Structure, mechanism and crystallographic fragment screening of the SARS-CoV-2 NSP13 helicase

Abstract

There is currently a lack of effective drugs to treat people infected with SARS-CoV-2, the cause of the global COVID-19 pandemic. The SARS-CoV-2 Non-structural protein 13 (NSP13) has been identified as a target for anti-virals due to its high sequence conservation and essential role in viral replication. Structural analysis reveals two “druggable” pockets on NSP13 that are among the most conserved sites in the entire SARS-CoV-2 proteome. Here we present crystal structures of SARS-CoV-2 NSP13 solved in the APO form and in the presence of both phosphate and a non-hydrolysable ATP analog. Comparisons of these structures reveal details of conformational changes that provide insights into the helicase mechanism and possible modes of inhibition. To identify starting points for drug development we have performed a crystallographic fragment screen against NSP13. The screen reveals 65 fragment hits across 52 datasets opening the way to structure guided development of novel antiviral agents.

Molecular dynamics simulations of the flexibility and inhibition of SARS-CoV-2 NSP 13 helicase

“…These residues are in a similar region to one of the binding pockets identified from our simulations (pocket 3). Our results from the PCA analysis support the conclusion from Newman et al, [ 22 ] that this region could be a good target for allosteric inhibitor design.…”

“…The proposed allosteric site is in a different location on the protein than some of the previously proposed allosteric sites for SARS-Cov and SARS-Cov-2 NSP13 helicase [ 29 , 33 ]. X-ray crystallographic fragment screening found three possible allosteric sites: between the stalk and ZBD domains, between the stalk and 1B domains, and between domains 1A and 2A [ 22 ]. The fragments in this third site (as determined from the 5RMB and 5RMF structures) makes contacts with residues GLN281, LYS430, PRO434, and PHY 437 on domain 1A and residues TYR457, ASP458, and LYS460 on domain 2A.…”

“…The helicases from the two viruses have a close homology, differing only by one residue (570 which is an ILE in SARS-CoV and VAL in SARS-CoV-2). After our simulations were set-up and running, crystal structures of SARS-CoV-2 NSP13 were published [ 22 ]. The apo structure (PDB ID 7NIO ) has two chains, with a Cα root-mean-square deviation (RMSD) of 0.15 nm.…”

“…1 shows the SWISS-MODEL [ 21 ] predicted structure based on 6JYT. Recently, Newman, et al , determined the X-ray structure of SARS-CoV-2 NSP13 helicase, for the apo protein and in complex with phosphate, adenosine 5′-triphosphate, ATP, and other ligands [ 22 ]. The catalytic function of the helicase is to separate double stranded nucleic acids; the protein hydrolyzes nucleoside triphospate molecules (most often ATP), a chemical reaction that ultimately produces the mechanical energy necessary for the helicase to change conformations every step in the unwinding process [ 23 ].…”

Potential Adjuvant Therapeutic Effect of Lactobacillus plantarum Probio-88 Postbiotics against SARS-COV-2

“…The residues at the C-terminal of PlnE, such as Val35, Gly32, and Phe31, formed several hydrogen bonds with Gln537, Arg443, and Lys 288 at the ATP binding site of SARS-COV-2 nsp 13 (Figure 8a). Newman and co-workers recently reported Gln537 and Arg443 as a crucial binding site for ATP hydrolysis [18]. In addition, the Gly43 located at the N-terminal of PlnE also formed a hydrogen bond interaction with one of the ss-RNA interfacing residues, namely Arg212 of SARS-COV-2 nsp 13 (Figure 8a).…”