looping cycles, which do not require catalysts and can be recycled. Over the course of the STAP project, we (1) developed and deeply characterized oxide materials for N 2 separation; (2) developed a method for the synthesis of metal nitrides, producing a series of quaternary compounds that have been heretofore unreported; (3) modeled, designed, and fabricated bench-scale tube and on-sun reactors for the N 2 production step and demonstrated the ability to separate N 2 over multiple cycles in the tube reactor; (4) designed and fabricated a bench-scale Ammonia Synthesis Reactor (ASR) and demonstrated the proof of concept of NH 3 synthesis via a novel looping process using metal nitrides over multiple cycles; and (5) completed a systems- and technoeconomic analysis showing the feasibility of ammonia production on a larger scale via the STAP process. The development of renewable, low-cost NH 3 will be of great interest to the chemicals industry, particularly agricultural sectors. The CSP industry should be both an important customer and potential end-user of this technology, as it affords the capability of synthesizing a promising thermochemical storage material on-site. Since the NH 3 synthesis step also requires H 2 , there will exist a symbiotic relationship between this technology and solar-thermochemical water-splitting applications. Green ammonia synthesis will result in the decarbonization of a hydrocarbon-intensive industry, helping to meet the Administration goal of industrial decarbonization by 2050. The resulting decrease in CO 2 and related pollutants will improve health and well-being of society, particularly for those living in the vicinity of commercial production plants.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2022

Isotope effects at the origin of life: Fingerprints of the Strecker synthesis

We report strecker synthesis creates α-amino acids from prebiotically plausible substrates (cyanide, ammonia, and aldehydes) and is widely hypothesized to be a key mechanism in the chemistry that led to life on Earth and on other planets. To better constrain the synthetic environments and precursors of abiotic α-amino acids, and to determine unique signatures of abiogenic amino acids, we measured the molecular-averaged and site-specific carbon and nitrogen isotope effects for the Strecker synthesis of alanine. The reaction steps of the Strecker synthesis can be divided into two groups: an initial series of reversible amination and nitrile-addition reactions (‘equilibration’) and a second series of irreversible hydrolysis reactions (‘hydrolysis’). The equilibration of cyanide, acetaldehyde, and ammonia with the intermediate, α-aminopropionitrile (α-APN), has a measured 55.1‰ equilibrium nitrogen isotope effect between the 15 N-rich amine nitrogen in α-aminopropionitrile and the 15 N-poor ammonia and a 20.0‰ equilibrium carbon isotope effect between the 13 C-poor C-2 site in α-aminopropionitrile and the 13 C-rich carbonyl carbon in acetaldehyde. The first irreversible hydrolysis step is inferred to have an up to 10‰ normal carbon fractionation (i.e., faster for 12 C, slower for 13 C) for the whole molecule, but it also has one or more side reactions that deplete the reactive α-APN reservoir by up to 15‰. The second hydrolysis step has a 15.4‰ normal kinetic isotope effect on the amide (C-1) site of alaninamide, which becomes the carboxyl site of alanine. Other α-amino acids will likely experience similar nitrogen isotope fractionations between ammonia and their amine sites, and similar carbon isotope fractionations between the carbonyl carbon in reactant aldehydes or ketones and the intermediate α-aminonitrile, and between cyanide and the carboxyl site. Therefore, these isotope effects allow us to predict the carbon and nitrogen isotopic contents and intramolecular structures of α-amino acids formed by Strecker synthesis based on their substrates’ isotopic compositions, or to infer the isotopic compositions of substrates from which amino acids formed, for example in the case of the amino-acid-rich carbonaceous chondrites. The site-specific C and N isotopic compositions of amino acids formed by Strecker chemistry contrast with those typical of terrestrial biosynthetic amino acids, so these data also provide a means of discriminating between biogenic and abiogenic α-amino acids.

