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At least 199 records · Page 11
All sourcesNASA NTRSDOE OSTI
DOE OSTI2025

Steric effects of central dogma processes on the compaction and segregation of bacterial nucleoids

The bacterial cytoplasm is characterized by a distinctive membrane-less organelle, the nucleoid, which harbors the chromosomal DNA. Here, we investigate the steric effects of dynamic processes associated with transcription and translation on the structure of this organelle using coarse-grained molecular dynamics simulations that incorporate out-of-equilibrium reactions. Our model captures the scale of the entire cell and incorporates a reaction-diffusion system for ribosomes and polyribosomes, coupling their nonequilibrium kinetics to DNA excluded volume interactions. Our findings demonstrate that out-of-equilibrium reactions increase the size of the nucleoid and the number of ribosomes in this subcellular region. In addition, we show that nucleoid size is proportional to transcriptional activity. Our model reproduces the time-dependent change in nucleoid size observed in rifampicin treatment experiments, where the pool of polyribosomes is depleted. Furthermore, we find that these active processes are essential for complete sister chromosome separation and correct nucleoid positioning within the cell. Overall, our study reveals the effects of the central dogma processes on the internal organization and localization of bacterial nucleoids.

Chang, Mu-Hung [Univ. of Tennessee, Knoxville, TN ↗
DOE OSTI2021

Bacterial microcompartments: tiny organelles with big potential

Organization of metabolic processes within the space of a cell is critical for the survival of many organisms. In bacteria, spatial organization is achieved via proteinaceous organelles called bacterial microcompartments, which encapsulate pathway enzymes, substrates, and co-factors to drive the safe and efficient metabolism of niche carbon sources. Microcompartments are self-assembled from shell proteins that encapsulate a core comprising various enzymes. Here, this review discusses how recent advances in understanding microcompartment structure and assembly have informed engineering efforts to repurpose compartments and compartment-based structures for non-native functions. These advances, both in understanding of the native structure and function of compartments, as well as in the engineering of new functions, will pave the way for the use of these structures in bacterial cell factories.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Geology and elevation shape bacterial assembly in Antarctic endolithic communities

Ice free areas of continental Antarctica are among the coldest and driest environments on Earth, and yet, they support surprisingly diverse and highly adapted microbial communities. Endolithic growth is one of the key adaptations to such extreme environments and often represents the dominant life-form. Despite growing scientific interest, little is known of the mechanisms that influence the assembly of endolithic microbiomes across these harsh environments. Here, we used metagenomics to examine the diversity and assembly of endolithic bacterial communities across Antarctica within different rock types and over a large elevation range. While granite supported richer and more heterogeneous communities than sandstone, elevation had no apparent effect on taxonomic richness, regardless of rock type. Conversely, elevation was clearly associated with turnover in community composition, with the deterministic process of variable selection driving microbial assembly along the elevation gradient. The turnover associated with elevation was modulated by geology, whereby for a given elevation difference, turnover was consistently larger between communities inhabiting different rock types. Overall, selection imposed by elevation and geology appeared stronger than turnover related to other spatially-structured environmental drivers. Our findings indicate that at the cold-arid limit of life on Earth, geology and elevation are key determinants of endolithic bacterial heterogeneity. This also suggests that warming temperatures may threaten the persistence of such extreme-adapted organisms.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2023

Revising the dynamic energy budget theory with a new reserve mobilization rule and three example applications to bacterial growth

