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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 127 records · Page 7
All sourcesNASA NTRSDOE OSTI
DOE OSTI2025

Integrated lipidomic and proteomic profiling reveals metabolic network disruption by SARS-CoV-2 variants

The rapid evolution of SARS-CoV-2 has produced myriad viral strains with increasing transmissibility and capacity for immune evasion. While effective vaccination campaigns have reduced the fatalities associated with SARS-CoV-2, infections continue, and a detailed understanding of how this virus manipulates host biochemical pathways remains elusive. We asked both whether the patterns of host lipid rewiring remained consistent across variants and whether the changes in the abundance of lipid classes are related to changes in the expression of the enzymes involved in their biosynthesis. We compared global nontargeted lipidomics on A549-ACE2 cells infected with the delta variant (B.1.617.2), or the omicron (B.1.1.529) variant to our previous results of global nontargeted lipidomics on A549-ACE2 cells infected with the original WA1 strain and further performed quantitative proteomics to assess changes in the host proteome. We found that metabolic rewiring, both on the lipid and the enzymatic level, is remarkably consistent across all three variants. We further mapped changes in the expression of host metabolic enzymes, linking enzyme expression to alterations in the abundance of specific lipids during infection. This analysis identified key proteins related to virus-mediated changes in lipid abundance, including fatty acid synthase (FASN), lysosomal acid lipase (LIPA), and ORMDL, a regulator of sphingolipid biosynthesis. These integrated lipidomic and proteomic experiments shed light on the importance of the complex network of host metabolism networks that support SARS-CoV-2 infection and suggest that lipid metabolism may be a promising avenue for uncovering conserved therapeutic targets.

SARS-CoV-2↗
DOE OSTI2023

Pan-Coronavirus CRISPR-CasRx Effector System Significantly Reduces Viable Titer in HCoV-OC43, HCoV-229E, and SARS-CoV-2

CRISPR-based technology has become widely used as an antiviral strategy, including as a broad-spectrum human coronavirus (HCoV) therapeutic. In this work, we have designed a CRISPR-CasRx effector system with guide RNAs (gRNAs) that are cross-reactive among several HCoV species. We tested the efficacy of this pan-coronavirus effector system by evaluating the reduction in viral viability associated with different CRISPR targets in HCoV-OC43, HCoV-229E, and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We determined that several CRISPR targets significantly reduce viral titer, despite the presence of single nucleotide polymorphisms in the gRNA when compared with a non-targeting, negative control gRNA. CRISPR targets reduced viral titer between 85% and >99% in HCoV-OC43, between 78% and >99% in HCoV-229E, and between 70% and 94% in SARS-CoV-2 when compared with an untreated virus control. These data establish a proof-of-concept for a pan-coronavirus CRISPR effector system that is capable of reducing viable virus in both Risk Group 2 and Risk Group 3 HCoV pathogens.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2020

Room-temperature neutron and X-ray data collection of 3CL M pro from SARS-CoV-2

The replication of SARS-CoV-2 produces two large polyproteins, pp1a and pp1ab, that are inactive until cleavage by the viral chymotrypsin-like cysteine protease enzyme (3CL M pro ) into a series of smaller functional proteins. At the heart of 3CL M pro is an unusual catalytic dyad formed by the side chains of His41 and Cys145 and a coordinated water molecule. The catalytic mechanism by which the enzyme operates is still unknown, as crucial information on the protonation states within the active site is unclear. To experimentally determine the protonation states of the catalytic site and of the other residues in the substrate-binding cavity, and to visualize the hydrogen-bonding networks throughout the enzyme, room-temperature neutron and X-ray data were collected from a large H/D-exchanged crystal of ligand-free (apo) 3CL M pro .

