DDQ/Fe(NO3)3-Catalyzed Aerobic Synthesis of 3-Acyl Indoles and an In Silico Study for the Binding Affinity of N-Tosyl-3-acyl Indoles toward RdRp against SARS-CoV-2.

In the present study, we herein report a DDQ-catalyzed new protocol for the synthesis of substituted 3-acylindoles. Being a potential system for virtual hydrogen storage, introduction of catalytic DDQ in combination with Fe(NO3)3·9H2O and molecular oxygen as co-catalysts offers a regioselective oxo-functionalization of C-3 alkyl-/aryllidine indolines even with scale-up investigations. Intermediate isolation, their spectroscopic characterization, and the density functional theory calculations indicate that the method involves dehydrogenative allylic hydroxylation and 1,3-functional group isomerization/aromatization followed by terminal oxidation to afford 3-acylindoles quantitatively with very high regioselectivity. This method is very general for a large number of substrates with varieties of functional groups tolerance emerging high-yield outcome. Moreover, molecular docking studies were performed for some selected ligands with an RNA-dependent RNA polymerase complex (RdRp complex) of SARS-CoV-2 to illustrate the binding potential of those ligands. The docking results revealed that few of the ligands possess the potential to inhibit the RdRp of SARS-Cov-2 with binding energies (-6.7 to -8.19 kcal/mol), which are comparably higher with respect to the reported binding energies of the conventional re-purposed drugs such as Remdesivir, Ribavirin, and so forth (-4 to -7 kcal/mol).

SARS-CoV-2 hijacks cellular kinase CDK2 to promote viral RNA synthesis.

The coronavirus disease 2019 (COVID-19) pandemic has devastated global health. Identifying key host factors essential for SARS-CoV-2 RNA replication is expected to unravel cellular targets for the development of broad-spectrum antiviral drugs which have been quested for the preparedness of future viral outbreaks. Here, we have identified host proteins that associate with nonstructural protein 12 (nsp12), the RNA-dependent RNA polymerase (RdRp) of SARS-CoV-2 using a mass spectrometry (MS)-based proteomic approach. Among the candidate factors, CDK2 (Cyclin-dependent kinase 2), a member of cyclin-dependent kinases, interacts with nsp12 and causes its phosphorylation at T20, thus facilitating the assembly of the RdRp complex consisting of nsp12, nsp7 and nsp8 and promoting efficient synthesis of viral RNA. The crucial role of CDK2 in viral RdRp function is further supported by our observation that CDK2 inhibitors potently impair viral RNA synthesis and SARS-CoV-2 infection. Taken together, we have discovered CDK2 as a key host factor of SARS-CoV-2 RdRp complex, thus serving a promising target for the development of SARS-CoV-2 RdRp inhibitors.

SARS-CoV-2 hijacks cellular kinase CDK2 to promote viral RNA synthesis.

The coronavirus disease 2019 (COVID-19) pandemic has devastated global health. Identifying key host factors essential for SARS-CoV-2 RNA replication is expected to unravel cellular targets for the development of broad-spectrum antiviral drugs which have been quested for the preparedness of future viral outbreaks. Here, we have identified host proteins that associate with nonstructural protein 12 (nsp12), the RNA-dependent RNA polymerase (RdRp) of SARS-CoV-2 using a mass spectrometry (MS)-based proteomic approach. Among the candidate factors, CDK2 (Cyclin-dependent kinase 2), a member of cyclin-dependent kinases, interacts with nsp12 and causes its phosphorylation at T20, thus facilitating the assembly of the RdRp complex consisting of nsp12, nsp7 and nsp8 and promoting efficient synthesis of viral RNA. The crucial role of CDK2 in viral RdRp function is further supported by our observation that CDK2 inhibitors potently impair viral RNA synthesis and SARS-CoV-2 infection. Taken together, we have discovered CDK2 as a key host factor of SARS-CoV-2 RdRp complex, thus serving a promising target for the development of SARS-CoV-2 RdRp inhibitors.

DDQ/Fe(NO3)3-Catalyzed Aerobic Synthesis of 3-Acyl Indoles and an In Silico Study for the Binding Affinity of N-Tosyl-3-acyl Indoles toward RdRp against SARS-CoV-2.