Dynamic energy budget (DEB) theory has been applied to model a wide range of organisms, including microbes. In the standard DEB model, biomass is partitioned into reserve and structural compartments, where reserve biomass is mobilized in a pseudolinear manner (while the reserve biomass density, defined as the ratio between reserve and structural biomass, decays linearly) to drive maintenance and the growth of structural biomass (and extracellular enzyme production if it is considered). However, the linear dynamics of the reserve biomass density makes the standard DEB model incapable of explaining the slowdown of microbial growth at high reserve density that is caused by macromolecular crowding effect which reduces biochemical reaction rates (a typical situation occurs when microbes are experiencing severe moisture stress) and is inconsistent with the observation that intracellular enzymatic reactions generally follow non-linear kinetics. By partitioning biomass into reserve, kinetic, and structural compartments, we show here that the Equilibrium Chemistry Approximation (ECA) kinetics can be used to represent enzymatically catalyzed reserve biomass mobilization that can then drive the kinetic and structural biomass synthesis. This revised DEB model better represents the tradeoff in ribosome allocation for structural growth and internal enzyme production, is structurally compatible with metabolic models of cell individuals, and includes the standard DEB model and the popular compromise model as special cases for representing population growth. We then applied the revised DEB model to interpret components of bacterial respiration, their dependence on substrate availability, and emergent microbial carbon use efficiency dynamics for an exponentially growing population. We found that the revised DEB model enables a better understanding of bacterial substrates use (carbon in our examples) than that can be derived from a few other models in the literature. In particular, the revised DEB model explains why carbon use efficiency may first increase, then plateau, and finally decrease with growth rate (and substrate uptake rate), as a function of proteomics. Additionally, the revised DEB model explains why the kinetic biomass compartment needs to be divided to reasonably incorporate proteomic control of microbial growth.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Moisture-mineral interactions drive bacterial and organic matter turnover in glacier-sourced riparian sediments undergoing pedogenesis

Glacial recession is occurring at unprecedented rates resulting in increased sediment accumula-tions in some riverine ecosystems. Increased sediment deposition has implications for ecosystem stability (e.g., floods and river paths) and environmental services (e.g., carbon sequestration). Soils and sediments have an enormous potential to retain carbon (C), predominantly due to sorp-tion to mineral surfaces. However, C persistence may be sensitive to climate-change induced temperature and moisture variations. We coupled ultrahigh resolution organic matter composition classification with bacterial characterization and respiration measurements to test the combined effects of temperature (4 vs 20°C) and moisture (50 vs 100% water-filled pore space) on C turn-over in sediments maintained under different mineralogical conditions (illite-amended vs non-amended). Here we show that the inhibition of CO 2 emissions from the combined effect of in-creased moisture content and illite was reflected in the turnover of key molecular signatures, such as the nominal oxidation state of C, often irrespective of temperature. However, shifts in bacteri-al communities from a coupled moisture-mineral interaction, were temperature-dependent. Our results highlight the importance of moisture in driving mineral-organic interactions and suggest that C in clay-rich, water-saturated sediments is both thermodynamically unfavorable and miner-al-protected from microbial consumption.

58 GEOSCIENCES↗
DOE OSTI2021

In Situ, Protein-Mediated Generation of a Photochemically Active Chlorophyll Analogue in a Mutant Bacterial Photosynthetic Reaction Center

All possible natural amino acids have been substituted for the native LeuL185 positioned near the B-side bacteriopheophytin (H B ) in the bacterial reaction center (RC) from Rhodobacter sphaeroides. Additional mutations are present that enhance electron transfer to the normally inactive B-side cofactors. About half the isolated RCs with Glu at L185 contain a magnesium chlorin (C B ) in place of H B . The chlorin is not the common BChl a oxidation product 3-desvinyl-3-acetyl chlorophyll a with a C-C bond in ring D and a C=C bond in ring B, but has properties consistent with reversal of these bond orders, giving 17,18-didehydro BChl a. In such RCs, charge-separated state P + C B – forms with ~5% yield. The other half of the GluL185-containing RCs have a bacteriochlorophyll a (BChl a) denoted β B in place of H B . Residues His, Asp, Asn and Gln at L185 yield RCs with ≥85% β B in the H B site, while most other amino acids result in RCs that retain H B (≥95%). To our knowledge, neither bacterial RCs that harbor five BChl a and one chlorophyll analog nor ones with six BChl a have been reported previously. The finding that altering the local environment within a cofactor binding site of a transmembrane complex leads to in situ generation of a photoactive chlorin with an unusual ring oxidation pattern suggests new strategies for amino-acid control over pigment type at specific sites in photosynthetic proteins.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2026