58 GEOSCIENCES↗
DOE OSTI2020

Material Discovery and High Throughput Exploration of Ru Based Catalysts for Low Temperature Ammonia Decomposition

High throughput experimentation has the capability to generate massive, multidimensional datasets, allowing for the discovery of novel catalytic materials. Here, we show the synthesis and catalytic screening of over 100 unique Ru-Metal-K based bimetallic catalysts for low temperature ammonia decomposition, with a Ru loading between 1–3 wt% Ru and a fixed K loading of 12 wt% K, supported on γ-Al2O3. Bimetallic catalysts containing Sc, Sr, Hf, Y, Mg, Zr, Ta, or Ca in addition to Ru were found to have excellent ammonia decomposition activity when compared to state-of-the-art catalysts in literature. Furthermore, the Ru content could be reduced to 1 wt% Ru, a factor of four decrease, with the addition of Sr, Y, Zr, or Hf, where these secondary metals have not been previously explored for ammonia decomposition. The bimetallic interactions between Ru and the secondary metal, specifically RuSrK and RuFeK, were investigated in detail to elucidate the reaction kinetics and surface properties of both high and low performing catalysts. The RuSrK catalyst had a turnover frequency of 1.78 s−1, while RuFeK had a turnover frequency of only 0.28 s−1 under identical operating conditions. Based on their apparent activation energies and number of surface sites, the RuSrK had a factor of two lower activation energy than the RuFeK, while also possessing an equivalent number of surface sites, which suggests that the Sr promotes ammonia decomposition in the presence of Ru by modifying the active sites of Ru.

McCullough, Katherine (ORCID:0000000197525740)↗
DOE OSTI2022

Molecular Simulations Probing the Adsorption and Diffusion of Ammonia, Nitrogen, Hydrogen, and Their Mixtures in Bulk MFI Zeolite and MFI Nanosheets at High Temperature and Pressure

Recent advances in the synthesis of MFI zeolite nanosheets have led to highly selective membranes that are promising candidates for small-scale ammonia separation from ammonia/nitrogen/hydrogen mixtures in distributed green ammonia production plants. Using force-field-based molecular simulations, we evaluate the performance of bulk all-silica MFI zeolite and a 3 nm thick all-silica MFI nanosheet for ammonia/nitrogen/hydrogen separations at moderate temperature and pressure conditions (T = 373 K, p = 5 bar) as well as conditions relevant for a membrane-based reactor–separator process (T = 523 or 623 K, p = 80 bar). Isobaric–isothermal Gibbs ensemble Monte Carlo (GEMC) simulations were carried out to understand the selective adsorption behavior for these mixtures. Molecular dynamics simulations in the canonical ensemble were performed to examine the transport behavior of ammonia in a mixture with nitrogen or hydrogen at loadings obtained from the GEMC simulations. Our results show that both bulk MFI and the nanosheet with explicit surface silanols are highly selective toward ammonia adsorption, but the adsorption selectivity decreases by factors of 4 and 10 for ammonia/nitrogen and ammonia/hydrogen mixtures as the temperature is increased from 373 to 623 K. Conversely, the diffusion selectivities toward ammonia are more favorable at process-relevant temperatures and for the MFI nanosheet. Here, at T = 523 K and p = 80 bar, our simulations predict overall separation factors of 3.8 ± 0.2 and 4.5 ± 0.3 for ammonia/nitrogen and ammonia/hydrogen mixtures, respectively, in the bulk MFI zeolite, and separation factors of 8.6 ± 0.4 and 12.5 ± 0.8, respectively, for the MFI nanosheet.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2022

Catalytic Processes to Accelerate Decarbonization in a Net-Zero Carbon World

Reducing carbon dioxide emissions is one of the critical challenges to mitigate global climate change, which is having detrimental impacts on society and the environment. Fossil fuel combustion in transportation, power generation, and industrial processes is the dominant contributor to carbon emissions. Over the past decades, sustainable solutions and strategies have been investigated and developed to enable decarbonization. Catalysis plays an essential role to address this global challenge by increasing energy efficiency, reducing carbon emissions, capturing carbon dioxide, and utilizing clean energy sources to displace fossil fuels. In this Review, the role of catalysis in reducing energy demand was discussed, enhancing process efficiency, displacing carbon-intensive feedstocks and products, and therefore, reducing carbon emissions. Here, recent advances in catalyst development were summarized, focusing on applications to enhance industrial processes efficiency and enable utilization of clean energy sources. Emerging approaches in catalysis were reviewed, including the manufacture of iron and steel, direct air capture of CO 2 , production of ethylene, ammonia, and sustainable aviation fuels, plastic recycling, and the synthesis of biobased plastics. The Review was concluded with suggested research directions to achieve a carbon net-zero world.