In the present study, we herein report a DDQ-catalyzed new protocol for the synthesis of substituted 3-acylindoles. Being a potential system for virtual hydrogen storage, introduction of catalytic DDQ in combination with Fe(NO3)3·9H2O and molecular oxygen as co-catalysts offers a regioselective oxo-functionalization of C-3 alkyl-/aryllidine indolines even with scale-up investigations. Intermediate isolation, their spectroscopic characterization, and the density functional theory calculations indicate that the method involves dehydrogenative allylic hydroxylation and 1,3-functional group isomerization/aromatization followed by terminal oxidation to afford 3-acylindoles quantitatively with very high regioselectivity. This method is very general for a large number of substrates with varieties of functional groups tolerance emerging high-yield outcome. Moreover, molecular docking studies were performed for some selected ligands with an RNA-dependent RNA polymerase complex (RdRp complex) of SARS-CoV-2 to illustrate the binding potential of those ligands. The docking results revealed that few of the ligands possess the potential to inhibit the RdRp of SARS-Cov-2 with binding energies (-6.7 to -8.19 kcal/mol), which are comparably higher with respect to the reported binding energies of the conventional re-purposed drugs such as Remdesivir, Ribavirin, and so forth (-4 to -7 kcal/mol).

On the Recognition of Natural Substrate CTP and Endogenous Inhibitor ddhCTP of SARS-CoV-2 RNA-Dependent RNA Polymerase: A Molecular Dynamics Study.

The novel coronavirus SARS-CoV-2 is the causative agent of the COVID-19 outbreak that is affecting the entire planet. As the pandemic is still spreading worldwide, with multiple mutations of the virus, it is of interest and of help to employ computational methods for identifying potential inhibitors of the enzymes responsible for viral replication. Attractive antiviral nucleotide analogue RNA-dependent RNA polymerase (RdRp) chain terminator inhibitors are investigated with this purpose. This study, based on molecular dynamics (MD) simulations, addresses the important aspects of the incorporation of an endogenously synthesized nucleoside triphosphate, ddhCTP, in comparison with the natural nucleobase cytidine triphosphate (CTP) in RdRp. The ddhCTP species is the product of the viperin antiviral protein as part of the innate immune response. The absence of the ribose 3'-OH in ddhCTP could have important implications in its inhibitory mechanism of RdRp. We built an in silico model of the RNA strand embedded in RdRp using experimental methods, starting from the cryo-electron microscopy structure and exploiting the information obtained by spectrometry on the RNA sequence. We determined that the model was stable during the MD simulation time. The obtained results provide deeper insights into the incorporation of nucleoside triphosphates, whose molecular mechanism by the RdRp active site still remains elusive.

A novel virtual screening procedure identifies Pralatrexate as inhibitor of SARS-CoV-2 RdRp and it reduces viral replication in vitro.

The spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus poses serious threats to the global public health and leads to worldwide crisis. No effective drug or vaccine is readily available. The viral RNA-dependent RNA polymerase (RdRp) is a promising therapeutic target. A hybrid drug screening procedure was proposed and applied to identify potential drug candidates targeting RdRp from 1906 approved drugs. Among the four selected market available drug candidates, Pralatrexate and Azithromycin were confirmed to effectively inhibit SARS-CoV-2 replication in vitro with EC50 values of 0.008µM and 9.453 µM, respectively. For the first time, our study discovered that Pralatrexate is able to potently inhibit SARS-CoV-2 replication with a stronger inhibitory activity than Remdesivir within the same experimental conditions. The paper demonstrates the feasibility of fast and accurate anti-viral drug screening for inhibitors of SARS-CoV-2 and provides potential therapeutic agents against COVID-19.

Targeting the RdRp of Emerging RNA Viruses: The Structure-Based Drug Design Challenge.

The RNA-dependent RNA polymerase (RdRp) is an essential enzyme for the viral replication process, catalyzing the viral RNA synthesis using a metal ion-dependent mechanism. In recent years, RdRp has emerged as an optimal target for the development of antiviral drugs, as demonstrated by recent approvals of sofosbuvir and remdesivir against Hepatitis C virus (HCV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), respectively. In this work, we overview the main sequence and structural features of the RdRp of emerging RNA viruses such as Coronaviruses, Flaviviruses, and HCV, as well as inhibition strategies implemented so far. While analyzing the structural information available on the RdRp of emerging RNA viruses, we provide examples of success stories such as for HCV and SARS-CoV-2. In contrast, Flaviviruses' story has raised attention about how the lack of structural details on catalytically-competent or ligand-bound RdRp strongly hampers the application of structure-based drug design, either in repurposing and conventional approaches.

