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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 541 records · Page 30
All sourcesNASA NTRSDOE OSTI
DOE OSTI2023

Differences in spore size and atmospheric survival shape stark contrasts in the dispersal dynamics of two closely related fungal pathogens

A frequently ignored but critical aspect of microbial dispersal is survival in the atmosphere. Here, we exposed spores of two closely related, morphologically dissimilar, and economically important fungal pathogens to typical atmospheric environments and modeled their movement in the troposphere. Alternaria solani conidia are nearly 10 times larger than A. alternata conidia, but in our experiments, most died within 24 h, while over half of A. alternata conidia remained viable on day 12. Next, we modeled the movement of spores across North America. We predict 99% of the larger A. solani conidia settle within 24 h, with a maximum dispersal distance of 100 km. By contrast, most A. alternata conidia remain airborne for more than 12 days, and dispersal over long distances(2000 km) is likely. Counterintuitively, the larger A. solani conidia survive poorly, as compared to smaller A. alternata conidia, but also land sooner and move over shorter distances.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Tapping the treasure trove of atypical phages

With advancements in genomics technologies, a vast diversity of ‘atypical’ phages, that is, with single-stranded DNA or RNA genomes, are being uncovered from different ecosystems. Though these efforts have revealed the existence and prevalence of these nonmodel phages, computational approaches often fail to associate these phages with their specific bacterial host(s), while the lack of methods to isolate these phages has limited our ability to characterize infectivity pathways and new gene function. In this review, we call for the development of generalizable experimental methods to better capture this understudied viral diversity via isolation and study them through gene-level characterization and engineering. Establishing a diverse set of new ‘atypical’ phage model systems has the potential to provide many new biotechnologies, including potential uses of these atypical phages in halting the spread of antibiotic resistance and engineering of microbial communities for beneficial outcomes.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Compositional and temporal division of labor modulates mixed sugar fermentation by an engineered yeast consortium

Abstract Synthetic microbial communities have emerged as an attractive route for chemical bioprocessing. They are argued to be superior to single strains through microbial division of labor (DOL), but the exact mechanism by which DOL confers advantages remains unclear. Here, we utilize a synthetic Saccharomyces cerevisiae consortium along with mathematical modeling to achieve tunable mixed sugar fermentation to overcome the limitations of single-strain fermentation. The consortium involves two strains with each specializing in glucose or xylose utilization for ethanol production. By controlling initial community composition, DOL allows fine tuning of fermentation dynamics and product generation. By altering inoculation delay, DOL provides additional programmability to parallelly regulate fermentation characteristics and product yield. Mathematical models capture observed experimental findings and further offer guidance for subsequent fermentation optimization. This study demonstrates the functional potential of DOL in bioprocessing and provides insight into the rational design of engineered ecosystems for various applications.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2026

High yield production of 3-hydroxypropionic acid using Issatchenkia orientalis

Biomanufacturing provides a more sustainable alternative to fossil-based chemical manufacturing. 3-Hydroxypropionic acid (3HP) is a top Department of Energy value-added chemical and precursor to bioplastics, yet cost-effective microbial production remains elusive. Here, we establish the acid-tolerant yeast Issatchenkia orientalis as a robust host for low-pH 3HP biosynthesis. Genome-scale modeling identifies the β-alanine pathway as optimal, offering the highest theoretical yield and lowest oxygen requirement. Thermodynamic analysis confirms its favorability under acidic conditions. Using sequence similarity network analysis, we discover highly active aspartate 1-decarboxylase (PAND), β-alanine-pyruvate aminotransferase (BAPAT), and 3HP dehydrogenase (YDFG), which significantly improve the pathway efficiency. Next, to further elevate the production, pathway optimization through multi-copy PAND integration, byproduct elimination (knockouts of pyruvate decarboxylase and glycerol-3-phosphate dehydrogenase), and reinforcement of aspartate flux by overexpression of pyruvate carboxylase and aspartate amino transferase improves the titer to 29 g/L in shake flasks. Fed-batch fermentation at pH 4 with low-cost corn steep liquor medium further increases the production to 92 g/L with 0.7 g/g yield and 0.55 g/L/h productivity. Techno-economic analysis indicates that such performance could potentially enable a financially viable process for sustainable acrylic acid production. This work establishes I. orientalis as a next-generation platform for cost-effective 3HP production and paves the way toward industrial commercialization.