Nanosecond Structure of Radical Pair Intermediates from High-Frequency Quantum Oscillations: Insight into the Q A •– to Q B Electron Transfer Step in Purple Bacterial Photosynthesis

We demonstrate the validity of our approach to deduce, from the anisotropy of quantum oscillations, the geometry of short-lived radical pair intermediates in photosynthesis. A global fit of a two-dimensional W-band (94 GHz) electron paramagnetic resonance (EPR) experiment provides the same global minimum values for the geometry of the A-side radical pair P 700 •+ A 1A •− in photosystem I (PSI) as observed in a previous Q-band (34 GHz) EPR study, yet with a significantly increased convergence rate of 62%. This demonstrates that the global fit yields the correct radical pair geometry even at Q-band frequencies. With this information, we revisit our previous Q-band study of the cofactor arrangement of P 865 •+ Q A •− , the stabilized charge-separated state in purple bacterial reaction centers (RCs). Analysis of calculated two-dimensional data sets of P 865 •+ Q A •− reveals that the quantum oscillation technique is unaffected by a mirror ambiguity in disordered solids and thus can provide unambiguous solutions for all five Euler angles of the radical pair geometry. This enables us to elucidate the Q A •− to Q B electron transfer step in purple bacterial photosynthesis, the subject of controversial discussions for more than 25 years. Our results show that this electron transfer step involves a gating mechanism requiring a 60° rotation of the headgroup of Q A •− in its binding pocket.

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

Release Factor Inhibiting Antimicrobial Peptides Improve Nonstandard Amino Acid Incorporation in Wild-type Bacterial Cells

We report a tunable chemical genetics approach for enhancing genetic code expansion in different wild-type bacterial strains that employs apidaecin-like, anti-microbial peptides observed to temporarily sequester and thereby inhibit Release Factor 1 (RF1). In a concentration-dependent matter, these peptides granted a conditional lambda phage resistance to a recoded Escherichia coli strain with non-essential RF1 activity and promoted multi-site non-standard amino acid (nsAA) incorporation at in-frame amber stop codons in vivo and in vitro. When exogenously added, the peptides stimulated specific nsAA incorporation in a variety of sensitive, wild-type (RF1+) strains including Agrobacterium tumefaciens, a species in which nsAA incorporation has not been previously reported. Improvement in nsAA incorporation was typically 2–15-fold in E. coli BL21, MG1655, DH10B strains and A. tumefaciens with the >20-fold improvement observed in probiotic E. coli Nissle 1917. In-cell expression of these peptides promoted multi-site nsAA incorporation in transcripts with up to 6 amber codons, with a >35-fold increase in BL21 showing moderate toxicity. Leveraging this RF1 sensitivity allowed multiplexed partial recoding of MG1655 and DH10B that rapidly resulted in resistant strains that showed an additional ~2 fold boost to nsAA incorporation independent of the peptide. Lastly, in-cell expression of an apidaecin-like peptide library allowed the discovery of new peptide variants with reduced toxicity that still improved multi-site nsAA incorporation >25-fold. In parallel to genetic reprogramming efforts, these new approaches can facilitate genetic code expansion technologies in a variety of wild-type bacterial strains.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Metabolomics Analysis of Bacterial Pathogen Burkholderia thailandensis and Mammalian Host Cells in Co-culture