36 MATERIALS SCIENCE↗
DOE OSTI2022

Proteomic and metabolic disturbances in lignin-modified Brachypodium distachyon

Abstract Lignin biosynthesis begins with the deamination of phenylalanine and tyrosine (Tyr) as a key branch point between primary and secondary metabolism in land plants. Here, we used a systems biology approach to investigate the global metabolic responses to lignin pathway perturbations in the model grass Brachypodium distachyon. We identified the lignin biosynthetic protein families and found that ammonia-lyases (ALs) are among the most abundant proteins in lignifying tissues in grasses. Integrated metabolomic and proteomic data support a link between lignin biosynthesis and primary metabolism mediated by the ammonia released from ALs that is recycled for the synthesis of amino acids via glutamine. RNA interference knockdown of lignin genes confirmed that the route of the canonical pathway using shikimate ester intermediates is not essential for lignin formation in Brachypodium, and there is an alternative pathway from Tyr via sinapic acid for the synthesis of syringyl lignin involving yet uncharacterized enzymatic steps. Our findings support a model in which plant ALs play a central role in coordinating the allocation of carbon for lignin synthesis and the nitrogen available for plant growth. Collectively, these data also emphasize the value of integrative multiomic analyses to advance our understanding of plant metabolism.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Poly(acrylic acid) Enabling the Synthesis of Highly Uniform Silica Nanoparticles of sub-100 nm

Robust synthesis of small colloidal silica (SiO 2 ) nanoparticles with narrow size distributions (<5%) is challenging because the possible involvement of continuous nucleation and growth of SiO 2 in the widely used Stöber process usually results in silica nanoparticles with large sizes and broad size distributions. Promoting heterogeneous nucleation and growth of silica is promising to tackle the challenges by in-situ forming transient colloidal seeds that provide nucleation surfaces for silica condensate. Here in this work, poly(acrylic acid) (PAA) is introduced to the Stöber synthesis solutions, which are usually composed of ethanol (containing a trace amount of water), tetraethyl orthosilicate (TEOS), and ammonia, to mediate the nucleation of TEOS hydrolysates and growth of SiO 2 nanoparticles. PAA reacts with ammonia to form soft PAA-NH 3 complex colloids that serve as transient seeds to facilitate heterogeneous nucleation and growth of SiO 2 nanoparticles after adding TEOS to the synthesis solution. The number of PAA-NH 3 complex colloidal particles, which strongly depends on the molar ratio of PAA and ammonia, determines the number of the resulting SiO 2 nanoparticles and thus the size of the SiO 2 nanoparticles correspondingly according to the supply of TEOS. Highly uniform SiO 2 nanoparticles with finely tuned diameters in the sub-100 region have been successfully synthesized, and the synthesis protocol is feasible to be scaled up while maintaining the high quality of SiO 2 nanoparticles.

heterogeneous nucleation and growth of nanoparticl↗
DOE OSTI2024

Impact of Silicon Impurity on the Hydrometallurgical Recovery of NCM622 Cathode

Hydrometallurgy is one of the best approaches to date for recycling LIBs due to its high efficiency, low energy usage, and industrial scalability. However, impurities have always been a thorny issue because they could have unintended impacts on the recovered cathode materials. This research marks the first systematic investigation into the influence of silicon impurity on the LiNi 0.6 Co 0.2 Mn 0.2 O 2 (NCM622) cathode obtained from hydrometallurgical recycling. Here we find that silicon nanoparticles will be nucleated at the center of the precursor particles during co-precipitation synthesis, and the silicon core will slowly dissolve in the surrounding ammonia, creating a special hollow structure in the particle. More importantly, the dissolution of silicon impurity will eventually lead to the deposition of silicates in the cathode material, which is an unfavorable result. Test data indicate that NCM622 cathode with 5 at% silicon has a capacity of 148.8 mAh g −1 after 100 cycles at 1/3 C, approximately 10 mAh g −1 lower than the virgin. Despite being relatively mild, the adverse influence of silicon impurity in hydrometallurgical recycling still requires attention.

25 ENERGY STORAGE↗
DOE OSTI2024

Underpotential Deposition of 3D Transition Metals: Versatile Electrosynthesis of Single-Atom Catalysts on Oxidized Carbon Supports

Use of single-atom catalysts (SACs) has become a popular strategy for tuning activity and selectivity toward specific pathways. However, conventional SAC synthesis methods require high temperatures and pressures, complicated procedures, and expensive equipment. Recently, underpotential deposition (UPD) has been investigated as a promising alternative, yielding high-loading SAC electrodes under ambient conditions and within minutes. Yet only few studies have employed UPD to synthesize SACs, and all have been limited to UPD of Cu. In this work, a flexible UPD approach for synthesis of mono- and bi-metallic Cu, Fe, Co, and Ni SACs directly on oxidized, commercially available carbon electrodes is reported. The UPD mechanism is investigated using in situ X-ray absorption spectroscopy and, finally, the catalytic performance of a UPD-synthesized Co SAC is assessed for electrochemical nitrate reduction to ammonia. In conclusion, the findings expand upon the usefulness and versatility of UPD for SAC synthesis, with hopes of enabling future research toward realization of fast, reliable, and fully electrified SAC synthesis processes.