In silico studies evidenced the role of structurally diverse plant secondary metabolites in reducing SARS-CoV-2 pathogenesis.

Plants are endowed with a large pool of structurally diverse small molecules known as secondary metabolites. The present study aims to virtually screen these plant secondary metabolites (PSM) for their possible anti-SARS-CoV-2 properties targeting four proteins/ enzymes which govern viral pathogenesis. Results of molecular docking with 4,704 ligands against four target proteins, and data analysis revealed a unique pattern of structurally similar PSM interacting with the target proteins. Among the top-ranked PSM which recorded lower binding energy (BE), > 50% were triterpenoids which interacted strongly with viral spike protein-receptor binding domain, > 32% molecules which showed better interaction with the active site of human transmembrane serine protease were belongs to flavonoids and their glycosides, > 16% of flavonol glycosides and > 16% anthocyanidins recorded lower BE against active site of viral main protease and > 13% flavonol glycoside strongly interacted with active site of viral RNA-dependent RNA polymerase. The primary concern about these PSM is their bioavailability. However, several PSM recorded higher bioavailability score and found fulfilling most of the drug-likeness characters as per Lipinski's rule (Coagulin K, Kamalachalcone C, Ginkgetin, Isoginkgetin, 3,3'-Biplumbagin, Chrysophanein, Aromoline, etc.). Natural occurrence, bio-transformation, bioavailability of selected PSM and their interaction with the target site of selected proteins were discussed in detail. Present study provides a platform for researchers to explore the possible use of selected PSM to prevent/ cure the COVID-19 by subjecting them for thorough in vitro and in vivo evaluation for the capabilities to interfering with the process of viral host cell recognition, entry and replication.

Viral macrodomains: a structural and evolutionary assessment of the pharmacological potential.

Viral macrodomains possess the ability to counteract host ADP-ribosylation, a post-translational modification implicated in the creation of an antiviral environment via immune response regulation. This brought them into focus as promising therapeutic targets, albeit the close homology to some of the human macrodomains raised concerns regarding potential cross-reactivity and adverse effects for the host. Here, we evaluate the structure and function of the macrodomain of SARS-CoV-2, the causative agent of COVID-19. We show that it can antagonize ADP-ribosylation by PARP14, a cellular (ADP-ribosyl)transferase necessary for the restriction of coronaviral infections. Furthermore, our structural studies together with ligand modelling revealed the structural basis for poly(ADP-ribose) binding and hydrolysis, an emerging new aspect of viral macrodomain biology. These new insights were used in an extensive evolutionary analysis aimed at evaluating the druggability of viral macrodomains not only from the Coronaviridae but also Togaviridae and Iridoviridae genera (causing diseases such as Chikungunya and infectious spleen and kidney necrosis virus disease, respectively). We found that they contain conserved features, distinct from their human counterparts, which may be exploited during drug design.

Remdesivir targets a structurally analogous region of the Ebola virus and SARS-CoV-2 polymerases.

Remdesivir is a broad-spectrum antiviral nucleotide prodrug that has been clinically evaluated in Ebola virus patients and recently received emergency use authorization (EUA) for treatment of COVID-19. With approvals from the Federal Select Agent Program and the Centers for Disease Control and Prevention's Institutional Biosecurity Board, we characterized the resistance profile of remdesivir by serially passaging Ebola virus under remdesivir selection; we generated lineages with low-level reduced susceptibility to remdesivir after 35 passages. We found that a single amino acid substitution, F548S, in the Ebola virus polymerase conferred low-level reduced susceptibility to remdesivir. The F548 residue is highly conserved in filoviruses but should be subject to specific surveillance among novel filoviruses, in newly emerging variants in ongoing outbreaks, and also in Ebola virus patients undergoing remdesivir therapy. Homology modeling suggests that the Ebola virus polymerase F548 residue lies in the F-motif of the polymerase active site, a region that was previously identified as susceptible to resistance mutations in coronaviruses. Our data suggest that molecular surveillance of this region of the polymerase in remdesivir-treated COVID-19 patients is also warranted.