Biotechnology↗
DOE OSTI2023

High-throughput genetic engineering of nonmodel and undomesticated bacteria via iterative site-specific genome integration

Efficient genome engineering is critical to understand and use microbial functions. Despite recent development of tools such as CRISPR-Cas gene editing, efficient integration of exogenous DNA with well-characterized functions remains limited to model bacteria. Here, we describe serine recombinase–assisted genome engineering, or SAGE, an easy-to-use, highly efficient, and extensible technology that enables selection marker–free, site-specific genome integration of up to 10 DNA constructs, often with efficiency on par with or superior to replicating plasmids. SAGE uses no replicating plasmids and thus lacks the host range limitations of other genome engineering technologies. We demonstrate the value of SAGE by characterizing genome integration efficiency in five bacteria that span multiple taxonomy groups and biotechnology applications and by identifying more than 95 heterologous promoters in each host with consistent transcription across environmental and genetic contexts. We anticipate that SAGE will rapidly expand the number of industrial and environmental bacteria compatible with high-throughput genetics and synthetic biology.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Global distribution, formation and fate of mineral‐associated soil organic matter under a changing climate: A trait‐based perspective

Abstract Soil organic matter (SOM) is the largest actively cycling reservoir of terrestrial carbon (C), and the majority of SOM in Earth's mineral soils (~65%) is mineral‐associated organic matter (MAOM). Thus, the formation and fate of MAOM can exert substantial influence on the global C cycle. To predict future changes to Earth's climate, it is critical to mechanistically understand the processes by which MAOM is formed and decomposed, and to accurately represent this process‐based understanding in biogeochemical and Earth system models. In this review, we use a trait‐based framework to synthesize the interacting roles of plants, soil micro‐organisms, and the mineral matrix in regulating MAOM formation and decomposition. Our proposed framework differentiates between plant and microbial traits that influence total OM inputs to the soil (‘feedstock traits’) versus traits that influence the proportion of OM inputs that are ultimately incorporated into MAOM (‘MAOM formation traits’). We discuss how these feedstock and MAOM formation traits may be altered by warming, altered precipitation and elevated carbon dioxide. At a planetary scale, these feedstock and MAOM formation traits help shape the distribution of MAOM across Earth's biomes, and modulate biome‐specific responses of MAOM to climate change. We leverage a global synthesis of MAOM measurements to provide estimates of the total amount of MAOM‐C globally (~840–1540 Pg C; 34%–51% of total terrestrial organic C), and its distribution across Earth's biomes. We show that MAOM‐C concentration is highest in temperate forests and grasslands, and lowest in shrublands and savannas. Grasslands and croplands have the highest proportion of soil organic carbon (SOC) in the MAOM fraction (i.e. the MAOM‐C:SOC ratio), while boreal forests and tundra have the lowest MAOM‐C:SOC ratio. Drawing on our trait framework, we then review experimental data and posit the effects of climate change on MAOM pools in different biomes. We conclude by discussing how MAOM is integrated into soil C models, and how feedstock and MAOM formation traits may be included in these models. We also summarize the projected fate of MAOM under climate change scenarios (Representative Concentration Pathways 4.5 and 8.5) and discuss key model uncertainties. Read the free Plain Language Summary for this article on the Journal blog.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2023

Biochemical Process Modeling and Simulation (BPMS)

