Mikrobiol. Z. 2022; 84(1):34-43.
doi: https://doi.org/10.15407/microbiolj84.01.034

In silico Identification of a Viral Surface Glycoprotein Site Suitable for the Development of
Low Molecular Weight Inhibitors for Various Variants of the SARS-CoV-2

A.A. Zaremba, P.Y. Zaremba, F.V. Muchnyk, G.V. Baranova, S.D. Zahorodnia

Zabolotny Institute of Microbiology and Virology, NAS of Ukraine
154 Acad. Zabolotny Str., Kyiv, 03143, Ukraine

Severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) is a new coronavirus that today has an extremely significant impact on both global economy and society as a whole, due to its pandemic status and risk of complications. Therefore, understanding the molecular features of the interaction of receptor binding domain (RBD), which determines most of the dangerous properties of this pathogen, with human angiotensin-converting enzyme 2 (hACE2) is an important step in the process of developing a successful strategy to combat SARS-CoV-2. In addition, given the significant rate of accumulation of mutations in RBD, it makes sense to consider its different variants. Goal. Identification of a pocket potentially suitable for the search for low molecular weight inhibitors of interaction of different variants of SARS-CoV-2 RBD and hACE2. Methods. The initial structure of different variants of the RBD/hACE2 complex was obtained from Protein Data Bank (PDB). Separate RBD variants were isolated from the same data. To obtain the Y453F mutant, variant P.1 was mutagenized in PyMol 1.8. The construction of the system, which included the resulting associate or individual protein, solvent and physiological concentration of sodium chloride, was performed using CHARMM-GUI (graphical user interface for CHARMM) tools according to the standard protocol for glycoproteins. The actual simulation and balancing of the system was performed in GROMACS (GROningen MAchine for Chemical Simulation) version 2019.6 for 50 ns. Results. The interface of RBD/hACE2 interaction is formed by amino acids Q24, D30, H34, E35, E37, Y41, Y83, K353, D355 and R393 – for hACE2 and K417, Y453, F486, N487, Y489, Q493, Q498, T500, N501, Y505 – for RBD. However, it is heterogeneous and can be divided into two subinterfaces, and each includes its own pool of interactions: hACE2 Q24/Y83 + RBD N487, hACE2 H34 + RBD Y453, hACE2 E35 + RBD Q493 and hACE2 D30 + RBD K417 – for N- terminal relative to H1 hACE2 subinterface and hACE2 E37/R393 + RBD Y505, hACE2 K353 + RBD Q498/G502 and hACE2 D355 + RBD T500 – for C-terminal. According to the considered N501Y mutation, changes are observed in the mentioned interaction patterns – hydrogen bonds of hACE2 Q42 + RBD Q498, hACE2 K31 + RBD Q493 and hACE2 K31 + RBD F490 are formed, and hACE2 H34 + RBD Y453 is lost. Similar aberrations, except for the hydrogen bond with F490, are observed in the case of the N501Y + Y453F variant. Despite significant changes in the pool of interactions, the gross number of hydrogen bonds for the complexes of all three variants is relatively stable and ranges from 9 to 10. Between the defined interaction subinterfaces for all considered variants of RBD are characterized by the presence of a pocket, which is formed by residues R403, Y453, Q493, S494, Y495, G496, F497, Q498, N501 and Y505 conditionally original variant. According to the results of the molecular dynamics simulation, the Y453F replacement has little effect on the overall topology of the cavity, but sufficiently reduces the polarity of the pocket part of its localization, which leads to the impossibility of forming any polar interactions. In contrast, N501Y, due to the larger size of the tyrosine radical and the presence of parahydroxyl, forms two equivalent mutually exclusive hydrogen bonds with the carbonyls of the peptide groups G496 and Y495. Additional stabilization of the Y501 is provided by interplanar stacking with the Y505. In addition to the anchored position in ~ 25% of the trajectory there is another “open” conformation Y501. At which the radical of this tyrosine does not interact with the rest of the protein. Conclusions. 1) The interface of interaction of SARS-CoV-2 RBD with hACE2 is not continuous and it can be conditionally divided into two subiterfaces: N-terminal and C-terminal. Each is characterized by its own pattern of connections and changes according to the RBD N501Y and Y453F replacements we have considered. However, despite the presence of significant molecular rearrangements caused by N501Y and Y453F, the total number of hydrogen bonds is relatively the same for all mutants. 2) Between the identified interaction subinterfaces, SARS-CoV-2 RBD contains caveola, which due to its location may be potentially suitable for finding promising candidates for drugs aimed at inhibiting the interaction of this protein with hACE2. In this case, the substitutions of N501Y and Y453F have a significant impact on the topology of a particular pocket and can potentially modify the activity of inhibitors directed to this area.