Both Boceprevir and GC376 efficaciously inhibit SARS-CoV-2 by targeting its main protease.

COVID-19 was declared a pandemic on March 11 by WHO, due to its great threat to global public health. The coronavirus main protease (Mpro, also called 3CLpro) is essential for processing and maturation of the viral polyprotein, therefore recognized as an attractive drug target. Here we show that a clinically approved anti-HCV drug, Boceprevir, and a pre-clinical inhibitor against feline infectious peritonitis (corona) virus (FIPV), GC376, both efficaciously inhibit SARS-CoV-2 in Vero cells by targeting Mpro. Moreover, combined application of GC376 with Remdesivir, a nucleotide analogue that inhibits viral RNA dependent RNA polymerase (RdRp), results in sterilizing additive effect. Further structural analysis reveals binding of both inhibitors to the catalytically active side of SARS-CoV-2 protease Mpro as main mechanism of inhibition. Our findings may provide critical information for the optimization and design of more potent inhibitors against the emerging SARS-CoV-2 virus.

Is oseltamivir suitable for fighting against COVID-19: In silico assessment, in vitro and retrospective study.

BACKGROUND: Oseltamivir is a first-line antiviral drug, especially in primary hospitals. During the ongoing outbreak of coronavirus disease 2019 (COVID-19), most patients with COVID-19 who are symptomatic have used oseltamivir. Considering its popular and important role as an antiviral drug, it is necessary to evaluate oseltamivir in the treatment of COVID-19. OBJECTIVE: To evaluate the effect of oseltamivir against COVID-19. METHODS: Swiss-model was used to construct the structure of the N-terminal RNA-binding domain (NRBD) of the nucleoprotein (NC), papain-like protease (PLpro), and RNA-directed RNA polymerase (RdRp) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). TM-align program was performed to compare the structure of the viral proteins with the structure of the neuraminidase of influenza A. Molecular docking was used to analyze the theoretical possibility of effective binding of oseltamivir with the active centers of the viral proteins. In vitro study was used to evaluate the antiviral efficiency of oseltamivir against SARS-CoV-2. By clinical case analysis, we statistically evaluated whether the history of oseltamivir use influenced the progression of the disease. RESULTS: The structures of NRBD, PLpro, and RdRp were built successfully. The results from TM-align suggested that the S protein, NRBD, 3C-like protease (3CLpro), PLPrO, and RdRp were structurally similar to the influenza A neuraminidase, with TM-scores of 0.30077, 0.19254, 0.28766, 0.30666, and 0.34047, respectively. Interestingly, the active center of 3CL pro was found to be similar to the active center from the neuraminidase of influenza A. Through an analysis of molecular docking, we discovered that oseltamivir carboxylic acid was more favorable to bind to the active site of 3CLpro effectively, but its inhibitory effect was not strong compared with the positive group. Finally, we used in vitro study and retrospective case analysis to verify our speculations. We found that oseltamivir is ineffective against SARS-CoV-2 in vitro study and the clinical use of oseltamivir did not improve the patients' symptoms and signs and did not slow the disease progression. CONCLUSIONS: We consider that oseltamivir isn't suitable for the treatment of COVID-19. During the outbreak of novel coronavirus, when oseltamivir is not effective for the patients after they take it, health workers should be highly vigilant about the possibility of COVID-19.

Molecular modelling of the antiviral action of Resveratrol derivatives against the activity of two novel SARS CoV-2 and 2019-nCoV receptors.

The pandemic threat of COVID-19 causes serious concern for people and world organizations. The effect of Coronavirus disease on the lifestyle and economic status of humans is undeniable, and all of the researchers (biologists, pharmacists, physicians, and chemists) can help decrease its destructive effects. The molecular docking approach can provide a fast prediction of the positive influence the targets on the COVID-19 outbreak. In this work, we choose resveratrol (RV) derivatives (22 cases) and two newly released coordinate structures for COVID-19 as receptors [Papain-like Protease of SARS CoV-2 (PBD ID: 6W9C) and 2019-nCoV RNA-dependent RNA Polymerase (PBD ID: 6M71)]. The results show that conformational isomerism is significant and useful parameter for docking results. A wide spectrum of interactions such as Van der Waals, conventional hydrogen bond, Pi-donor hydrogen bond, Pi-Cation, Pi-sigma, Pi-Pi stacked, Amide-Pi stacked and Pi-Alkyl is detected via docking of RV derivatives and COVID-19 receptors. The potential inhibition effect of RV-13 (-184.99 kj/mol), and RV-12 (-173.76 kj/mol) is achieved at maximum value for 6W9C and 6M71, respectively.

