Engineering Papers⌕ Search

SEARCH · Engineering Papers

Results for “PHAGE DATA”

Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 55 records · Page 3
All sourcesNASA NTRSDOE OSTI
DOE OSTI2025

Nanobody screening and machine learning guided identification of cross-variant anti-SARS-CoV-2 neutralizing heavy-chain only antibodies

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) continues to persist, demonstrating the risks posed by emerging infectious diseases to national security, public health, and the economy. Development of new vaccines and antibodies for emerging viral threats requires substantial resources and time, and traditional development platforms for vaccines and antibodies are often too slow to combat continuously evolving immunological escape variants, reducing their efficacy over time. Previously, we designed a next-generation synthetic humanized nanobody (Nb) phage display library and demonstrated that this library could be used to rapidly identify highly specific and potent neutralizing heavy chain-only antibodies (HCAbs) with prophylactic and therapeutic efficacy in vivo against the original SARS-CoV-2. In this study, we used a combination of high throughput screening and machine learning (ML) models to identify HCAbs with potent efficacy against SARS-CoV-2 viral variants of interest (VOIs) and concern (VOCs). To start, we screened our highly diverse Nb phage display library against several pre-Omicron VOI and VOC receptor binding domains (RBDs) to identify panels of cross-reactive HCAbs. Using HCAb affinity for SARS-CoV-2 VOI and VOCs (pre-Omicron variants) and model features from other published data, we were able to develop a ML model that successfully identified HCAbs with efficacy against Omicron variants, independent of our experimental biopanning workflow. This biopanning informed ML approach reduced the experimental screening burden by 78% to 90% for the Omicron BA.5 and Omicron BA.1 variants, respectively. The combined approach can be applied to other emerging viruses with pandemic potential to rapidly identify effective therapeutic antibodies against emerging variants.

Antibodies↗
DOE OSTI2024

VIBES: a workflow for annotating and visualizing viral sequences integrated into bacterial genomes

Abstract Bacteriophages are viruses that infect bacteria. Many bacteriophages integrate their genomes into the bacterial chromosome and become prophages. Prophages may substantially burden or benefit host bacteria fitness, acting in some cases as parasites and in others as mutualists. Some prophages have been demonstrated to increase host virulence. The increasing ease of bacterial genome sequencing provides an opportunity to deeply explore prophage prevalence and insertion sites. Here we present VIBES (Viral Integrations in Bacterial genomES), a workflow intended to automate prophage annotation in complete bacterial genome sequences. VIBES provides additional context to prophage annotations by annotating bacterial genes and viral proteins in user-provided bacterial and viral genomes. The VIBES pipeline is implemented as a Nextflow-driven workflow, providing a simple, unified interface for execution on local, cluster and cloud computing environments. For each step of the pipeline, a container including all necessary software dependencies is provided. VIBES produces results in simple tab-separated format and generates intuitive and interactive visualizations for data exploration. Despite VIBES’s primary emphasis on prophage annotation, its generic alignment-based design allows it to be deployed as a general-purpose sequence similarity search manager. We demonstrate the utility of the VIBES prophage annotation workflow by searching for 178 Pf phage genomes across 1072 Pseudomonas spp. genomes.

59 BASIC BIOLOGICAL SCIENCES↗
NASA NTRS1991

Microbial growth and physiology in space - A review

An overview of microbial behavior in closed environments is given with attention to data related to simulated microgravity and actual space flight. Microbes are described in terms of antibiotic sensitivity, subcellular structure, and physiology, and the combined effects are considered of weightlessness and cosmic radiation on human immunity to such microorganisms. Space flight results report such effects as increased phage induction, accelerated microbial growth rates, and the increased risk of disease communication and microbial exchange aboard confining spacecraft. Ultrastructural changes are also noted in the nuclei, cell membranes, and cytoplasmic streaming, and it appears that antibiotic sensitivity is reduced under both actual and simulated conditions of spaceflight.