3CL Mpro↗
DOE OSTI2020

Structural basis of RNA cap modification by SARS-CoV-2

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), the causative agent of COVID-19 illness, has caused millions of infections worldwide. In SARS coronaviruses, the non-structural protein 16 (nsp16), in conjunction with nsp10, methylates the 5'-end of virally encoded mRNAs to mimic cellular mRNAs, thus protecting the virus from host innate immune restriction. We report here the high-resolution structure of a ternary complex of SARS-CoV-2 nsp16 and nsp10 in the presence of cognate RNA substrate analogue and methyl donor, S-adenosyl methionine (SAM). The nsp16/nsp10 heterodimer is captured in the act of 2'-O methylation of the ribose sugar of the first nucleotide of SARS-CoV-2 mRNA. We observe large conformational changes associated with substrate binding as the enzyme transitions from a binary to a ternary state. This induced fit model provides mechanistic insights into the 2'-O methylation of the viral mRNA cap. We also discover a distant (25 Å) ligand-binding site unique to SARS-CoV-2, which can alternatively be targeted, in addition to RNA cap and SAM pockets, for antiviral development.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

A metal ion orients SARS-CoV-2 mRNA to ensure accurate 2'- O methylation of its first nucleotide

The SARS-CoV-2 nsp16/nsp10 enzyme complex modifies the 2'-OH of the first transcribed nucleotide of the viral mRNA by covalently attaching a methyl group to it. The 2'-O methylation of the first nucleotide converts the status of mRNA cap from Cap-0 to Cap-1, and thus, helps the virus evade immune surveillance in host cells. Here, we report two structures of nsp16/nsp10 representing pre- and post-release states of the RNA product (Cap-1). We observe overall widening of the enzyme upon product formation, and an inward twisting motion in the substrate binding region upon product release. These conformational changes reset the enzyme for the next round of catalysis. The structures also identify a unique binding mode and the importance of a divalent metal ion for 2'-O methylation. We also describe underlying structural basis for the perturbed enzymatic activity of a clinical variant of SARS-CoV-2, and a previous SARS-CoV outbreak strain.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2022

The Local Topological Free Energy of the SARS-CoV-2 Spike Protein

The novel coronavirus SARS-CoV-2 infects human cells using a mechanism that involves binding and structural rearrangement of its Spike protein. Understanding protein rearrangement and identifying specific amino acids where mutations affect protein rearrangement has attracted much attention for drug development. In this manuscript, we use a mathematical method to characterize the local topology/geometry of the SARS-CoV-2 Spike protein backbone. Our results show that local conformational changes in the FP, HR1, and CH domains are associated with global conformational changes in the RBD domain. The SARS-CoV-2 variants analyzed in this manuscript (alpha, beta, gamma, delta Mink, G614, N501) show differences in the local conformations of the FP, HR1, and CH domains as well. Finally, most mutations of concern are either in or in the vicinity of high local topological free energy conformations, suggesting that high local topological free energy conformations could be targets for mutations with significant impact of protein function. Namely, the residues 484, 570, 614, 796, and 969, which are present in variants of concern and are targeted as important in protein function, are predicted as such from our model.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2023

Intensity of sample processing methods impacts wastewater SARS-CoV-2 whole genome amplicon sequencing outcomes

Wastewater SARS-CoV-2 surveillance has been deployed since the beginning of the COVID-19 pandemic to monitor the dynamics in virus burden in local communities. Genomic surveillance of SARS-CoV-2 in wastewater, particularly efforts aimed at whole genome sequencing for variant tracking and identification, are still challenging due to low target concentration, complex microbial and chemical background, and lack of robust nucleic acid recovery experimental procedures. The intrinsic sample limitations are inherent to wastewater and are thus unavoidable. Here, we use a statistical approach that couples correlation analyses to a random forest-based machine learning algorithm to evaluate potentially important factors associated with wastewater SARS-CoV-2 whole genome amplicon sequencing outcomes, with a specific focus on the breadth of genome coverage. We collected 182 composite and grab wastewater samples from the Chicago area between November 2020 to October 2021. Samples were processed using a mixture of processing methods reflecting different homogenization intensities (HA + Zymo beads, HA + glass beads, and Nanotrap), and were sequenced using one of the two library preparation kits (the Illumina COVIDseq kit and the QIAseq DIRECT kit). Technical factors evaluated using statistical and machine learning approaches include sample types, certain sample intrinsic features, and processing and sequencing methods. The results suggested that sample processing methods could be a predominant factor affecting sequencing outcomes, and library preparation kits was considered a minor factor. Finally, a synthetic SARS-CoV-2 RNA spike-in experiment was performed to validate the impact from processing methods and suggested that the intensity of the processing methods could lead to different RNA fragmentation