The Tier 1 HHS/USDA Select Agent Burkholderia pseudomallei is a bacterial pathogen that is highly virulent when introduced into the respiratory tract and intrinsically resistant to many antibiotics. Transcriptomic- and proteomic-based methodologies have been used to investigate mechanisms of virulence employed by B. pseudomallei and Burkholderia thailandensis, a convenient surrogate; however, analysis of the pathogen and host metabolomes during infection is lacking. Changes in the metabolites produced can be a result of altered gene expression and/or post-transcriptional processes. Thus, metabolomics complements transcriptomics and proteomics by providing a chemical readout of a biological phenotype, which serves as a snapshot of an organism’s physiological state. However, the poor signal from bacterial metabolites in the context of infection poses a challenge in their detection and robust annotation. In this work, we coupled mammalian cell culture-based metabolomics with feature-based molecular networking of mono- and co-cultures to annotate the pathogen’s secondary metabolome during infection of mammalian cells. These methods enabled us to identify several key secondary metabolites produced by B. thailandensis during infection of airway epithelial and macrophage cell lines. Additionally, the use of in silico approaches provided insights into shifts in host biochemical pathways relevant to defense against infection. Using chemical class enrichment analysis, for example, we identified changes in a number of host-derived compounds including immune lipids such as prostaglandins, which were detected exclusively upon pathogen challenge. Taken together, our findings indicate that co-culture of B. thailandensis with mammalian cells alters the metabolome of both pathogen and host and provides a new dimension of information for in-depth analysis of the host–pathogen interactions underlying Burkholderia infection.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2025

Designing Peptide Fossils That Model the Evolution of the Bacterial Ferredoxin Fold

Electron transfer coupled to redox chemistry is at the heart of metabolism. The proteins responsible for moving electrons (protein electron carriers) must have emerged at the origin of life. The small iron–sulfur-binding bacterial ferredoxins were likely among these first proteins. Embedded within the ferredoxin sequence and structure is a symmetry that points to an ancient gene duplication event. Little is understood about the nature of ferredoxins prior to this duplication event or what environmental factors may have driven the selection for more complex forms. The deep-time molecular history of ferredoxins goes back billions of years and cannot be reconstructed by phylogenetic analyses based on amino acid sequences. Here, we use structure-guided protein design to model a fossil half-ferredoxin stage in the evolution of this fold, the semidoxins, and their symmetric full-length counterparts, the symdoxins. Semidoxin designs homodimerize, exhibiting structural, thermodynamic, and electrochemical behaviors in most cases identical to cognate symdoxins. However, the semi- and symdoxin fossil stages behave differently when incorporated into an in vivo electron transfer complementation assay. Both can support bacterial growth dependent on protein expression. Growth rates of bacteria expressing the semidoxins are much more sensitive to oxygen than those of bacteria expressing symdoxins. Motivated by the in vivo functionality of designed semidoxins, we identified putative naturally occurring semidoxins in extant anaerobic microorganisms. This is consistent with the observed in vivo oxygen sensitivity of the semidoxin designs. One natural semidoxin is shown to be folded and redox active. However, it exists as a mixture of monomers and dimers, suggesting a potential connection between semidoxins and even simpler single iron–sulfur cluster-binding peptides.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Routes and rates of bacterial dispersal impact surface soil microbiome composition and functioning

Abstract Recent evidence suggests that, similar to larger organisms, dispersal is a key driver of microbiome assembly; however, our understanding of the rates and taxonomic composition of microbial dispersal in natural environments is limited. Here, we characterized the rate and composition of bacteria dispersing into surface soil via three dispersal routes (from the air above the vegetation, from nearby vegetation and leaf litter near the soil surface, and from the bulk soil and litter below the top layer). We then quantified the impact of those routes on microbial community composition and functioning in the topmost litter layer. The bacterial dispersal rate onto the surface layer was low (7900 cells/cm2/day) relative to the abundance of the resident community. While bacteria dispersed through all three routes at the same rate, only dispersal from above and near the soil surface impacted microbiome composition, suggesting that the composition, not rate, of dispersal influenced community assembly. Dispersal also impacted microbiome functioning. When exposed to dispersal, leaf litter decomposed faster than when dispersal was excluded, although neither decomposition rate nor litter chemistry differed by route. Overall, we conclude that the dispersal routes transport distinct bacterial communities that differentially influence the composition of the surface soil microbiome.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

