A novel coronavirus severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) outbreak has caused a global coronavirus disease 2019 (COVID-19) pandemic, resulting in tens of thousands of ...infections and thousands of deaths worldwide. The RNA-dependent RNA polymerase (RdRp), also named nsp12 is the central component of coronaviral replication and transcription machinery, and it appears to be a primary target for the antiviral drug remdesivir. We report the cryo-electron microscopy structure of COVID-19 virus full-length nsp12 in complex with cofactors nsp7 and nsp8 at 2.9-angstrom resolution. In addition to the conserved architecture of the polymerase core of the viral polymerase family, nsp12 possesses a newly identified β-hairpin domain at its N terminus. A comparative analysis model shows how remdesivir binds to this polymerase. The structure provides a basis for the design of new antiviral therapeutics that target viral RdRp.
SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) is the etiological agent responsible for the global COVID-19 (coronavirus disease 2019) outbreak. The main protease of SARS-CoV-2, M
, is ...a key enzyme that plays a pivotal role in mediating viral replication and transcription. We designed and synthesized two lead compounds (
and
) targeting M
Both exhibited excellent inhibitory activity and potent anti-SARS-CoV-2 infection activity. The x-ray crystal structures of SARS-CoV-2 M
in complex with
or
, both determined at a resolution of 1.5 angstroms, showed that the aldehyde groups of
and
are covalently bound to cysteine 145 of M
Both compounds showed good pharmacokinetic properties in vivo, and
also exhibited low toxicity, which suggests that these compounds are promising drug candidates.
Nucleotide analog inhibitors, including broad-spectrum remdesivir and favipiravir, have shown promise in in vitro assays and some clinical studies for COVID-19 treatment, this despite an incomplete ...mechanistic understanding of the viral RNA-dependent RNA polymerase nsp12 drug interactions. Here, we examine the molecular basis of SARS-CoV-2 RNA replication by determining the cryo-EM structures of the stalled pre- and post- translocated polymerase complexes. Compared with the apo complex, the structures show notable structural rearrangements happening to nsp12 and its co-factors nsp7 and nsp8 to accommodate the nucleic acid, whereas there are highly conserved residues in nsp12, positioning the template and primer for an in-line attack on the incoming nucleotide. Furthermore, we investigate the inhibition mechanism of the triphosphate metabolite of remdesivir through structural and kinetic analyses. A transition model from the nsp7-nsp8 hexadecameric primase complex to the nsp12-nsp7-nsp8 polymerase complex is also proposed to provide clues for the understanding of the coronavirus transcription and replication machinery.
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•Structures of SARS-CoV-2 RNA polymerase in complexes with RNA revealed•Conformational changes in nsp8 and its interaction with the exiting RNA are observed•Incorporation and delayed-chain-termination mechanism of remdesivir is elucidated•Transition model from primase complex to polymerase complex is proposed
Cryo-EM structures of the SARS-CoV-2 RNA polymerase in complexes with RNA, before and after RNA translocation, reveals structural rearrangements that the RNA-dependent RNA polymerase (RdRp) nsp12 and its co-factors (nsp7 and nsp8) undergo to accommodate nucleic acid binding. Further insights into how the complex is inhibited by remdesivir, and into the primase to polymerase transition, are also presented.
Abstract
Bandgap control is of central importance for semiconductor technologies. The traditional means of control is to dope the lattice chemically, electrically or optically with charge carriers. ...Here, we demonstrate a widely tunable bandgap (renormalisation up to 550 meV at room-temperature) in two-dimensional (2D) semiconductors by coherently doping the lattice with plasmonic hot electrons. In particular, we integrate tungsten-disulfide (WS
2
) monolayers into a self-assembled plasmonic crystal, which enables coherent coupling between semiconductor excitons and plasmon resonances. Accompanying this process, the plasmon-induced hot electrons can repeatedly fill the WS
2
conduction band, leading to population inversion and a significant reconstruction in band structures and exciton relaxations. Our findings provide an effective measure to engineer optical responses of 2D semiconductors, allowing flexibilities in design and optimisation of photonic and optoelectronic devices.