Keywords: SARS-CoV-2, receptor binding domain (RBD), mutation N501Y, mutation Y453F, human angiotensin-converting enzyme 2 (hACE2), GROMACS, CHARMM-GUI.

Full text (PDF, in English)

  1. Syed A, Khan A, Gosai F, Asif A, Dhillon S. Gastrointestinal pathophysiology of SARS-CoV-2 – a literature review. Journal of community hospital internal medicine perspectives. 2020; 10(6):523–28. https://doi.org/10.1080/20009666.2020.1811556
  2. Dong E, Du H, Gardner L. An interactive webbased dashboard to track COVID-19 in real time. Lancet Infect Dis. 2020; 20(5):533–34. https://doi.org/10.1016/S1473-3099(20)30120-1
  3. Chan-Yeung M, Xu RH. SARS: epidemiology. Respirology. 2003; 8(1):9–14. https://doi.org/10.1046/j.1440-1843.2003.00518.x
  4. Sanyaolu A, Okorie C, Marinkovic A, Haider N, Abbasi AF, Jaferi U, Prakash S, Balendra V. The emerging SARS-CoV-2 variants of concern. Ther Adv Infect Dis. 2021; 8:20499361211024372. https://doi.org/10.1177/20499361211024372
  5. Berman HM, Westbrook J, Feng Z, Gilliland G, Bhat TN, Weissig H, Shindyalov IN, Bourne PE. The Protein Data  Bank. Nucleic Acids Research. 2000; 28:235–42. https://doi.org/10.1093/nar/28.1.235
  6. The PyMOL (1.7.4.4 Edu) Molecular Graphics System, Version 1.8 Schrödinger, LLC.
  7. Lee J, Cheng X, Swails JM, Yeom MS, Eastman PK, Lemkul JA, et al. CHARMM-GUI Input Generator for NAMD, GROMACS, AMBER, OpenMM, and CHARMM/OpenMM Simulations using the CHARMM36 Additive Force Field. J Chem Theory Comput. 2016; 12:405–13. https://doi.org/10.1021/acs.jctc.5b00935
  8. Abraham MJ, Murtola T, Schulz R, Páll S, Smith JC, Hess B, Lindahl E. GROMACS: High performance molecular simulations through multi-level parallelism from laptops to supercomputers. SoftwareX. 2015; 1–2:19–25. https://doi.org/10.1016/j.softx.2015.06.001
  9. Duan L, Zheng Q, Zhang H, Niu Y, Lou Y, Wang H. The SARS-CoV-2 Spike Glycoprotein Biosynthesis, Structure, Function, and Antigenicity: Implications for the Design of Spike-Based Vaccine Immunogens. Front Immunol. 2020; 11:576622. https://doi.org/10.3389/fimmu.2020.576622
  10. Ou X, Liu Y, Lei X, Li P, Mi D, Ren L, Guo L, Guo R, Chen T, Hu J, Xiang Z, Mu Z, Chen X, Chen J, Hu K, Jin Q, Wang J, Qian Z. Characterization of spike glycoprotein of SARS-CoV-2 on virus entry and its immune cross-reactivity with SARS-CoV. Nat Commun. 2020; 11(1):1620. https://doi.org/10.1038/s41467-020-15562-9
  11. Walls AC, Park YJ, Tortorici MA, Wall A, McGuire AT, Veesler D. Structure, Function, and Antigenicity of the SARS-CoV-2 Spike Glycoprotein. Cell. 2020; 181(2):281–92. https://doi.org/10.1016/j.cell.2020.02.058
  12. Behloul N, Baha S, Guo Y, Yang Z, Shi R, Meng J. In silico identification of strong binders of the SARS-CoV-2 receptor-binding domain. Eur J Pharmacol. 2021; 890:173701. https://doi.org/10.1016/j.ejphar.2020.173701