A catalog of the diversity and ubiquity of bacterial microcompartments

Abstract Bacterial microcompartments (BMCs) are organelles that segregate segments of metabolic pathways which are incompatible with surrounding metabolism. BMCs consist of a selectively permeable shell, composed of three types of structurally conserved proteins, together with sequestered enzymes that vary among functionally distinct BMCs. Genes encoding shell proteins are typically clustered with those for the encapsulated enzymes. Here, we report that the number of identifiable BMC loci has increased twenty-fold since the last comprehensive census of 2014, and the number of distinct BMC types has doubled. The new BMC types expand the range of compartmentalized catalysis and suggest that there is more BMC biochemistry yet to be discovered. Our comprehensive catalog of BMCs provides a framework for their identification, correlation with bacterial niche adaptation, experimental characterization, and development of BMC-based nanoarchitectures for biomedical and bioengineering applications.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2023

Previously uncharacterized rectangular bacterial structures in the dolphin mouth

Much remains to be explored regarding the diversity of uncultured, host-associated microbes. Here, we describe rectangular bacterial structures (RBSs) in the mouths of bottlenose dolphins. DNA staining revealed multiple paired bands within RBSs, suggesting the presence of cells dividing along the longitudinal axis. Cryogenic transmission electron microscopy and tomography showed parallel membrane-bound segments that are likely cells, encapsulated by an S-layer-like periodic surface covering. RBSs displayed unusual pilus-like appendages with bundles of threads splayed at the tips. We present multiple lines of evidence, including genomic DNA sequencing of micromanipulated RBSs, 16S rRNA gene sequencing, and fluorescence in situ hybridization, suggesting that RBSs are bacterial and distinct from the genera Simonsiella and Conchiformibius (family Neisseriaceae), with which they share similar morphology and division patterning. Our findings highlight the diversity of novel microbial forms and lifestyles that await characterization using tools complementary to genomics such as microscopy.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Growth phase estimation for abundant bacterial populations sampled longitudinally from human stool metagenomes

Longitudinal sampling of the stool has yielded important insights into the ecological dynamics of the human gut microbiome. However, human stool samples are available approximately once per day, while commensal population doubling times are likely on the order of minutes-to-hours. Despite this mismatch in timescales, much of the prior work on human gut microbiome time series modeling has assumed that day-to-day fluctuations in taxon abundances are related to population growth or death rates, which is likely not the case. Here, we propose an alternative model of the human gut as a stationary system, where population dynamics occur internally and the bacterial population sizes measured in a bolus of stool represent a steady-state endpoint of these dynamics. We formalize this idea as stochastic logistic growth. We show how this model provides a path toward estimating the growth phases of gut bacterial populations in situ. We validate our model predictions using an in vitro Escherichia coli growth experiment. Finally, we show how this method can be applied to densely-sampled human stool metagenomic time series data. We discuss how these growth phase estimates may be used to better inform metabolic modeling in flow-through ecosystems, like animal guts or industrial bioreactors.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Spatiotemporal development of expanding bacterial colonies driven by emergent mechanical constraints and nutrient gradients

Abstract Bacterial colonies growing on solid surfaces can exhibit robust expansion kinetics, with constant radial growth and saturating vertical expansion, suggesting a common developmental program. Here, we study this process forEscherichia colicells using a combination of modeling and experiments. We show that linear radial colony expansion is set by the verticalization of interior cells due to mechanical constraints rather than radial nutrient gradients as commonly assumed. In contrast, vertical expansion slows down from an initial linear regime even while radial expansion continues linearly. This vertical slowdown is due to limitation of cell growth caused by vertical nutrient gradients, exacerbated by concurrent oxygen depletion. Starvation in the colony interior results in a distinct death zone which sets in as vertical expansion slows down, with the death zone increasing in size along with the expanding colony. Thus, our study reveals complex heterogeneity within simple monoclonal bacterial colonies, especially along the vertical dimension. The intricate dynamics of such emergent behavior can be understood quantitatively from an interplay of mechanical constraints and nutrient gradients arising from obligatory metabolic processes.