60 APPLIED LIFE SCIENCES↗
DOE OSTI2020

Crystal structures of SARS-CoV-2 ADP-ribose phosphatase: from the apo form to ligand complexes

Among 15 nonstructural proteins (Nsps), the newly emerging Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2) encodes a large, multidomain Nsp3. One of its units is the ADP-ribose phosphatase domain (ADRP; also known as the macrodomain, MacroD), which is believed to interfere with the host immune response. Such a function appears to be linked to the ability of the protein to remove ADP-ribose from ADP-ribosylated proteins and RNA, yet the precise role and molecular targets of the enzyme remain unknown. Here, five high-resolution (1.07–2.01 Å) crystal structures corresponding to the apo form of the protein and its complexes with 2-(N-morpholino)ethanesulfonic acid (MES), AMP and ADP-ribose have been determined. The protein is shown to undergo conformational changes to adapt to the ligand in the manner previously observed in close homologues from other viruses. A conserved water molecule is also identified that may participate in hydrolysis. This work builds foundations for future structure-based research on ADRP, including the search for potential antiviral therapeutics.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Effect of natural mutations of SARS-CoV-2 on spike structure, conformation, and antigenicity

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants with multiple spike mutations enable increased transmission and antibody resistance. We combined cryo–electron microscopy (cryo-EM), binding, and computational analyses to study variant spikes, including one that was involved in transmission between minks and humans, and others that originated and spread in human populations. All variants showed increased angiotensin-converting enzyme 2 (ACE2) receptor binding and increased propensity for receptor binding domain (RBD)–up states. While adaptation to mink resulted in spike destabilization, the B.1.1.7 (UK) spike balanced stabilizing and destabilizing mutations. A local destabilizing effect of the RBD E484K mutation was implicated in resistance of the B.1.1.28/P.1 (Brazil) and B.1.351 (South Africa) variants to neutralizing antibodies. Our studies revealed allosteric effects of mutations and mechanistic differences that drive either interspecies transmission or escape from antibody neutralization.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

CovTransformer: A transformer model for SARS-CoV-2 lineage frequency forecasting

With hundreds of SARS-CoV-2 lineages circulating in the global population, there is an ongoing need for predicting and forecasting lineage frequencies and thus identifying rapidly expanding lineages. Accurate prediction would allow for more focused experimental efforts to understand pathogenicity of future dominating lineages and characterize the extent of their immune escape. Here, we first show that the inherent noise and biases in lineage frequency data make a commonly-used regression-based approach unreliable. To address this weakness, we constructed a machine learning model for SARS-CoV-2 lineage frequency forecasting, called CovTransformer, based on the transformer architecture. We designed our model to navigate challenges such as a limited amount of data with high levels of noise and bias. We first trained and tested the model using data from the UK and the USA, and then tested the generalization ability of the model to many other countries and US states. Remarkably, the trained model makes accurate predictions two months into the future with high levels of accuracy both globally (in 31 countries with high levels of sequencing effort) and at the US-state level. Our model performed substantially better than a widely used forecasting tool, the multinomial regression model implemented in Nextstrain, demonstrating its utility in SARS-CoV-2 monitoring. Assuming a newly emerged lineage is identified and assigned, our test using retrospective data shows that our model is able to identify the dominating lineages 7 weeks in advance on average before they became dominant. Overall, our work demonstrates that transformer models represent a promising approach for SARS-CoV-2 forecasting and pandemic monitoring.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2022

