Structural analysis, virtual screening and molecular simulation to identify potential inhibitors targeting 2'-O-ribose methyltransferase of SARS-CoV-2 coronavirus

Abstract

SARS-CoV-2, an emerging coronavirus, has spread rapidly around the world, resulting in over ten million cases and more than half a million deaths as of July 1, 2020. Effective treatments and vaccines for SARS-CoV-2 infection do not currently exist. Previous studies demonstrated that nonstructural protein 16 (nsp16) of coronavirus is an S-adenosyl methionine (SAM)-dependent 2'-O-methyltransferase (2'-O-MTase) that has an important role in viral replication and prevents recognition by the host innate immune system. In the present study, we employed structural analysis, virtual screening, and molecular simulation approaches to identify clinically investigated and approved drugs which can act as promising inhibitors against nsp16 2'-O-MTase of SARS-CoV-2. Comparative analysis of primary amino acid sequences and crystal structures of seven human CoVs defined the key residues for nsp16 2-O'-MTase functions. Virtual screening and docking analysis ranked the potential inhibitors of nsp16 from more than 4,500 clinically investigated and approved drugs. Furthermore, molecular dynamics simulations were carried out on eight top candidates, including Hesperidin, Rimegepant, Gs-9667, and Sonedenoson, to calculate various structural parameters and understand the dynamic behavior of the drug-protein complexes. Our studies provided the foundation to further test and repurpose these candidate drugs experimentally and/or clinically for COVID-19 treatment.Communicated by Ramaswamy H. Sarma.

Keywords: KDKE motif; SARS-CoV-2; inhibitor; methyltransferase; molecular dynamics simulation; nsp16; virtual screening.

Figure 1.

Comparative analysis of primary amino acid sequences and crystal structures of CoV 2’-O-MTases. (A) Schematic presentation of the SARS-CoV-2 genome organization. Expression of two open reading frames (ORF1a and ORF1b) yields 16 nsps, including 2’-O-MTase nsp16. S, E, M, and N indicate the four structural proteins: spike, envelope, membrane, and nucleocapsid. (B) Sequence alignment of nsp16 proteins derived from genome sequences of the following: SARS-CoV-2, SARS-CoV-1, MERS-CoV, HCoV-OC43, HCoV-HKU1, HCoV-NL63, and HCoV-229E. The secondary structure of SARS-CoV-2 nsp16 is shown above. Residues with 100% conservation are indicated in solid red boxes and those with identity of 70% or higher are depicted in light red color. The red stars indicate the conserved KDKE motif in 2’-O-MTases. (C) Surface representation of the SAM binding pocket and the RNA binding groove in SARS-CoV-2 nsp16 with coloring according to the electrostatic potential. The surface electrostatic potential diagram (±5 kT/e) in SARS-CoV-2 nsp16 (PDB: 6W4H Chain A) was generated by PyMol; the blue areas represent positively charged areas, while the red areas represent negatively charged areas. (D) The KDKE catalytic tetrad motif is located at the bottom of the RNA binding groove of nsp16.

Figure 2.

Comparison of 2’-O-MTase domain of human CMTR1 with nsp16 of SARS-CoV-2. Top: Surface representation with electrostatic potentials showing the RNA binding groove and the SAM binding pocket of human CMTR1 (PDB: 4N48). Bottom: Superposition of the KDKE catalytic tetrad motifs of nsp16 of SARS-CoV-2 (PDB: 6W4H) and 2’-O-MTase domain of human CMTR1 (PDB: 4N48).

Figure 3.

(A) Similarity chart of the 1,380 top-scoring compounds for the SARS-CoV-2 nsp16 using the FragFp descriptors in DataWarrior. Compounds with high chemical similarity are connected by lines. Stereoisomers of the same drug such as Hesperidin (38 Stereoisomers) are clustered together. Compounds are colored according to their predicted AutoDock Vina score. (B) Neighbor tree shows seven drugs that have FragFp similarity scores to SAM greater than 0.8. Drugs are colored according to their FragFp similarity score to SAM. (C) Pie charts show target and pathway classes of hit compounds that were queried in the Probes and Drugs Portal.

Figure 4.

Interaction of four drugs (Hesperidin, Osi-027, Gs-9667 and Sonedenoson) with SAM binding pocket of SARS-CoV-2 nsp16. Left: Surface representation with charge showing predicted binding of SARS-CoV-2 nsp16 with the four drugs in the SAM binding pocket. The drugs are depicted by sticks. Right: 3 D representation of 2’-O-MTase active site residues interacting with Hesperidin, Osi-027, Gs-9667 and Sonedenoson.

Figure 5.

Plot of MD simulation trajectories of free 2’-O-MTase and drug-protein complexes during 100 ns simulation. RMSD: Root Mean Square Deviation; RMSF: Root Mean Square Fluctuation; SASA: Solvent Accessible Surface Area; H-Bond: hydrogen bond.

Figure 6.

Principal Component Analysis (PCA) of free 2’-O-MTase, along with eight drug- and SAM-protein complexes. The bottom shows a superimposed plot of all ten PCA analyses.

Figure 7.

The per-residue free energy contribution spectrums of nsp16 in four compound-protein complexes. Only residues contributing above a ± 10 kcal/mol threshold were colored and labeled.