Cioletti, Louis A.↗
DOE OSTI2018

Engineered synthetic antibodies as probes to quantify the energetic contributions of ligand binding to conformational changes in proteins

Conformational changes in proteins due to ligand binding are ubiquitous in biological processes and are integral to many biological systems. However, it is often challenging to link ligand-induced conformational changes to a resulting biological function because it is difficult to distinguish between the energetic components associated with ligand binding and those due to structural rearrangements. Consequently, here, we used a unique approach exploiting conformation-specific and regio-specific synthetic antibodies (sABs) to probe the energetic contributions of ligand binding to conformation changes. Using maltose-binding protein (MBP) as a model system, customized phage-display selections were performed to generate sABs that stabilize MBP in different conformational states, modulating ligand-binding affinity in competitive, allosteric, or peristeric manners. We determined that the binding of a closed conformation–specific sAB (sAB-11M) to MBP in the absence of maltose is entropically driven, providing new insight into designing antibody-stabilized protein interactions. Crystal structures of sABs bound to MBP, together with biophysical data, delineate the basis of free energy differences between different conformational states and confirm the use of the sABs as energy probes for dissecting enthalpic and entropic contributions to conformational transitions. Our work provides a foundation for investigating the energetic contributions of distinct conformational dynamics to specific biological outputs. We anticipate that our approach also may be valuable for analyzing the energy landscapes of regulatory proteins controlling biological responses to environmental changes.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Structure analysis of the telomere resolvase from the Lyme disease spirochete Borrelia garinii reveals functional divergence of its C-terminal domain

Borrelia spirochetes are the causative agents of Lyme disease and relapsing fever, two of the most common tick-borne illnesses. A characteristic feature of these spirochetes is their highly segmented genomes which consists of a linear chromosome and a mixture of up to approximately 24 linear and circular extrachromosomal plasmids. The complexity of this genomic arrangement requires multiple strategies for efficient replication and partitioning during cell division, including the generation of hairpin ends found on linear replicons mediated by the essential enzyme ResT, a telomere resolvase. Using an integrative structural biology approach employing advanced modelling, circular dichroism, X-ray crystallography and small-angle X-ray scattering, we have generated high resolution structural data on ResT from B. garinii. Our data provides the first high-resolution structures of ResT from Borrelia spirochetes and revealed active site positioning in the catalytic domain. We also demonstrate that the C-terminal domain of ResT is required for both transesterification steps of telomere resolution, and is a requirement for DNA binding, distinguishing ResT from other telomere resolvases from phage and bacteria. These results advance our understanding of the molecular function of this essential enzyme involved in genome maintenance in Borrelia pathogens.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Metagenomic compendium of 189,680 DNA viruses from the human gut microbiome

Bacteriophages have important roles in the ecology of the human gut microbiome but are under-represented in reference databases. To address this problem, we assembled the Metagenomic Gut Virus catalogue that comprises 189,680 viral genomes from 11,810 publicly available human stool metagenomes. Over 75% of genomes represent double-stranded DNA phages that infect members of the Bacteroidia and Clostridia classes. Based on sequence clustering we identified 54,118 candidate viral species, 92% of which were not found in existing databases. The Metagenomic Gut Virus catalogue improves detection of viruses in stool metagenomes and accounts for nearly 40% of CRISPR spacers found in human gut Bacteria and Archaea. We also produced a catalogue of 459,375 viral protein clusters to explore the functional potential of the gut virome. This revealed tens of thousands of diversity-generating retroelements, which use error-prone reverse transcription to mutate target genes and may be involved in the molecular arms race between phages and their bacterial hosts.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Predicting variable gene content in Escherichia coli using conserved genes