Abstract
Cytochromes
bd
are ubiquitous amongst prokaryotes including many human-pathogenic bacteria. Such complexes are targets for the development of antimicrobial drugs. However, an understanding ...of the relationship between the structure and functional mechanisms of these oxidases is incomplete. Here, we have determined the 2.8 Å structure of
Mycobacterium smegmatis
cytochrome
bd
by single-particle cryo-electron microscopy. This
bd
oxidase consists of two subunits CydA and CydB, that adopt a pseudo two-fold symmetrical arrangement. The structural topology of its Q-loop domain, whose function is to bind the substrate, quinol, is significantly different compared to the C-terminal region reported for cytochromes
bd
from
Geobacillus thermodenitrificans
(
G. th
) and
Escherichia coli
(
E. coli
). In addition, we have identified two potential oxygen access channels in the structure and shown that similar tunnels also exist in
G. th
and
E. coli
cytochromes
bd
. This study provides insights to develop a framework for the rational design of antituberculosis compounds that block the oxygen access channels of this oxidase.
Abstract
Accumulating mutations in the SARS-CoV-2 Spike (S) protein can increase the possibility of immune escape, challenging the present COVID-19 prophylaxis and clinical interventions. Here, 3 ...receptor binding domain (RBD) specific monoclonal antibodies (mAbs), 58G6, 510A5 and 13G9, with high neutralizing potency blocking authentic SARS-CoV-2 virus display remarkable efficacy against authentic B.1.351 virus. Surprisingly, structural analysis has revealed that 58G6 and 13G9 both recognize the steric region S
470–495
on the RBD, overlapping the E484K mutation presented in B.1.351. Also, 58G6 directly binds to another region S
450–458
in the RBD. Significantly, 58G6 and 510A5 both demonstrate prophylactic efficacy against authentic SARS-CoV-2 and B.1.351 viruses in the transgenic mice expressing human ACE2 (hACE2), protecting weight loss and reducing virus loads. Together, we have evidenced 2 potent neutralizing Abs with unique mechanism targeting authentic SARS-CoV-2 mutants, which can be promising candidates to fulfill the urgent needs for the prolonged COVID-19 pandemic.
Abstract
Realizing optical manipulation of microscopic objects is crucial in the research fields of life science, condensed matter physics, and physical chemistry. In non-liquid environments, this ...task is commonly regarded as difficult due to strong adhesive surface force (~µN) attached to solid interfaces that makes tiny optical driven force (~pN) insignificant. Here, by recognizing the microscopic interaction mechanism between friction force—the parallel component of surface force on a contact surface—and thermoelastic waves induced by pulsed optical absorption, we establish a general principle enabling the actuation of micro-objects on dry frictional surfaces based on the opto-thermo-mechanical effects. Theoretically, we predict that nanosecond pulsed optical absorption with mW-scale peak power is sufficient to tame µN-scale friction force. Experimentally, we demonstrate the two-dimensional spiral motion of gold plates on micro-fibers driven by nanosecond laser pulses, and reveal the rules of motion control. Our results pave the way for the future development of micro-scale actuators in non-liquid environments.
Recombinant adeno-associated viruses (rAAVs) have emerged as promising gene therapy vectors due to their proven efficacy and safety in clinical applications. In non-human primates (NHPs), rAAVs are ...administered via suprachoroidal injection at a higher dose. However, high doses of rAAVs tend to increase additional safety risks. Here, we present a novel AAV capsid (AAVv128), which exhibits significantly enhanced transduction efficiency for photoreceptors and retinal pigment epithelial (RPE) cells, along with a broader distribution across the layers of retinal tissues in different animal models (mice, rabbits, and NHPs) following intraocular injection. Notably, the suprachoroidal delivery of AAVv128-anti-VEGF vector completely suppresses the Grade IV lesions in a laser-induced choroidal neovascularization (CNV) NHP model for neovascular age-related macular degeneration (nAMD). Furthermore, cryo-EM analysis at 2.1 Å resolution reveals that the critical residues of AAVv128 exhibit a more robust advantage in AAV binding, the nuclear uptake and endosome escaping. Collectively, our findings highlight the potential of AAVv128 as a next generation ocular gene therapy vector, particularly using the suprachoroidal delivery route.
