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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 667 records · Page 37
All sourcesNASA NTRSDOE OSTI
DOE OSTI2023

Atomistic structure of the SARS-CoV-2 pseudoknot in solution from SAXS-driven molecular dynamics

SARS-CoV-2 depends on -1 programmed ribosomal frameshifting (-1 PRF) to express proteins essential for its replication. The RNA pseudoknot stimulating -1 PRF is thus an attractive drug target. However, the structural models of this pseudoknot obtained from cryo-EM and crystallography differ in some important features, leaving the pseudoknot structure unclear. We measured the solution structure of the pseudoknot using small-angle X-ray scattering (SAXS). The measured profile did not agree with profiles computed from the previously solved structures. Beginning with each of these solved structures, we used the SAXS data to direct all atom molecular dynamics (MD) simulations to improve the agreement in profiles. In all cases, this refinement resulted in a bent conformation that more closely resembled the cryo-EM structures than the crystal structure. Applying the same approach to a point mutant abolishing -1 PRF revealed a notably more bent structure with reoriented helices. This work clarifies the dynamic structures of the SARS-CoV-2 pseudoknot in solution.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2024

A phage nucleus-associated RNA-binding protein is required for jumbo phage infection

Abstract Large-genome bacteriophages (jumbo phages) of the proposed family Chimalliviridae assemble a nucleus-like compartment bounded by a protein shell that protects the replicating phage genome from host-encoded restriction enzymes and DNA-targeting CRISPR-Cas nucleases. While the nuclear shell provides broad protection against host nucleases, it necessitates transport of mRNA out of the nucleus-like compartment for translation by host ribosomes, and transport of specific proteins into the nucleus-like compartment to support DNA replication and mRNA transcription. Here, we identify a conserved phage nuclear shell-associated protein that we term Chimallin C (ChmC), which adopts a nucleic acid-binding fold, binds RNA with high affinity in vitro, and binds phage mRNAs in infected cells. ChmC also forms phase-separated condensates with RNA in vitro. Targeted knockdown of ChmC using mRNA-targeting dCas13d results in accumulation of phage-encoded mRNAs in the phage nucleus, reduces phage protein production, and compromises virion assembly. Taken together, our data show that the conserved ChmC protein plays crucial roles in the viral life cycle, potentially by facilitating phage mRNA translocation through the nuclear shell to promote protein production and virion development.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

BGC Atlas: a web resource for exploring the global chemical diversity encoded in bacterial genomes

Secondary metabolites are compounds not essential for an organism’s development, but provide significant ecological and physiological benefits. These compounds have applications in medicine, biotechnology and agriculture. Their production is encoded in biosynthetic gene clusters (BGCs), groups of genes collectively directing their biosynthesis. The advent of metagenomics has allowed researchers to study BGCs directly from environmental samples, identifying numerous previously unknown BGCs encoding unprecedented chemistry. Here, we present the BGC Atlas (https://bgc-atlas.cs.uni-tuebingen.de), a web resource that facilitates the exploration and analysis of BGC diversity in metagenomes. The BGC Atlas identifies and clusters BGCs from publicly available datasets, offering a centralized database and a web interface for metadata-aware exploration of BGCs and gene cluster families (GCFs). We analyzed over 35 000 datasets from MGnify, identifying nearly 1.8 million BGCs, which were clustered into GCFs. The analysis showed that ribosomally synthesized and post-translationally modified peptides are the most abundant compound class, with most GCFs exhibiting high environmental specificity. We believe that our tool will enable researchers to easily explore and analyze the BGC diversity in environmental samples, significantly enhancing our understanding of bacterial secondary metabolites, and promote the identification of ecological and evolutionary factors shaping the biosynthetic potential of microbial communities.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2013

POGO-DB—a database of pairwise-comparisons of genomes and conserved orthologous genes

POGO-DB (http://pogo.ece.drexel.edu/) provides an easy platform for comparative microbial genomics. POGO-DB allows users to compare genomes using pre-computed metrics that were derived from extensive computationally intensive BLAST comparisons of >2000 microbes. These metrics include (i) average protein sequence identity across all orthologs shared by two genomes, (ii) genomic fluidity (a measure of gene content dissimilarity), (iii) number of ‘orthologs’ shared between two genomes, (iv) pairwise identity of the 16S ribosomal RNA genes and (v) pairwise identity of an additional 73 marker genes present in >90% prokaryotes. Users can visualize these metrics against each other in a 2D plot for exploratory analysis of genome similarity and of how different aspects of genome similarity relate to each other. The results of these comparisons are fully downloadable. In addition, users can download raw BLAST results for all or user-selected comparisons. Therefore, we provide users with full flexibility to carry out their own downstream analyses, by creating easy access to data that would normally require heavy computational resources to generate. POGO-DB should prove highly useful for researchers interested in comparative microbiology and benefit the microbiome/metagenomic communities by providing the information needed to select suitable phylogenetic marker genes within particular lineages.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2013