Having the ability to predict the protein-encoding gene content of an incomplete genome or metagenome-assembled genome is important for a variety of bioinformatic tasks. In this study, as a proof of concept, we built machine learning classifiers for predicting variable gene content in Escherichia coli genomes using only the nucleotide k-mers from a set of 100 conserved genes as features. Protein families were used to define orthologs, and a single classifier was built for predicting the presence or absence of each protein family occurring in 10%–90% of all E. coli genomes. The resulting set of 3,259 extreme gradient boosting classifiers had a per-genome average macro F1 score of 0.944 [0.943–0.945, 95% CI]. We show that the F1 scores are stable across multi-locus sequence types and that the trend can be recapitulated by sampling a smaller number of core genes or diverse input genomes. Surprisingly, the presence or absence of poorly annotated proteins, including “hypothetical proteins” was accurately predicted (F1 = 0.902 [0.898–0.906, 95% CI]). Models for proteins with horizontal gene transfer-related functions had slightly lower F1 scores but were still accurate (F1s = 0.895, 0.872, 0.824, and 0.841 for transposon, phage, plasmid, and antimicrobial resistance-related functions, respectively). Finally, using a holdout set of 419 diverse E. coli genomes that were isolated from freshwater environmental sources, we observed an average per-genome F1 score of 0.880 [0.876–0.883, 95% CI], demonstrating the extensibility of the models. Overall, this study provides a framework for predicting variable gene content using a limited amount of input sequence data.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Synthetic antibodies against BRIL as universal fiducial marks for single-particle cryoEM structure determination of membrane proteins

We propose the concept of universal fiducials based on a set of pre-made semi-synthetic antibodies (sABs) generated by customized phage display selections against the fusion protein BRIL, an engineered variant of apocytochrome b562a. These sABs can bind to BRIL fused either into the loops or termini of different GPCRs, ion channels, receptors and transporters without disrupting their structure. A crystal structure of BRIL in complex with an affinity-matured sAB (BAG2) that bound to all systems tested delineates the footprint of interaction. Negative stain and cryoEM data of several examples of BRIL-membrane protein chimera highlight the effectiveness of the sABs as universal fiducial marks. Taken together with a cryoEM structure of sAB bound human nicotinic acetylcholine receptor, this work demonstrates that these anti-BRIL sABs can greatly enhance the particle properties leading to improved cryoEM outcomes, especially for challenging membrane proteins.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Genomic and phenotypic comparison of two variants of multidrug-resistant Salmonella enterica serovar Heidelberg isolated during the 2015–2017 multi-state outbreak in cattle

Salmonella enterica subspecies enterica serovar Heidelberg (Salmonella Heidelberg) has caused several multistate foodborne outbreaks in the United States, largely associated with the consumption of poultry. However, a 2015–2017 multidrug-resistant (MDR) Salmonella Heidelberg outbreak was linked to contact with dairy beef calves. Traceback investigations revealed calves infected with outbreak strains of Salmonella Heidelberg exhibited symptoms of disease frequently followed by death from septicemia. To investigate virulence characteristics of Salmonella Heidelberg as a pathogen in bovine, two variants with distinct pulse-field gel electrophoresis (PFGE) patterns that differed in morbidity and mortality during the multistate outbreak were genotypically and phenotypically characterized and compared. Strain SX 245 with PFGE pattern JF6X01.0523 was identified as a dominant and highly pathogenic variant causing high morbidity and mortality in affected calves, whereas strain SX 244 with PFGE pattern JF6X01.0590 was classified as a low pathogenic variant causing less morbidity and mortality. Comparison of whole-genome sequences determined that SX 245 lacked ~200 genes present in SX 244, including genes associated with the IncI1 plasmid and phages; SX 244 lacked eight genes present in SX 245 including a second YdiV Anti-FlhC(2)FlhD(4) factor, a lysin motif domain containing protein, and a pentapeptide repeat protein. RNA-sequencing revealed fimbriae-related, flagella-related, and chemotaxis genes had increased expression in SX 245 compared to SX 244. Furthermore, SX 245 displayed higher invasion of human and bovine epithelial cells than SX 244. These data suggest that the presence and up-regulation of genes involved in type 1 fimbriae production, flagellar regulation and biogenesis, and chemotaxis may play a role in the increased pathogenicity and host range expansion of the Salmonella Heidelberg isolates involved in the bovine-related outbreak.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Mechanisms of SARS-CoV-2 neutralization by shark variable new antigen receptors elucidated through X-ray crystallography