Structural basis of SARS-CoV-2 Omicron immune evasion and receptor engagement

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant of concern evades antibody-mediated immunity that comes from vaccination or infection with earlier variants due to accumulation of numerous spike mutations. To understand the Omicron antigenic shift, we determined cryo–electron microscopy and x-ray crystal structures of the spike protein and the receptor-binding domain bound to the broadly neutralizing sarbecovirus monoclonal antibody (mAb) S309 (the parent mAb of sotrovimab) and to the human ACE2 receptor. We provide a blueprint for understanding the marked reduction of binding of other therapeutic mAbs that leads to dampened neutralizing activity. Remodeling of interactions between the Omicron receptor-binding domain and human ACE2 likely explains the enhanced affinity for the host receptor relative to the ancestral virus.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2023

Iterative computational design and crystallographic screening identifies potent inhibitors targeting the Nsp3 macrodomain of SARS-CoV-2

The nonstructural protein 3 (NSP3) of the severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) contains a conserved macrodomain enzyme (Mac1) that is critical for pathogenesis and lethality. While small-molecule inhibitors of Mac1 have great therapeutic potential, at the outset of the COVID-19 pandemic, there were no well-validated inhibitors for this protein nor, indeed, the macrodomain enzyme family, making this target a pharmacological orphan. Here, we report the structure-based discovery and development of several different chemical scaffolds exhibiting low- to sub-micromolar affinity for Mac1 through iterations of computer-aided design, structural characterization by ultra-high-resolution protein crystallography, and binding evaluation. Potent scaffolds were designed with in silico fragment linkage and by ultra-large library docking of over 450 million molecules. Both techniques leverage the computational exploration of tangible chemical space and are applicable to other pharmacological orphans. Overall, 160 ligands in 119 different scaffolds were discovered, and 153 Mac1-ligand complex crystal structures were determined, typically to 1 Å resolution or better. Our analyses discovered selective and cell-permeable molecules, unexpected ligand-mediated conformational changes within the active site, and key inhibitor motifs that will template future drug development against Mac1.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

2'-O methylation of RNA cap in SARS-CoV-2 captured by serial crystallography

The genome of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) coronavirus has a capping modification at the 5'-untranslated region (UTR) to prevent its degradation by host nucleases. These modifications are performed by the Nsp10/14 and Nsp10/16 heterodimers using S-adenosylmethionine as the methyl donor. Nsp10/16 heterodimer is responsible for the methylation at the ribose 2'-O position of the first nucleotide. To investigate the conformational changes of the complex during 2'-O methyltransferase activity, we used a fixed-target serial synchrotron crystallography method at room temperature. We determined crystal structures of Nsp10/16 with substrates and products that revealed the states before and after methylation, occurring within the crystals during the experiments. Here we report the crystal structure of Nsp10/16 in complex with Cap-1 analog ( m7 GpppA m2'-O ). Inhibition of Nsp16 activity may reduce viral proliferation, making this protein an attractive drug target.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Covalent narlaprevir- and boceprevir-derived hybrid inhibitors of SARS-CoV-2 main protease

Emerging SARS-CoV-2 variants continue to threaten the effectiveness of COVID-19 vaccines, and small-molecule antivirals can provide an important therapeutic treatment option. The viral main protease (Mpro) is critical for virus replication and thus is considered an attractive drug target. We performed the design and characterization of three covalent hybrid inhibitors BBH-1, BBH-2 and NBH-2 created by splicing components of hepatitis C protease inhibitors boceprevir and narlaprevir, and known SARS-CoV-1 protease inhibitors. A joint X-ray/neutron structure of the M pro /BBH-1 complex demonstrates that a Cys145 thiolate reaction with the inhibitor’s keto-warhead creates a negatively charged oxyanion. Protonation states of the ionizable residues in the M pro active site adapt to the inhibitor, which appears to be an intrinsic property of M pro . Structural comparisons of the hybrid inhibitors with PF-07321332 reveal unconventional F···O interactions of PF-07321332 with Mpro which may explain its more favorable enthalpy of binding. BBH-1, BBH-2 and NBH-2 exhibit comparable antiviral properties in vitro relative to PF-07321332, making them good candidates for further design of improved antivirals.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2022