Rapid metabolic pathway assembly and modification using serine integrase site-specific recombination

Synthetic biology requires effective methods to assemble DNA parts into devices and to modify these devices once made. Here we demonstrate a convenient rapid procedure for DNA fragment assembly using site-specific recombination by ΦC31 integrase. Using six orthogonal attP/attB recombination site pairs with different overlap sequences, we can assemble up to five DNA fragments in a defined order and insert them into a plasmid vector in a single recombination reaction. ΦC31 integrase-mediated assembly is highly efficient, allowing production of large libraries suitable for combinatorial gene assembly strategies. The resultant assemblies contain arrays of DNA cassettes separated by recombination sites, which can be used to manipulate the assembly by further recombination. We illustrate the utility of these procedures to (i) assemble functional metabolic pathways containing three, four or five genes; (ii) optimize productivity of two model metabolic pathways by combinatorial assembly with randomization of gene order or ribosome binding site strength; and (iii) modify an assembled metabolic pathway by gene replacement or addition.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2014

The tmRNA website

We report that the transfer-messenger RNA (tmRNA) and its partner protein SmpB act together in resolving problems arising when translating bacterial ribosomes reach the end of mRNA with no stop codon. Their genes have been found in nearly all bacterial genomes and in some organelles. The tmRNA Website serves tmRNA sequences, alignments and feature annotations, and has recently moved to http: //bioinformatics.sandia.gov/tmrna/. New features include software used to find the sequences, an update raising the number of unique tmRNA sequences from 492 to 1716, and a database of SmpB sequences which are served along with the tmRNA sequence from the same organism.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2014

Transcript level and sequence determinants of protein abundance and noise in Escherichia coli

The range over which a protein is expressed, and its cell-to-cell variability, is often thought to be linked to the demand for its activity. Steady-state protein level is determined by multiple mechanisms controlling transcription and translation, many of which are limited by DNA- and RNA-encoded signals that affect initiation, elongation and termination of polymerases and ribosomes. We performed a comprehensive analysis of >100 sequence features to derive a predictive model composed of a minimal non-redundant set of factors explaining 66% of the total variation of protein abundance observed in >800 genes in Escherichia coli. The model suggests that protein abundance is primarily determined by the transcript level (53%) and by effectors of translation elongation (12%), whereas only a small fraction of the variation is explained by translational initiation (1%). Our analyses uncover a new sequence determinant, not previously described, affecting translation initiation and suggest that elongation rate is affected by both codon biases and specific amino acid composition. We also show that transcription and translation efficiency may have an effect on expression noise, which is more similar than previously assumed.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2015

Functional dichotomy in the 16S rRNA (m 1 A1408) methyltransferase family and control of catalytic activity via a novel tryptophan mediated loop reorganization

Methylation of the bacterial small ribosomal subunit (16S) rRNA on the N1 position of A1408 confers exceptionally high-level resistance to a broad spectrum of aminoglycoside antibiotics. Here, we present a detailed structural and functional analysis of the Catenulisporales acidiphilia 16S rRNA (m 1 A1408) methyltransferase (‘CacKam’). The apo CacKam structure closely resembles other m1A1408 methyltransferases within its conserved SAM-binding fold but the region linking core β strands 6 and 7 (the ‘β6/7 linker’) has a unique, extended structure that partially occludes the putative 16S rRNA binding surface, and sequesters the conserved and functionally critical W203 outside of the CacKam active site. Substitution of conserved residues in the SAM binding pocket reveals a functional dichotomy in the 16S rRNA (m 1 A1408) methyltransferase family, with two apparently distinct molecular mechanisms coupling cosubstrate/ substrate binding to catalytic activity. Our results additionally suggest that CacKam exploits the W203-mediated remodeling of the β6/7 linker as a novel mechanism to control 30S substrate recognition and enzymatic turnover.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Expansion of the tmRNA sequence database and new tools for search and visualization