Single-domain Variable New Antigen Receptors (VNARs) from the immune system of sharks are the smallest naturally occurring binding domains found in nature. Possessing flexible paratopes that can recognize protein motifs inaccessible to classical antibodies, VNARs have yet to be exploited for the development of SARS-CoV-2 therapeutics. Here, we detail the identification of a series of VNARs from a VNAR phage display library screened against the SARS-CoV-2 receptor binding domain (RBD). The ability of the VNARs to neutralize pseudotype and authentic live SARS-CoV-2 virus rivalled or exceeded that of full-length immunoglobulins and other single-domain antibodies. Crystallographic analysis of two VNARs found that they recognized separate epitopes on the RBD and had distinctly different mechanisms of virus neutralization unique to VNARs. Structural and biochemical data suggest that VNARs would be effective therapeutic agents against emerging SARS-CoV-2 mutants, including the Delta variant, and coronaviruses across multiple phylogenetic lineages. This study highlights the utility of VNARs as effective therapeutics against coronaviruses and may serve as a critical milestone for nearing a paradigm shift of the greater biologic landscape.

60 APPLIED LIFE SCIENCES↗
NASA NTRS1990

Preliminary crystallographic examination of a novel fungal lysozyme from Chalaropsis

The lysozyme from the fungus of the Chalaropsis species has been crystallized. This lysozyme displays no sequence homology with avian, phage, or mammalian lysozymes, however, preliminary studies indicate significant sequence homology with the bacterial lysozyme from Streptomyces. Both enzymes are unusual in possessing beta-1,4-N-acetylmuramidase and beta-1,4-N,6-O-diacetylmuramidase activity. The crystals grow from solutions of ammonium sulfate during growth periods from several months to a year. The space group is P2(1)2(1)2(1) with a = 34.0 A, b = 42.6 A, c = 122.1 A. Preliminary data indicate that there is 1 molecule/asymmetric unit.

Carter, Daniel C.↗
DOE OSTI2017

Rhesus macaque and mouse models for down-selecting circumsporozoite protein based malaria vaccines differ significantly in immunogenicity and functional outcomes

Background: Non-human primates, such as the rhesus macaques, are the preferred model for down-selecting human malaria vaccine formulations, but the rhesus model is expensive and does not allow for direct efficacy testing of human malaria vaccines. Transgenic rodent parasites expressing genes of human Plasmodium are now routinely used for efficacy studies of human malaria vaccines. Mice have however rarely predicted success in human malaria trials and there is scepticism whether mouse studies alone are sufficient to move a vaccine candidate into the clinic. Methods: A comparison of immunogenicity, fine-specificity and functional activity of two Alum-adjuvanted Plasmodium falciparum circumsporozoite protein (CSP)-based vaccines was conducted in mouse and rhesus models. One vaccine was a soluble recombinant protein (CSP) and the other was the same CSP covalently conjugated to the Qβ phage particle (Qβ-CSP). Results: Mice showed different kinetics of antibody responses and different sensitivity to the NANP-repeat and N-terminal epitopes as compared to rhesus. While mice failed to discern differences between the protective efficacy of CSP versus Qβ-CSP vaccine following direct challenge with transgenic Plasmodium berghei parasites, rhesus serum from the Qβ-CSP-vaccinated animals induced higher in vivo sporozoite neutralization activity. Conclusions: Despite some immunologic parallels between models, these data demonstrate that differences between the immune responses induced in the two models risk conflicting decisions regarding potential vaccine utility in humans. In combination with historical observations, the data presented here suggest that although murine models may be useful for some purposes, non-human primate models may be more likely to predict the human response to investigational vaccines.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2020

Small and mighty: adaptation of superphylum Patescibacteria to groundwater environment drives their genome simplicity