Science & Technology - Other Topics↗
DOE OSTI2025

A blueprint for biomolecular condensation driven by bacterial microcompartment encapsulation peptides

Bacterial microcompartments are protein organelles with diverse metabolic capabilities. Their functional diversity is determined by an enzymatic core that is sequestered within a structurally conserved protein shell architecture. Segregation of protein cargo into the bacterial microcompartment is enabled by encapsulation peptides, which are short helical domains fused to core proteins through a disordered linker. Here, we investigate how encapsulation peptides drive multicomponent cargo assembly into biomolecular condensates. In vitro experiments supported by molecular dynamics simulations demonstrate the importance of both conserved hydrophobic packing and electrostatic interactions in stabilizing trimeric encapsulation peptide bundles. Topological rearrangements of encapsulation peptide domains can drive programmable liquid- or gel-like partitioning in vitro and in vivo. This partitioning is found to be encapsulation peptide-specific, modular, and can co-assemble at least three fluorescent reporters. In summary, we describe the molecular features necessary to drive biomolecular condensation using a widespread peptide tag. This work can serve as a blueprint for implementing encapsulation peptide biotechnology across diverse applications.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Colistin resistance plasmids dually enhance bacterial virulence and antibiotic resistance via surface polysaccharide biosynthesis

Plasmids carrying the mobilized colistin-resistance gene mcr-1 are prevalent among multidrug-resistant Gram-negative pathogens, yet their broad impact on bacterial physiology and virulence remains unclear. Here, we demonstrate that acquisition of an mcr-1 plasmid concurrently increases antimicrobial resistance and pathogenicity in Escherichia coli. On the same plasmid, the XRE-family transcriptional regulator EcaR cooperates with MCR-1 to activate the wec operon, driving biosynthesis of two surface polysaccharides: enterobacterial common antigen (ECA) and a high-molecular-weight O-chain. Expression of these surface polysaccharides increases bile resistance and virulence in a murine model and further elevates colistin resistance. MCR-1 enhances transcription of upstream genes in the wec operon, whereas EcaR directly activates an internal promoter (PwecE) to induce downstream gene expression. Thus, both components are required for surface polysaccharide expression, and deletion of either abolishes the phenotype. Genomic analysis of publicly available mcr plasmids reveals widespread co-occurrence of mcr-1 and ecaR on IncI2 and IncX4 plasmids, indicating their functional complementarity. These findings uncover a mechanism by which resistance plasmids remodel the bacterial surface, linking horizontal gene transfer to coordinated regulation of antimicrobial resistance and virulence.

Antimicrobial resistance↗
DOE OSTI2021

Genomic and functional analyses of fungal and bacterial consortia that enable lignocellulose breakdown in goat gut microbiomes

Abstract The herbivore digestive tract is home to a complex community of anaerobic microbes that work together to break down lignocellulose. These microbiota are an untapped resource of strains, pathways and enzymes that could be applied to convert plant waste into sugar substrates for green biotechnology. We carried out more than 400 parallel enrichment experiments from goat faeces to determine how substrate and antibiotic selection influence membership, activity, stability and chemical productivity of herbivore gut communities. We assembled 719 high-quality metagenome-assembled genomes (MAGs) that are unique at the species level. More than 90% of these MAGs are from previously unidentified herbivore gut microorganisms. Microbial consortia dominated by anaerobic fungi outperformed bacterially dominated consortia in terms of both methane production and extent of cellulose degradation, which indicates that fungi have an important role in methane release. Metabolic pathway reconstructions from MAGs of 737 bacteria, archaea and fungi suggest that cross-domain partnerships between fungi and methanogens enabled production of acetate, formate and methane, whereas bacterially dominated consortia mainly produced short-chain fatty acids, including propionate and butyrate. Analyses of carbohydrate-active enzyme domains present in each anaerobic consortium suggest that anaerobic bacteria and fungi employ mostly complementary hydrolytic strategies. The division of labour among herbivore anaerobes to degrade plant biomass could be harnessed for industrial bioprocessing.

59 BASIC BIOLOGICAL SCIENCES↗