Abstract Transfer–messenger RNA (tmRNA) contributes essential tRNA-like and mRNA-like functions during the process of trans-translation, a mechanism of quality control for the translating bacterial ribosome. Proper tmRNA identification benefits the study of trans-translation and also the study of genomic islands, which frequently use the tmRNA gene as an integration site. Automated tmRNA gene identification tools are available, but manual inspection is still important for eliminating false positives. We have increased our database of precisely mapped tmRNA sequences over 50-fold to 97 179 unique sequences. Group I introns had previously been found integrated within a single subsite within the TψC-loop; they have now been identified at four distinct subsites, suggesting multiple founding events of invasion of tmRNA genes by group I introns, all in the same vicinity. tmRNA genes were found in metagenomic archaeal genomes, perhaps a result of misbinning of bacterial sequences during genome assembly. With the expanded database, we have produced new covariance models for improved tmRNA sequence search and new secondary structure visualization tools.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

CABO-16S—a Combined Archaea, Bacteria, Organelle 16S rRNA database framework for amplicon analysis of prokaryotes and eukaryotes in environmental samples

Abstract Identification of both prokaryotic and eukaryotic microorganisms in environmental samples is currently challenged by the need for additional sequencing to obtain separate 16S and 18S ribosomal RNA (rRNA) amplicons or the constraints imposed by “universal” primers. Organellar 16S rRNA sequences are amplified and sequenced along with prokaryote 16S rRNA and provide an alternative method to identify eukaryotic microorganisms. CABO-16S combines bacterial and archaeal sequences from the SILVA database with 16S rRNA sequences of plastids and other organelles from the PR2 database to enable identification of all 16S rRNA sequences. Comparison of CABO-16S with SILVA 138.2 results in equivalent taxonomic classification of mock communities and increased classification of diverse environmental samples. In particular, identification of phototrophic eukaryotes in shallow seagrass environments, marine waters, and lake waters was increased. The CABO-16S framework allows users to add custom sequences for further classification of underrepresented clades and can be easily updated with future releases of reference databases. Addition of sequences obtained from Sanger sequencing of methane seep sediments and curated sequences of the polyphyletic SEEP-SRB1 clade resulted in differentiation of syntrophic and non-syntrophic SEEP-SRB1 in hydrothermal vent sediments. CABO-16S highlights the benefit of combining and amending existing training sets when studying microorganisms in diverse environments.

Eitel, Eryn M. (ORCID:0009000723919297)↗
DOE OSTI2018

DRMY1, a Myb-Like Protein, Regulates Cell Expansion and Seed Production in Arabidopsis thaliana

Abstract Plant organ development to a specific size and shape is controlled by cell proliferation and cell expansion. Here, we identify a novel Myb-like Arabidopsis gene, Development Related Myb-like1 (DRMY1), which controls cell expansion in both vegetative and reproductive organs. DRMY1 is strongly expressed in developing organs and its expression is reduced by ethylene while it is induced by ABA. DRMY1 has a Myb-like DNA-binding domain, which is predominantly localized in the nucleus and does not exhibit transcriptional activation activity. The loss-of-function T-DNA insertion mutant drmy1 shows reduced organ growth and cell expansion, which is associated with changes in the cell wall matrix polysaccharides. Interestingly, overexpression of DRMY1 in Arabidopsis does not lead to enhanced organ growth. Expression of genes involved in cell wall biosynthesis/remodeling, ribosome biogenesis and in ethylene and ABA signaling pathways is changed with the deficiency of DRMY1. Our results suggest that DRMY1 plays an essential role in organ development by regulating cell expansion either directly by affecting cell wall architecture and/or cytoplasmic growth or indirectly through the ethylene and/or ABA signaling pathways.

Cell Biology↗
DOE OSTI2026

Metabolic flux, metabolite, and transcript analysis uncover reprogramming of metabolism toward higher seed oil

Overexpression of WRINKLED1 (WRI1), a master regulator of glycolysis and fatty acid biosynthesis, together with DIACYLGLYCEROL ACYLTRANSFERASE1 (DGAT1), which catalyzes the final step of triacylglycerol assembly, is a promising strategy for enhancing seed oil content. However, how these regulators coordinate system-wide metabolic reprogramming at the levels of gene expression, metabolite pools, and fluxes remains poorly understood. To address this, we performed 13 C-metabolic flux analysis, metabolomics, and transcriptomics on in vitro cultured pennycress (Thlaspi arvense L.) embryos overexpressing the native WRI1 and DGAT1 homologs. Here, in cultured embryos, WRI1/DGAT1 overexpression increased triacylglycerol accumulation by 28% while reducing protein content by 34%, relative to the wild type. Embryos showed ∼20-fold and 50-fold upregulation of WRI1 and DGAT1 along with induction of WRI1 target genes in glycolysis and fatty acid biosynthesis. Genes associated with photosynthesis and Calvin cycle functions were also upregulated, whereas genes encoding ribosomal proteins and seed storage proteins were strongly repressed, consistent with the observed lipid–protein tradeoff. Flux analysis revealed that enhanced triacylglycerol biosynthesis is supported by increased flux through the Rubisco shunt and cytosolic pyruvate kinase, while the oxidative pentose phosphate pathway and malic enzyme contributed little to NADPH or pyruvate supply. Metabolomic profiling revealed extensive perturbations in glycolytic intermediates, tricarboxylic acid cycle metabolites, and amino acids. In plant grown seeds, WRI1/DGAT1 lines also showed a modest but significant increase in total lipid content. Collectively, these findings reveal how WRI1 and DGAT1 reprogram central metabolism to enhance oil accumulation, with relevance to mature seeds.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Impact of inoculation practices on microbiota assembly and community stability in a fabricated ecosystem