The newly defined superphylum Patescibacteria such as Parcubacteria (OD1) and Microgenomates (OP11) has been found to be prevalent in groundwater, sediment, lake, and other aquifer environments. Recently increasing attention has been paid to this diverse superphylum including > 20 candidate phyla (a large part of the candidate phylum radiation, CPR) because it refreshed our view of the tree of life. However, adaptive traits contributing to its prevalence are still not well known. Results Here, we investigated the genomic features and metabolic pathways of Patescibacteria in groundwater through genome-resolved metagenomics analysis of >600 Gbp sequence data. We observed that, while the members of Patescibacteria have reduced genomes (~ 1 Mbp) exclusively, functions essential to growth and reproduction such as genetic information processing were retained. Surprisingly, they have sharply reduced redundant and nonessential functions, including specific metabolic activities and stress response systems. The Patescibacteria have ultra-small cells and simplified membrane structures, including flagellar assembly, transporters, and two-component systems. Despite the lack of CRISPR viral defense, the bacteria may evade predation through deletion of common membrane phage receptors and other alternative strategies, which may explain the low representation of prophage proteins in their genomes and lack of CRISPR. By establishing the linkages between bacterial features and the groundwater environmental conditions, our results provide important insights into the functions and evolution of this CPR group.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2019

PHANOTATE: a novel approach to gene identification in phage genomes

Motivation: Currently there are no tools specifically designed for annotating genes in phages. Several tools are available that have been adapted to run on phage genomes, but due to their underlying design, they are unable to capture the full complexity of phage genomes. Phages have adapted their genomes to be extremely compact, having adjacent genes that overlap and genes completely inside of other longer genes. This non-delineated genome structure makes it difficult for gene prediction using the currently available gene annotators. Here we present PHANOTATE, a novel method for gene calling specifically designed for phage genomes. Although the compact nature of genes in phages is a problem for current gene annotators, we exploit this property by treating a phage genome as a network of paths: where open reading frames are favorable, and overlaps and gaps are less favorable, but still possible. We represent this network of connections as a weighted graph, and use dynamic programing to find the optimal path. Results: We compare PHANOTATE to other gene callers by annotating a set of 2133 complete phage genomes from GenBank, using PHANOTATE and the three most popular gene callers. We found that the four programs agree on 82% of the total predicted genes, with PHANOTATE predicting more genes than the other three. We searched for these extra genes in both GenBank’s non-redundant protein database and all of the metagenomes in the sequence read archive, and found that they are present at levels that suggest that these are functional protein-coding genes. Availability and implementation: https://github.com/deprekate/PHANOTATE. Supplementary information: Supplementary data are available at Bioinformatics online.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Underlying data for characterization of phycocyanobilin (PCB) biosynthesis in Galdieria sulphuraria

This dataset contains data for studies of phycocyanobilin (PCB) synthesis in the red alga Galdieria sulphuraria. Red algae such as G. sulphuraria utilize phycobilisomes for light harvesting. The phycobilisomes of early-branching organisms such as G. sulphuraria or Cyanidioschyzon merolae contain PCB chromophores but not phycoerythrobilin (PEB), in contrast to the phycobilisomes of other red algae. The studies reported in this dataset examine biosynthesis of PCB in G. sulphuraria and C. merolae, starting from biliverdin IX-alpha (BV), the last known common precursor for PCB and PEB. In cyanobacteria, phages, green algae, and land plants, conversion of BV into PCB or PEB is carried out by a family of enzymes called ferredoxin-dependent bilin reductases (FDBRs). The current studies demonstrate that G. sulphuraria, but not C. merolae, require the action of an additional isomerase to synthesize PCB.

59 BASIC BIOLOGICAL SCIENCES↗
NASA NTRS1994

Continuous in vitro evolution of bacteriophage RNA polymerase promoters

Rapid in vitro evolution of bacteriophage T7, T3, and SP6 RNA polymerase promoters was achieved by a method that allows continuous enrichment of DNAs that contain functional promoter elements. This method exploits the ability of a special class of nucleic acid molecules to replicate continuously in the presence of both a reverse transcriptase and a DNA-dependent RNA polymerase. Replication involves the synthesis of both RNA and cDNA intermediates. The cDNA strand contains an embedded promoter sequence, which becomes converted to a functional double-stranded promoter element, leading to the production of RNA transcripts. Synthetic cDNAs, including those that contain randomized promoter sequences, can be used to initiate the amplification cycle. However, only those cDNAs that contain functional promoter sequences are able to produce RNA transcripts. Furthermore, each RNA transcript encodes the RNA polymerase promoter sequence that was responsible for initiation of its own transcription. Thus, the population of amplifying molecules quickly becomes enriched for those templates that encode functional promoters. Optimal promoter sequences for phage T7, T3, and SP6 RNA polymerase were identified after a 2-h amplification reaction, initiated in each case with a pool of synthetic cDNAs encoding greater than 10(10) promoter sequence variants.