The Biochemical Process Modeling and Simulation project aims to reduce the cost and time of research by applying theory, modeling, and simulation to the most relevant bottlenecks in the biochemical process. We use molecular modeling, quantum mechanics, metabolic modeling, fluid dynamics, and reaction-diffusion methods in close collaboration with pretreatment, hydrolysis, upgrading, and TEA. The project's outcomes are increased yields and efficiency of the biochemical process, added value to products, and reduced price of fuels by specifically targeting catalytic efficiency, reactor design, enzyme efficiency, and microbial design. We work closely with experimental projects to identify problems and iterate with experiments to find and refine solutions. By working with experimentalists, we decide on problems that can be solved with simulation that could otherwise not be solved or would take too long with experiment alone to reach BETO's targets. Over the years, we have produced solutions that have resulted in determining the most likely fatty-acid derivative for passive transport out of bacteria that upgrade biomass, and we have also designed enzyme mutations for enhanced lignin upgrading. Metabolic models have been developed to tune the activity of 2,3 butanediol production for the 2030 target. A computational method to deliver understanding of how complex omics data can be interpreted in the metabolic pathways of organisms used in the Agile Biofoundry. We have found methods to overcome specific barriers and continue to develop those methods. Our reactor studies have guided the design of both the microbes and reactors for aerobic and micro-aerobic production at all scales and have been instrumental in improving the accuracy of techno-economic analysis models. This project is essential in the process of selecting the final processes for 2030 SAF production targets. More specifically, recently, we have: 1) Predicted the strength of the basic structural interactions in commodity plastics to provide guidance for plastics upcycling strategies. 2) Developed computational tool to improve the characterization of lignin-derived compounds 3) Developed new methodologies to enable Machine Learning-based Directed Evolution for protein engineering. 4) Developed Machine Learning methods to predict protein promiscuity and mutations to further improve microbial and enzymatic driven processes and demonstrated the utility of ML approaches to engineering proteins from sparse experimental datasets. 5) Developed new methods to enable high-fidelity simulation of aerobic fermentation at industrial scale and resolving mismatch of time scales through subcycling/operator splitting 7) Identified the difficulty in preventing local high-oxygen conditions in industrial bubble columns, which leads to less-desirable acetoin production, suggesting future research directions in alternative reactor configurations (e.g loop reactors, shallow-channel reactors).

BIOMASS FUELS↗
DOE OSTI2025

Structured interactions drive abrupt transitions in the spatial organization of microbial communities

Bacteria possess diverse mechanisms to regulate their motility in response to environmental and physiological signals, enabling them to navigate complex habitats and adapt their behavior. Some of these mechanisms are species specific and enable cells to modulate their movement based on the ecological identity of neighboring species. Here, we introduce a model in which bacteria interact via local signals that either enhance or suppress the motility of neighboring cells depending on species type. Through large-scale simulations and a coarse-grained stochastic model, we demonstrate the emergence of a sharp transition driven by nucleation processes: increasing the density of motility-suppressing interactions drives the system from a fully mixed, motile phase to a state characterized by large, stationary bacterial clusters. Remarkably, in systems with a large number of interacting species, this transition can be triggered solely by altering the structure of the motility-regulation interaction matrix while maintaining species and interaction densities constant. In particular, we find that heterogeneous and modular interactions promote the transition more readily than homogeneous random ones. These findings add a dimension to the theory of motility-induced phase separation and contribute to the ongoing effort to understand microbial interactions, suggesting that structured, nonrandom ones may be key to reproducing commonly observed spatial patterns in microbial communities.

bacterial communities↗
DOE OSTI2023

Development of molecular cluster models to probe pyrite surface reactivity

Abstract The recent discovery that anaerobic methanogens can reductively dissolve pyrite and utilize dissolution products as a source of iron and sulfur to meet their biosynthetic demands for these elements prompted the development of atomic‐scale nanoparticle models, as maquettes of reactive surface sites, for describing the fundamental redox steps that take place at the mineral surface during reduction. The given report describes our computational approach for modeling n (FeS 2 ) nanoparticles originated from mineral bulk structure. These maquettes contain a comprehensive set of coordinatively unsaturated Fe (II) sites that are connected via a range of persulfide (S 2 2− ) ligation. In addition to the specific maquettes with n = 8, 18, and 32 FeS 2 units, we established guidelines for obtaining low‐energy structures by considering the pattern of ionic, covalent, and magnetic interactions among the metal and ligand sites. The developed models serve as computational nano‐reactors that can be used to describe the reductive dissolution mechanism of pyrite to better understand the reactive sites on the mineral, where microbial extracellular electron‐transfer reactions can occur.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2020

Stimulation of soil respiration by elevated CO 2 is enhanced under nitrogen limitation in a decade-long grassland study

Whether and how CO 2 and nitrogen (N) availability interact to influence carbon (C) cycling processes such as soil respiration remains a question of considerable uncertainty in projecting future C–climate feedbacks, which are strongly influenced by multiple global change drivers, including elevated atmospheric CO 2 concentrations (eCO 2 ) and increased N deposition. However, because decades of research on the responses of ecosystems to eCO 2 and N enrichment have been done largely independently, their interactive effects on soil respiratory CO 2 efflux remain unresolved. Here, we show that in a multifactor free-air CO 2 enrichment experiment, BioCON (Biodiversity, CO 2 , and N deposition) in Minnesota, the positive response of soil respiration to eCO 2 gradually strengthened at ambient (low) N supply but not enriched (high) N supply for the 12-y experimental period from 1998 to 2009. In contrast to earlier years, eCO 2 stimulated soil respiration twice as much at low than at high N supply from 2006 to 2009. In parallel, microbial C degradation genes were significantly boosted by eCO 2 at low but not high N supply. Further, incorporating those functional genes into a coupled C–N ecosystem model reduced model parameter uncertainty and improved the projections of the effects of different CO 2 and N levels on soil respiration. If our observed results generalize to other ecosystems, they imply widely positive effects of eCO 2 on soil respiration even in infertile systems.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2021