Studying plant-microbe-soil interactions is challenging due to their high complexity and variability in natural ecosystems. While fabricated ecosystems provide opportunities to recapitulate aspects of these systems in reduced complexity and controlled environments, inoculation can be a significant source of variation. To tackle this, we evaluated how different bacteria inoculation practices and plant harvesting time points affect the reproducibility of a microbial synthetic community (SynCom) in association with the model grass Brachypodium distachyon. We tested three microbial inoculation practices: seed inoculation, transplant inoculation, and seedling inoculation; and two harvesting points: early (14-day-old plants) and late (21 days post-inoculation). We grew our plants and bacterial strains in sterile devices (EcoFABs) and characterized the microbial community from root, rhizosphere, and sand using 16S ribosomal RNA gene sequencing. The results showed that inoculation practices significantly affected the rhizosphere microbial community only when harvesting at an early time point but not at the late stage. As the SynCom showed a persistent association with B. distachyon at 21 days post-inoculation regardless of inoculation practices, we assessed the reproducibility of each inoculation method and found that transplant inoculation showed the highest reproducibility. Moreover, plant biomass was not adversely affected by transplant inoculation treatment. We concluded that bacteria inoculation while transplanting coupled with a later harvesting time point gives the most reproducible microbial community in the EcoFAB-B. distachyon-SynCom fabricated ecosystem and recommend this method as a standardized protocol for use with fabricated ecosystem experimental systems.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Impact of Soil Salinity on the Cowpea Nodule-Microbiome and the Isolation of Halotolerant PGPR Strains to Promote Plant Growth under Salinity Stress

Cowpea is one of the major legumes cultivated in arid and semiarid regions of the world. Four soil-microbial samples (SS-1 through SS-4) collected from semiarid soils in Punjab, Pakistan were planted with cowpea (Vigna unguiculata) crops, which were grown under salinity stress to analyze bacterial composition in the rhizosphere and within nodules using cultivation-dependent and -independent methods. Two varieties, 603 and the salt-tolerant CB 46, were each inoculated with or without the four different native soil samples or grown in medium either N-deficient (-N) or supplemented with N (+N). Plants inoculated with soil samples SS-2 and SS-4 grew better than plants inoculated with SS-1- and SS-3 and grew comparably with the +N controls. Environmental DNA (eDNA) was isolated from SS-1 and SS-4, and, by 16S ribosomal RNA sequencing, the soil microbiomes consisted mainly of Actinobacteria, Firmicutes, Proteobacteria, and other nonproteobacterial genera. However, analysis of eDNA isolated from cowpea nodules established by the trap plants showed that the nodule microbiome consisted almost exclusively of proteobacterial sequences, particularly species of Bradyrhizobium. Bacteria were isolated from both soils and nodules, and 34 of the 51 isolates tested positive for plant-growth-promoting rhizobacteria traits in plate assays. Many could serve as future inocula for crops in arid soils. The discrepancy between the types of bacteria isolated by culturing bacteria isolated from surface-sterilized cowpea nodules (proteobacteria and nonproteobacteria) versus those detected by sequencing DNA isolated from the nodules (proteobacteria) from cowpea nodules (proteobacteria and nonproteobacteria) versus those detected in the nodule microbiome (proteobacteria) needs further study.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2013

Genomes of the class Erysipelotrichia clarify the firmicute origin of the class Mollicutes