  13. Gervasoni S, Vistoli G, Talarico C, Manelfi C, Beccari AR, Studer G, Tauriello G, Waterhouse AM, Schwede T, Pedretti AA. Comprehensive Mapping of the Druggable Cavities within the SARS-CoV-2 Therapeutically Relevant Proteins by Combining Pocket and Docking Searches as Implemented in Pockets 2.0. Int J Mol Sci. 2020; 21(14):5152. https://doi.org/10.3390/ijms21145152
  14. Feng S, Luan X, Wang Y, Wang H, Zhang Z, Wang Y, Tian Z, Liu M, Xiao Y, Zhao Y, Zhou R, Zhang S. Eltrombopag is a potential target for drug intervention in SARS-CoV-2 spike protein. Infect Genet Evol. 2020; 85:104419. https://doi.org/10.1016/j.meegid.2020.104419
  15. Hacisuleyman E, Hale C, Saito Y, Blachere NE, Bergh M, Conlon EG, Schaefer-Babajew DJ, DaSilva J, Muecksch F, Gaebler C, Lifton R, Nussenzweig MC, Hatziioannou T, Bieniasz PD, Darnell RB. Vaccine Breakthrough Infections with SARS-CoV-2 Variants. N Engl J Med. 2021; 384(23):2212–18. https://doi.org/10.1056/NEJMoa2105000
  16. Planas D, Veyer D, Baidaliuk A, Staropoli I, Guivel-Benhassine F, Rajah MM, Planchais C, Porrot F, Robillard N, Puech J, Prot M, Gallais F, Gantner P, Velay A, Le Guen J, Kassis-Chikhani N, Edriss D, Belec L, Seve A, Courtellemont L, Péré H, Hocqueloux L, Fafi-Kremer S, Prazuck T, Mouquet H, Bruel T, Simon-Lorière E, Rey FA, Schwartz O. Reduced sensitivity of SARS-CoV-2 variant Delta to antibody neutralization. Nature. 2021; 596(7871):276–80. https://doi.org/10.1038/s41586-021-03777-9
  17. Williams AH, Zhan CG. Fast Prediction of Binding Affinities of the SARS-CoV-2 Spike Protein Mutant N501Y (UK Variant) with ACE2 and Miniprotein Drug Candidates. J Phys Chem B. 2021; 125(17):4330–36. https://doi.org/10.1021/acs.jpcb.1c00869
  18. Aljindan RY, Al-Subaie AM, Al-Ohali AI, Kumar DT, Doss CGP, Kamaraj B. Investigation of nonsynonymous mutations in the spike protein of SARS-CoV-2 and its interaction with the ACE2 receptor by molecular docking and MM/GBSA approach. Comput Biol Med. 2021; 135:104654. https://doi.org/10.1016/j.compbiomed.2021.104654
  19. Supasa P, Zhou D, Dejnirattisai W, Liu C, Mentzer AJ, Ginn HM, Zhao Y, Duyvesteyn HME, Nutalai R, et al. Reduced neutralization of SARS-CoV-2 B.1.1.7 variant by convalescent and vaccine sera. Cell. 2021; 184(8):2201–11. https://doi.org/10.1016/j.cell.2021.02.033
  20. Hoffmann M, Kleine-Weber H, Schroeder S, Krüger N, Herrler T, Erichsen S, Schiergens TS, Herrler G, Wu NH, Nitsche A, Müller MA, Drosten C, Pöhlmann S. SARS-CoV-2 Cell Entry Depends on ACE2 and TMPRSS2 and Is Blocked by a Clinically Proven Protease Inhibitor. Cell. 2020; 181(2):271–80. https://doi.org/10.1016/j.cell.2020.02.052
  21. Trezza A, Iovinelli D, Santucci A, Prischi F, Spiga O. An integrated Miniprotein Drug Candidates. J Phys Chem B. 2021; 125(17):4330–36.