Aerobic bacterial methane synthesis

Significance Observations summarized herein contribute to an ongoing paradigm shift in microbial ecology, documenting an emergent property of ecosystem function that further challenges the perception that biogenic methane (CH 4 ) production is strictly an anaerobic process. Relevant metabolites, a model bacterial isolate, gene, and enzyme are identified, and we show how this property can conceivably be broadly distributed in the biosphere and contribute to global CH 4 emissions. Scientifically, this study will enable lines of investigation that will expand our understanding of CH 4 synthesis and emission in nature and illustrates how CH 4 synthesis may actually serve as a nexus for the C and N cycles in nature.

Wang, Qian↗
DOE OSTI2025

Microbes display broad diversity in cobamide preferences

ABSTRACT Cobamides, the vitamin B 12 (cobalamin) family of cofactors, are used by most organisms but produced by only a fraction of prokaryotes, and are thus considered key shared nutrients among microbes. Cobamides are structurally diverse, with multiple different cobamides found in most microbial communities. The ability to use different cobamides has been tested for several bacteria and microalgae, and nearly all show preferences for certain cobamides. This approach is limited by the commercial unavailability of cobamides other than cobalamin. Here, we have extracted and purified seven commercially unavailable cobamides to characterize bacterial cobamide preferences based on growth in specific cobamide-dependent conditions. The tested bacteria include engineered strains of Escherichia coli , Sinorhizobium meliloti , and Bacillus subtilis expressing native or heterologous cobamide-dependent enzymes, cultured under conditions that functionally isolate specific cobamide-dependent processes such as methionine synthesis. Comparison of these results to those of previous studies of diverse bacteria and microalgae revealed that a broad diversity of cobamide preferences exists not only across different organisms but also between different cobamide-dependent metabolic pathways within the same organism. The microbes differed in the cobamides that support growth most efficiently, cobamides that do not support growth, and the minimum cobamide concentrations required for growth. The latter differ by up to four orders of magnitude across organisms from different environments and by up to 20-fold between cobamide-dependent enzymes within the same organism. Given that cobamides are shared, required for use of specific growth substrates, and essential for central metabolism in certain organisms, cobamide preferences likely impact community structure and function. IMPORTANCE Nearly all bacteria are found in microbial communities with tens to thousands of other species. Molecular interactions such as metabolic cooperation and competition are key factors underlying community assembly and structure. Cobamides, the vitamin B 12 family of enzyme cofactors, are one such class of nutrients, produced by only a minority of prokaryotes but required by most microbes. A unique aspect of cobamides is their broad diversity, with nearly 20 structural forms identified in nature. Importantly, this structural diversity impacts growth as most bacteria that have been tested show preferences for specific cobamide forms. We measured cobamide-dependent growth in several model bacteria and compared the results to those of previous analyses of cobamide preference. We found that cobamide preferences vary widely across bacteria, showing the importance of characterizing these aspects of cobamide biology to understand the impact of cobamides on microbial communities.

Mok, Kenny C. (ORCID:0000000252276987)↗
DOE OSTI2023

Exposing new taxonomic variation with inflammation — a murine model-specific genome database for gut microbiome researchers