Nucleotide Analogues as Inhibitors of SARS-CoV-2 Polymerase, a Key Drug Target for COVID-19.

SARS-CoV-2 is responsible for the current COVID-19 pandemic. On the basis of our analysis of hepatitis C virus and coronavirus replication, and the molecular structures and activities of viral inhibitors, we previously demonstrated that three nucleotide analogues (the triphosphates of Sofosbuvir, Alovudine, and AZT) inhibit the SARS-CoV RNA-dependent RNA polymerase (RdRp). We also demonstrated that a library of additional nucleotide analogues terminate RNA synthesis catalyzed by the SARS-CoV-2 RdRp, a well-established drug target for COVID-19. Here, we used polymerase extension experiments to demonstrate that the active triphosphate form of Sofosbuvir (an FDA-approved hepatitis C drug) is incorporated by SARS-CoV-2 RdRp and blocks further incorporation. Using the molecular insight gained from the previous studies, we selected the active triphosphate forms of six other antiviral agents, Alovudine, Tenofovir alafenamide, AZT, Abacavir, Lamivudine, and Emtricitabine, for evaluation as inhibitors of the SARS-CoV-2 RdRp and demonstrated the ability of these viral polymerase inhibitors to be incorporated by SARS-CoV-2 RdRp, where they terminate further polymerase extension with varying efficiency. These results provide a molecular basis for inhibition of the SARS-CoV-2 RdRp by these nucleotide analogues. If sufficient efficacy of some of these FDA-approved drugs in inhibiting viral replication in cell culture is established, they may be explored as potential COVID-19 therapeutics.

Analysis of SARS-CoV-2 RNA-dependent RNA polymerase as a potential therapeutic drug target using a computational approach.

BACKGROUND: The Severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) outbreak originating in Wuhan, China, has raised global health concerns and the pandemic has now been reported on all inhabited continents. Hitherto, no antiviral drug is available to combat this viral outbreak. METHODS: Keeping in mind the urgency of the situation, the current study was designed to devise new strategies for drug discovery and/or repositioning against SARS-CoV-2. In the current study, RNA-dependent RNA polymerase (RdRp), which regulates viral replication, is proposed as a potential therapeutic target to inhibit viral infection. RESULTS: Evolutionary studies of whole-genome sequences of SARS-CoV-2 represent high similarity (> 90%) with other SARS viruses. Targeting the RdRp active sites, ASP760 and ASP761, by antiviral drugs could be a potential therapeutic option for inhibition of coronavirus RdRp, and thus viral replication. Target-based virtual screening and molecular docking results show that the antiviral Galidesivir and its structurally similar compounds have shown promise against SARS-CoV-2. CONCLUSIONS: The anti-polymerase drugs predicted here-CID123624208 and CID11687749-may be considered for in vitro and in vivo clinical trials.

RNA-Dependent RNA Polymerase as a Target for COVID-19 Drug Discovery.

COVID-19 respiratory disease caused by the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has rapidly become a global health issue since it emerged in December 2019. While great global efforts are underway to develop vaccines and to discover or repurpose therapeutic agents for this disease, as of this writing only the nucleoside drug remdesivir has been approved under Emergency Use Authorization to treat COVID-19. The RNA-dependent RNA polymerase (RdRP), a viral enzyme for viral RNA replication in host cells, is one of the most intriguing and promising drug targets for SARS-CoV-2 drug development. Because RdRP is a viral enzyme with no host cell homologs, selective SARS-CoV-2 RdRP inhibitors can be developed that have improved potency and fewer off-target effects against human host proteins and thus are safer and more effective therapeutics for treating COVID-19. This review focuses on biochemical enzyme and cell-based assays for RdRPs that could be used in high-throughput screening to discover new and repurposed drugs against SARS-CoV-2.

Intermolecular interaction among Remdesivir, RNA and RNA-dependent RNA polymerase of SARS-CoV-2 analyzed by fragment molecular orbital calculation.