36 MATERIALS SCIENCE↗
DOE OSTI2020

Ammonothermal Synthesis, Crystal Structure, and Vibrational Properties of the Doubly Deprotonated Calcium Guanidinate, CaC(NH) 3

We report the synthesis of the doubly deprotonated calcium guanidinate, CaC(NH) 3 , from liquid ammonia and its crystal structure as determined from powder X‐ray and neutron diffraction, and confirmed using CNH elemental analysis and infrared (IR) spectroscopy data. CaC(NH) 3 crystallizes in the hexagonal system with space group P 6 3 / m (no. 176) with Z = 2 and a = 5.2666(13) Å, c = 6.5881(6) Å and V = 158.25(4) Å 3 . We also compare the structural similarities and differences of this phase with the isotypical strontium and ytterbium compounds.

Ogutu, George↗
DOE OSTI2023

Nitrogen and phosphorous acquisition strategies drive coexistence patterns among archaeal lineages in soil

Soil represents the largest reservoir of Archaea on Earth. Present-day archaeal diversity in soils globally is dominated by members of the class Nitrososphaeria. The evolutionary radiation of this class is thought to reflect adaptations to a wide range of temperatures, pH, and other environmental conditions. However, the mechanisms that govern competition and coexistence among Nitrososphaeria lineages in soil remain poorly understood. Here we show that predominant soil Nitrososphaeria lineages compose a patchwork of gene inventory and expression profiles for ammonia, urea, and phosphate utilization. In contrast, carbon fixation, respiration, and ATP synthesis genes are conserved and expressed consistently among predominant phylotypes across 12 major evolutionary lineages commonly found in soil. In situ gene expression profiles closely resemble pure culture reference strains under optimal growth conditions. Together, these results reveal resource-based coexistence patterns among Nitrososphaeria lineages and suggest complementary ecophysiological niches associated with differential nutrient acquisition strategies among globally predominant archaeal lineages in soil.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Self-Assembling Cell-Free Systems for Scalable Bioconversion

This project focused on developing cell-free systems to directly express multi-enzyme catalysts and perform CO2 bioconversions for industrial chemical production. The use of cell-free expression (CFE) systems derived from bacterial lysates is emerging as a promising approach for biomanufacturing. CFEs are genetically programmable, permit the expression of toxic enzymes, and allow for rapid prototyping of metabolic pathways. Research Contributions: 1. Understanding the Area Investigated: This research advances the understanding of cell-free systems by demonstrating their capability to perform complex multi-enzyme reactions. By directly expressing multi-gene systems, CFEs avoid the high costs and inefficiencies associated with producing and purifying enzymes for multi-step pathways. 2. Technical Effectiveness and Economic Feasibility: The project successfully engineered a CFE-based multienzyme biocatalyst for the de novo synthesis of serine and glycine from CO2 equivalents (formate and bicarbonate) and ammonia. This method achieved a 30% conversion rate of formate into these industrially important amino acids. Additionally, an 8-gene CFE biocatalyst was developed to produce malate, conserving 43% of carbon that would otherwise be lost as CO2. This approach has the potential to reach higher carbon efficiency than microbial production. 3. Public Benefit: The cell-free production of chemicals like serine, glycine, and malate using electrochemically generated formate could significantly reduce CO2 emissions. For example, satisfying the global malate market with this method could avoid approximately 400,000 tons of CO2 emissions annually. This work demonstrates the potential of CFE systems to produce platform chemicals, contributing to environmental sustainability and reducing reliance on petrochemicals. Future Prospects: The CFE-based biocatalyst process could be extended to produce a variety of chemicals, including other industrial di-acids, aromatics, terpenes, alcohols, and polymers. This project showcases the capabilities of cell-free expression systems for prototyping carbon-conserving pathways and sustainably bioproducing platform chemicals, marking a significant step towards economically-viable industrial processes.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