The tree of life is paramount for achieving an integrated understanding of microbial evolution and the relationships between physiology, genealogy and genomics. It provides the framework for interpreting environmental sequence data, whether applied to microbial ecology or to human health. However, there remain many instances where there is ambiguity in our understanding of the phylogeny of major lineages, and/or confounding nomenclature. Here we apply recent genomic sequence data to examine the evolutionary history of members of the classes Mollicutes (phylum Tenericutes) and Erysipelotrichia (phylum Firmicutes). Consistent with previous analyses, we find evidence of a specific relationship between them in molecular phylogenies and signatures of the 16S rRNA, 23S rRNA, ribosomal proteins and aminoacyl-tRNA synthetase proteins. Furthermore, by mapping functions over the phylogenetic tree we find that the erysipelotrichia lineages are involved in various stages of genomic reduction, having lost (often repeatedly) a variety of metabolic functions and the ability to form endospores. Although molecular phylogeny has driven numerous taxonomic revisions, we find it puzzling that the most recent taxonomic revision of the phyla Firmicutes and Tenericutes has further separated them into distinct phyla, rather than reflecting their common roots.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2017

Reclassification of a Polynucleobacter cosmopolitanus strain isolated from tropical Lake Victoria as Polynucleobacter victoriensis sp. nov.

The genus Polynucleobacter (family Burkholderiaceae ) is phylogenetically subdivided into at least four subclusters. One of those, subcluster PnecC, was recognized as a cryptic species complex. Here we test by comparative genome analyses whether subcluster PnecD, currently solely represented by the species Polynucleobacter cosmopolitanus , also represents such a cryptic species complex. The genome sequences of the two P. cosmopolitanus strains, MWH-MoIso2T and MWH-VicM1, were determined. The latter strain was also characterized in the previous description of P. cosmopolitanus . These two strains originate from a temperate lake located in Austria and from the large tropical Lake Victoria located in East Africa, respectively. Strains MWH-MoIso2T and MWH-VicM1 possess quite small genomes of 1.78 and 1.63 Mbp, respectively, and share similar G+C values of 44.1 and 43.1 mol%, respectively. Both strains encode only a single copy of the ribosomal operon, and their 16S rRNA genes differ only in four positions, equalling a sequence similarity of 99.74 %. Both genomes possess characteristics indicating evolutionary genome streamlining, such as high coding densities of 93.9 and 94.6 % of bases, respectively. Average nucleotide identity (ANI) comparisons of the genomes of the two strains resulted in a value of 78.4 %, suggesting that each of the strains represents a separate species. Our investigation suggests that PnecD represents an additional cryptic species complex within the genus Polynucleobacter that was not resolved by 16S rRNA gene sequence analyses. We propose reclassification of strain MWH-VicM1 as Polynucleobacter victoriensis sp. nov., with type strain MWH-VicM1T(=DSM 21486T=JCM 32005T).

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Siderophores and secondary metabolites produced by Ganoderma adspersum

Ganoderma adspersum is a white-rot wood-degrading basidiomycete of ecological, biotechnological and medicinal interest. In addition to its role in lignin degradation, it produces bioactive metabolites with reported antimicrobial and antioxidant activities. However, the mechanisms of iron acquisition, including siderophore-mediated pathways, remain poorly characterized in Ganoderma species. Improved understanding of these systems is essential to elucidate their contributions to fungal physiology, secondary metabolism and ecological adaptation. In this study, the genome of G. adspersum was sequenced for the first time and screened for genes that may be involved in the production of secondary metabolites. A gene cluster was identified as potentially involved in iron uptake. In particular, genes related to non-ribosomal peptide synthetases were detected next to a gene encoding a monooxygenase and indicated a potential hydroxamate-family siderophore. Liquid chromatography (LC)-ES-MS analysis of secondary metabolites secreted by G. adspersum into the growth medium under iron-limiting conditions revealed a group of previously undescribed siderophores. Genome and MS/MS analysis suggested that these structures might be related to the coprinoferrin family of siderophores. Aside from siderophores, the genome and LC-MS analysis revealed G. adspersum to be a prolific producer of a variety of triterpenoids and sesquiterpenoids, in agreement with previous findings. This is the first description of the genome sequence of G. adspersum and its siderophores.

Reyes, Carolina [Laboratory for Cellulose and Wood↗
DOE OSTI2012

Phylogenomic analysis of polyketide synthase-encoding genes in Trichoderma

Members of the economically important ascomycete genus Trichoderma are ubiquitously distributed around the world. The mycoparasitic lifestyle and plant defense inducing interactions of Trichoderma spp. make them ideal biocontrol agents. Of the Trichoderma enzymes that produce secondary metabolites, some of which likely play important roles in biocontrol processes, polyketide synthases have garnered less attention that non-ribosomal peptide synthetases such as those that produce peptaibols. We have taken a phylogenomic approach to study the polyketide synthase repertoire encoded in the genomes of Trichoderma reesei, Trichoderma atroviride and Trichoderma virens. Our analysis lays a foundation for future research related to polyketide synthases within the Trichoderma genus and other filamentous fungi.

Trichoderma, polyketide, genomic, phylogenomic, fu↗