COVID-19, a disease caused by a new strain of coronavirus (SARS-CoV-2) originating from Wuhan, China, has now spread around the world, triggering a global pandemic, leaving the public eagerly awaiting the development of a specific medicine and vaccine. In response, aggressive efforts are underway around the world to overcome COVID-19. In this study, referencing the data published on the Protein Data Bank (PDB ID: 7BV2) on April 22, we conducted a detailed analysis of the interaction between the complex structures of the RNA-dependent RNA polymerase (RdRp) of SARS-CoV-2 and Remdesivir, an antiviral drug, from the quantum chemical perspective based on the fragment molecular orbital (FMO) method. In addition to the hydrogen bonding and intra-strand stacking between complementary strands as seen in normal base pairs, Remdesivir bound to the terminus of an primer-RNA strand was further stabilized by diagonal π-π stacking with the -1A' base of the complementary strand and an additional hydrogen bond with an intra-strand base, due to the effect of chemically modified functional group. Moreover, stable OH/π interaction is also formed with Thr687 of the RdRp. We quantitatively revealed the exhaustive interaction within the complex among Remdesivir, template-primer-RNA, RdRp and co-factors, and published the results in the FMODB database.

Phylogenetic Analysis and Structural Perspectives of RNA-Dependent RNA-Polymerase Inhibition from SARs-CoV-2 with Natural Products.

Most recently, an outbreak of severe pneumonia caused by the infection of SARS-CoV-2, a novel coronavirus first identified in Wuhan, China, imposes serious threats to public health. Upon infecting host cells, coronaviruses assemble a multi-subunit RNA-synthesis complex of viral non-structural proteins (nsp) responsible for the replication and transcription of the viral genome. Therefore, the role and inhibition of nsp12 are indispensable. A cryo-EM structure of RdRp from SARs-CoV-2 was used to identify novel drugs from Northern South African medicinal compounds database (NANPDB) by using computational virtual screening and molecular docking approaches. Considering Remdesivir as the control, 42 compounds were shortlisted to have docking score better than Remdesivir. The top 5 hits were validated by using molecular dynamics simulation approach and free energy calculations possess strong inhibitory properties than the Remdesivir. Thus, this study paved a way for designing novel drugs by decoding the architecture of an important enzyme and its inhibition with compounds from natural resources. This disclosing of necessary knowledge regarding the screening and the identification of top hits could help to design effective therapeutic candidates against the coronaviruses and design robust preventive measurements.

Interaction of hydroxychloroquine with SARS-CoV2 functional proteins using all-atoms non-equilibrium alchemical simulations.

Using a combination of enhanced sampling molecular dynamics techniques and non-equilibrium alchemical transformations with full atomistic details, we have shown that hydroxychloroquine (HCQ) may act as a mild inhibitor of important functional proteins for SARS-CoV2 replication, with potency increasing in the series PLpro, 3CLpro, RdRp. By analyzing the bound state configurations, we were able to improve the potency for the 3CLpro target, designing a novel HCQ-inspired compound, named PMP329, with predicted nanomolar activity. If confirmed in vitro, our results provide a molecular rationale for the use of HCQ or of strictly related derivatives in the treatment of Covid-19.

Structural and Biochemical Characterization of the nsp12-nsp7-nsp8 Core Polymerase Complex from SARS-CoV-2.

The ongoing global pandemic of coronavirus disease 2019 (COVID-19) has caused a huge number of human deaths. Currently, there are no specific drugs or vaccines available for this virus (SARS-CoV-2). The viral polymerase is a promising antiviral target. Here, we describe the near-atomic-resolution structure of the SARS-CoV-2 polymerase complex consisting of the nsp12 catalytic subunit and nsp7-nsp8 cofactors. This structure highly resembles the counterpart of SARS-CoV with conserved motifs for all viral RNA-dependent RNA polymerases and suggests a mechanism of activation by cofactors. Biochemical studies reveal reduced activity of the core polymerase complex and lower thermostability of individual subunits of SARS-CoV-2 compared with SARS-CoV. These findings provide important insights into RNA synthesis by coronavirus polymerase and indicate adaptation of SARS-CoV-2 toward humans with a relatively lower body temperature than the natural bat hosts.