The murine CBA/J mouse model widely supports immunology and enteric pathogen research. This model has illuminated Salmonella interactions with the gut microbiome since pathogen proliferation does not require disruptive pretreatment of the native microbiota, nor does it become systemic, thereby representing an analog to gastroenteritis disease progression in humans. Despite the value to broad research communities, microbiota in CBA/J mice are not represented in current murine microbiome genome catalogs. Here we present the first microbial and viral genomic catalog of the CBA/J murine gut microbiome. Using fecal microbial communities from untreated and Salmonella-infected, highly inflamed mice, we performed genomic reconstruction to determine the impacts on gut microbiome membership and functional potential. From high depth whole community sequencing (~ 42.4 Gbps/sample), we reconstructed 2281 bacterial and 4516 viral draft genomes. Salmonella challenge significantly altered gut membership in CBA/J mice, revealing 30 genera and 98 species that were conditionally rare and unsampled in non-inflamed mice. Additionally, inflamed communities were depleted in microbial genes that modulate host anti-inflammatory pathways and enriched in genes for respiratory energy generation. Our findings suggest decreases in butyrate concentrations during Salmonella infection corresponded to reductions in the relative abundance in members of the Alistipes. Strain-level comparison of CBA/J microbial genomes to prominent murine gut microbiome databases identified newly sampled lineages in this resource, while comparisons to human gut microbiomes extended the host relevance of dominant CBA/J inflammation-resistant strains. This CBA/J microbiome database provides the first genomic sampling of relevant, uncultivated microorganisms within the gut from this widely used laboratory model. Using this resource, we curated a functional, strain-resolved view on how Salmonella remodels intact murine gut communities, advancing pathobiome understanding beyond inferences from prior amplicon-based approaches. Salmonella-induced inflammation suppressed Alistipes and other dominant members, while rarer commensals like Lactobacillus and Enterococcus endure. The rare and novel species sampled across this inflammation gradient advance the utility of this microbiome resource to benefit the broad research needs of the CBA/J scientific community, and those using murine models for understanding the impact of inflammation on the gut microbiome more generally.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

The Model System Saccharomyces cerevisiae Versus Emerging Non-Model Yeasts for the Production of Biofuels

Microorganisms are effective platforms for the production of a variety of chemicals including biofuels, commodity chemicals, polymers and other natural products. However, deep cellular understanding is required for improvement of current biofuel cell factories to truly transform the Bioeconomy. Modifications in microbial metabolic pathways and increased resistance to various types of stress caused by the production of these chemicals are crucial in the generation of robust and efficient production hosts. Recent advances in systems and synthetic biology provide new tools for metabolic engineering to design strategies and construct optimal biocatalysts for the sustainable production of desired chemicals, especially in the case of ethanol and fatty acid production. Yeast is an efficient producer of bioethanol and most of the available synthetic biology tools have been developed for the industrial yeast Saccharomyces cerevisiae. Non-conventional yeast systems have several advantageous characteristics that are not easily engineered such as ethanol tolerance, low pH tolerance, thermotolerance, inhibitor tolerance, genetic diversity and so forth. Currently, synthetic biology is still in its initial steps for studies in non-conventional yeasts such as Yarrowia lipolytica, Kluyveromyces marxianus, Issatchenkia orientalis and Pichia pastoris. Therefore, the development and application of advanced synthetic engineering tools must also focus on these underexploited, non-conventional yeast species. Herein, we review the basic synthetic biology tools that can be applied to the standard S. cerevisiae model strain, as well as those that have been developed for non-conventional yeasts. In addition, we will discuss the recent advances employed to develop non-conventional yeast strains that are efficient for the production of a variety of chemicals through the use of metabolic engineering and synthetic biology.

09 BIOMASS FUELS↗
DOE OSTI2022

Ecosystems and Networks Integrated with Genes and Molecular Assemblies (ENIGMA). Final report

The goal of the Stahl lab is to use a combination of isotopic, genomic, and activity-based analyses in the field, of selected field isolates, and defined synthetic communities to identify sources and sinks of different nitrogen species in the field, focusing on the FRC at the ORNL. The most general research objective is to characterize variants in the pathway for denitrification among field isolates and synthetic microbial assemblies as a foundation for predicting the biotic and abiotic controls of nitrogen and carbon transformations in the field. Specifically, we have focused on characterizing environmental controls of nitrous oxide emissions, an important greenhouse gas and ozone depleting molecule. Studies have mapped the flux of nitrous oxide and controlling genetic elements within the FRC subsurface, indicating the importance of different enzyme variants in nitrous oxide production and consumption in controlling surface emissions. We have developed methods to quantify and catalog the isotopic signatures imparted on nitrous oxide by different nitrous oxide reductase variants in isolated organisms and model synthetic communities. We have developed methods to quantify and catalog the nitrous oxide affinity of field relevant isolates and their biomass yield when respiring nitrous oxide on different carbon substrates. We continue to expand the collection of field-relevant microbial isolates within ENIGMA by focused enrichment and isolation from field material. These objectives have been facilitated by the development of new technologies, including: 1) the development of a multiplexed automated optical density measurement system for quantifying growth kinetics of microorganisms dependent on gaseous substrates in monoculture or gas exchange in synthetic communities and 2) the development of a microcalorimetric analysis format to quantify the impact of environmental stress on microbial maintenance energy. Since field relevant processes are mediated by assemblies of organisms interacting with each other and their local environment, we have also collaborated with other ENIGMA investigators on the assembly and characterization of synthetic communities. These studies have evaluated the role of organisms lacking a complete pathway for denitrification in the control of field processes, measuring process rates of assemblies of organisms that individually lack the full denitrification pathway but mediate the complete denitrification when growing together. These efforts have so far resulted in the publication of 18 peer reviewed papers ranging from ecological and physiological drivers of pathway optimization to field-scale process characterization.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2025