Limited cross-variant immunity from SARS-CoV-2 Omicron without vaccination

SARS-CoV-2 Delta and Omicron are globally relevant variants of concern. Although individuals infected with Delta are at risk of developing severe lung disease, infection with Omicron often causes milder symptoms, especially in vaccinated individuals. The question arises of whether widespread Omicron infections could lead to future cross-variant protection, accelerating the end of the pandemic. Here we show that without vaccination, infection with Omicron induces a limited humoral immune response in mice and humans. Sera from mice overexpressing the human ACE2 receptor and infected with Omicron neutralize only Omicron, but not other variants of concern, whereas broader cross-variant neutralization was observed after WA1 and Delta infections. Unlike WA1 and Delta, Omicron replicates to low levels in the lungs and brains of infected animals, leading to mild disease with reduced expression of pro-inflammatory cytokines and diminished activation of lung-resident T cells. Sera from individuals who were unvaccinated and infected with Omicron show the same limited neutralization of only Omicron itself. By contrast, Omicron breakthrough infections induce overall higher neutralization titres against all variants of concern. Our results demonstrate that Omicron infection enhances pre-existing immunity elicited by vaccines but, on its own, may not confer broad protection against non-Omicron variants in unvaccinated individuals.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2020

Structural basis for potent neutralization of SARS-CoV-2 and role of antibody affinity maturation

SARS-CoV-2 is a betacoronavirus virus responsible for the COVID-19 pandemic. Here, we determine the X-ray crystal structure of a potent neutralizing monoclonal antibody, CV30, isolated from a patient infected with SARS-CoV-2, in complex with the receptor binding domain. The structure reveals that CV30 binds to an epitope that overlaps with the human ACE2 receptor binding motif providing a structural basis for its neutralization. CV30 also induces shedding of the S1 subunit, indicating an additional mechanism of neutralization. A germline reversion of CV30 results in a substantial reduction in both binding affinity and neutralization potential indicating the minimal somatic mutation is needed for potently neutralizing antibodies against SARS-CoV-2.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Cooperative multivalent receptor binding promotes exposure of the SARS-CoV-2 fusion machinery core

The molecular events that permit the spike glycoprotein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to bind and enter cells are important to understand for both fundamental and therapeutic reasons. Spike proteins consist of S1 and S2 domains, which recognize angiotensin-converting enzyme 2 (ACE2) receptors and contain the viral fusion machinery, respectively. Ostensibly, the binding of spike trimers to ACE2 receptors promotes dissociation of the S1 domains and exposure of the fusion machinery, although the molecular details of this process have yet to be observed. We report the development of bottom-up coarse-grained (CG) models consistent with cryo-electron tomography data, and the use of CG molecular dynamics simulations to investigate viral binding and S2 core exposure. We show that spike trimers cooperatively bind to multiple ACE2 dimers at virion-cell interfaces in a manner distinct from binding between soluble proteins, which processively induces S1 dissociation. We also simulate possible variant behavior using perturbed CG models, and find that ACE2-induced S1 dissociation is primarily sensitive to conformational state populations and the extent of S1/S2 cleavage, rather than ACE2 binding affinity. These simulations reveal an important concerted interaction between spike trimers and ACE2 dimers that primes the virus for membrane fusion and entry.

59 BASIC BIOLOGICAL SCIENCES↗