The SARS-CoV-2 Omicron variant evades most currently approved neutralizing antibodies (nAbs) and caused drastic decrease of plasma neutralizing activity elicited by vaccination or prior infection, ...urging the need for the development of pan-variant antivirals. Breakthrough infection induces a hybrid immunological response with potentially broad, potent and durable protection against variants, therefore, convalescent plasma from breakthrough infection may provide a broadened repertoire for identifying elite nAbs. We performed single-cell RNA sequencing (scRNA-seq) and BCR sequencing (scBCR-seq) of B cells from BA.1 breakthrough-infected patients who received 2 or 3 previous doses of inactivated vaccine. Elite nAbs, mainly derived from the IGHV2-5 and IGHV3-66/53 germlines, showed potent neutralizing activity across Wuhan-Hu-1, Delta, Omicron sublineages BA.1 and BA.2 at picomolar NT
values. Cryo-EM analysis revealed diverse modes of spike recognition and guides the design of cocktail therapy. A single injection of paired antibodies cocktail provided potent protection in the K18-hACE2 transgenic female mouse model of SARS-CoV-2 infection.
Chronic inflammatory response plays a prominent role in obesity-related nonalcoholic fatty liver disease (NAFLD). However, the intrahepatic triggering mechanism of inflammation remains obscure. This ...study aimed to elucidate the role of serum amyloid A1 (SAA1), an acute-phase response protein, in the obesity-induced hepatic inflammation and NAFLD.
Male mice were fed a high fat diet (HFD) for 16 weeks, and insulin resistance, hepatic steatosis, and inflammation in mice were monitored. Murine SAA1/2 was genetically manipulated to investigate the role of SAA1 in NAFLD.
We found that SAA1 was increased in the NAFLD liver in both humans and mice. Knockout of SAA1/2 or knockdown of hepatic SAA1/2 promoted energy expenditure and alleviated HFD-induced metabolic disorder, hepatic steatosis, and inflammation. Endogenous overexpression of SAA1 in hepatocytes by adeno-associated virus 8 (AAV8) transfection aggravated overnutrition-associated gain of body weight, insulin resistance, hepatic lipid accumulation, and liver injury, which were markedly alleviated by knockout of murine toll-like receptor 4 (TLR4). Mechanistically, SAA1 directly bound with TLR4/myeloid differentiation 2 (MD2) to induce TLR4 internalization, leading to the activation of nuclear factor (NF)-κB signaling and production of both SAA1 and other inflammatory cytokines, including interleukin (IL)-6 and C–C chemokine ligand (CCL2) in hepatocytes. Administration of HFD mice with an AAV8-shRNA-SAA1/2 showed a therapeutic effect on hepatic inflammation and NAFLD progression.
These results demonstrate that SAA1 triggers hepatic steatosis and intrahepatic inflammatory response by forming a SAA1/TLR4/NF-κB/SAA1 feedforward regulatory circuit, which, in turn, leads to NAFLD progression. SAA1 may act as a potential target for the disease intervention.
•SAA1/2 deficiency alleviates HFD-induced hepatic steatosis and inflammation in mice.•SAA1 aggravating overnutrition-associated hepatic steatosis and inflammation is dependent on TLR4.•SAA1 directly binds to TLR4/MD2 to induce TLR4 internalization, leading to the activation of NF-κB signaling .•SAA1/TLR4/NF-κB/SAA1 positive feedback in hepatocytes may be a potential target for obesity associated NAFLD.
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