Non-NASA Center↗
DOE OSTI2015

Metagenomic reconstructions of bacterial CRISPR loci constrain population histories

Bacterial CRISPR-Cas systems offer insight into recent population history because they rapidly incorporate, in a unidirectional manner, short fragments (spacers) from coexisting infective virus populations into host chromosomes. Immunity is achieved by sequence identity between transcripts of spacers and their targets. Here, we used metagenomics to study the stability and dynamics of the type I-E CRISPR-Cas locus of Leptospirillum group II bacteria in biofilms sampled over 5 years from an acid mine drainage (AMD) system. Despite recovery of 452,686 spacers from CRISPR amplicons and metagenomic data, rarefaction curves of spacers show no saturation. The vast repertoire of spacers is attributed to phage/plasmid population diversity and retention of old spacers, despite rapid evolution of the targeted phage/plasmid genome regions (proto-spacers). The oldest spacers (spacers found at the trailer end) are conserved for at least 5 years, and 12% of these retain perfect or near-perfect matches to proto-spacer targets. The majority of proto-spacer regions contain an AAG proto-spacer adjacent motif (PAM). Spacers throughout the locus target the same phage population (AMDV1), but there are blocks of consecutive spacers without AMDV1 target sequences. Results suggest long-term coexistence of Leptospirillum with AMDV1 and periods when AMDV1 was less dominant. Metagenomics can be applied to millions of cells in a single sample to provide an extremely large spacer inventory, allow identification of phage/plasmids and enable analysis of previous phage/plasmid exposure. As a result, this method can provide insights into prior bacterial environment and genetic interplay between hosts and their viruses.

59 BASIC BIOLOGICAL SCIENCES↗
NASA NTRS1994

Mutation induction in bacteria after heavy ion irradiation

From a compilation of experimental data on the mutagenic effects of heavy ions in bacteria, main conclusions have been drawn as follows: (1) The mutagenic efficacy of heavy ions in bacteria depends on physical and biological variables. Physical variables are the radiation dose, energy and charge of the ion; the biological variables are the bacterial strain, the repair genotype of bacteria, and the endpoint investigated (type of mutation, induction of enzymes related to mutagenesis); (2) The responses on dose or fluence are mainly linear or linear quadratic. The quadratic component, if found for low LET radiation, is gradually reduced with increasing LET; (3) At low values of Z and LET the cross section of mutation induction sigma m (as well as SOS response, sigma sos. and lambda phage induction, sigma lambda versus LET curves can be quite consistently described by a common function which increases up to approximately 100 keV/mu m. For higher LET values, the sigma(m) versus LET curves show the so-called 'hooks' observed also for other endpoints; (4) For light ions (Z is less than or equal to 4), the cross sections mostly decrease with increasing ion energy, which is probably related to the decrease of the specific energy departed by the ion inside the sensitive volume (cell). For ions in the range of Z = 10, sigma(m) is nearly independent on the ion energy. For heavier ions (Z is greater than or equal to 16), sigma(m) increases with the energy up to a maximum or saturation around 10 MeV/u. The increment becomes steeper with increasing atomic number of the ion. It correlates with the increasing track radius of the heavy ion; (5) The mutagenic efficiency per lethal event changes slightly with ion energy, if Z is small indicating a rough correlation between cellular lethality and mutation induction, only. For ions of higher Z this relation increases with energy, indicating a change in the 'mode' of radiation action from 'killing-prone' to 'mutation-prone'; and (6) Repair genotype substantially influences the radiation induced mutagenesis. Different mechanisms of mutation induction and/or different types of biologically significant lesions in wild type cells compared to repair deficient strains are a likely explanation.

Horneck, G.↗