Ecohydrological controls on root and microbial respiration in the East River watershed of Colorado

The main objective of this project was to conduct exploratory work to quantify how snow and rain water inputs influence the CO2 coming from the soil surface (soil CO2 flux), and its plant and microbial sources, in the East River watershed, near Crested Butte, Colorado. Knowledge gained from this effort laid the groundwork for a more comprehensive (ongoing) follow-on grant that is using a combination of experiments, field observations, machine learning and simulation modeling to fully disentangle these relationships. New field measurements were made at four locations along an elevational transect on Snodgrass Mountain at Rocky Mountain Biological Laboratory. These sites were chosen to differ in total snowpack and water table depth, and included both deciduous (aspen) and evergreen (spruce/fir) forest types. We used automated measurements of soil CO2 concentrations to quantify the total soil CO2 flux, and the vertical CO2 production within the soil profile at each site. Isotope (radiocarbon, 14C) measurements determined how much of the CO2 emitted from the soil surface came from plant respiration (root metabolism) versus microbial respiration (decomposition of soil organic matter) sources. Supporting data on plant phenology and microbial dynamics provided context for the observed variation in respiration sources. This work was motivated by our overarching hypothesis that quantifying belowground plant and microbial processes separately, and how they are influenced by snow and rain inputs, is necessary for understanding and predicting how the East River watershed ecosystems will respond to future environmental change.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2021

Efficient production of oxidized terpenoids via engineering fusion proteins of terpene synthase and cytochrome P450

The functionalization of terpenes using cytochrome P450 enzymes is a versatile route to the production of useful derivatives that can be further converted to value-added products. Many terpenes are hydrophobic and volatile making their availability as a substrate for P450 enzymes significantly limited during microbial production. In this study, we developed a strategy to improve the accessibility of terpene molecules for the P450 reaction by linking terpene synthase and P450 together. As a model system, fusion proteins of 1,8-cineole synthase (CS) and P450 cin were investigated and it showed an improved hydroxylation of the monoterpenoid 1,8-cineole up to 5.4-fold. Structural analysis of the CS-P450 cin fusion proteins by SEC-SAXS indicated a dimer formation with preferred orientations of the active sites of the two domains. We also applied the enzyme fusion strategy to the oxidation of a sesquiterpene epi-isozizaene and the fusion enzymes significantly improved albaflavenol production in engineered E. coli. From the analysis of positive and negative examples of the fusion strategy, we proposed key factors in structure-based prediction and evaluation of fusion enzymes. Developing fusion enzymes for terpene synthase and P450 presents an efficient strategy toward oxidation of hydrophobic terpene compounds. This strategy could be widely applicable to improve the biosynthetic titer of the functionalized products from hydrophobic terpene intermediates.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2026

metagRoot: a comprehensive database of protein families associated with plant root microbiomes

The plant root microbiome is vital in plant health, nutrient uptake, and environmental resilience. To explore and harness this diversity, we present metagRoot, a specialized and enriched database focused on the protein families of the plant root microbiome. MetagRoot integrates metagenomic, metatranscriptomic, and reference genome-derived protein data to characterize 71 091 enriched protein families, each containing at least 100 sequences. These families are annotated with multiple sequence alignments, CRISPR elements, hidden Markov models, taxonomic and functional classifications, ecosystem and geolocation metadata, and predicted 3D structures using AlphaFold2. MetagRoot is a powerful tool for decoding the molecular landscape of root-associated microbial communities and advancing microbiome-informed agricultural practices by enriching protein family information with ecological and structural context. The database is available at https://pavlopoulos-lab.org/metagroot/ or https://www.metagroot.org.

Chasapi, Maria N↗