key: cord-0706685-o3w7779t authors: Khater, Ibrahim; Nassar, Aaya title: SARS-CoV-2 variant surge and vaccine breakthrough infection: A computational analysis date: 2022-02-04 journal: Inform Med Unlocked DOI: 10.1016/j.imu.2022.100873 sha: 64f81566ce62e6080cb53f3414f59e45805562e2 doc_id: 706685 cord_uid: o3w7779t Coronavirus Delta variant was first detected in India in October of 2020, and it led to a massive second wave of COVID-19 cases in the country. Since then, the highly infectious Delta strain has been spreading globally. The Delta variant and its sub-lineages showed an increased infection rate with a reduced effect of the potential antibody neutralization. The current work is a modeled computational analysis of the mutated receptor-binding domain (RBD) of the SARS-CoV-2 B.1.617 lineage binding with ACE2 and GRP78 to understand the increased strain transmissibility. The cell-surface Glucose Regulated Protein 78 (GRP78) attached to the mutated ACE2-SARS-CoV-2 Spike RBD complex is modeled. The results showed that GRP78 β-substrate-binding domain weakly binds to the wild-type RBD combined with angiotensin-converting enzyme 2 (ACE2) within the SARS-CoV-2 Spike RBD-ACE2 complex. Both GRP78 and ACE2 bind approximately in the same region on the wild-type SARS-CoV-2 Spike RBD surface. On the other hand, GRP78 strongly binds to the mutated SARS-CoV-2 Spike RBD in the RBD-ACE2 complex through the α-substrate-binding domain instead of β-substrate-binding domain in a different region from that of ACE2. The current findings suggest that blocking the main ACE2 pathway may not prevent the interactions between GRP78 and the mutated SARS-CoV-2 Spike RBD, which might introduce an additional avenue into the virus invasion for the host cell if the ACE2 pathway is blocked by the neutralized antibodies. Hence, the peptide satpdb10668 has been proposed as a potential inhibitor of SARS-CoV-2 attachment and virus invasion into the host cell. Viruses are changing constantly due to mutations and new variants are likely to emerge. Occasionally, new variants appear, and they may persist or disappear. Some variants appear to spread more quickly and easily than others. With coronavirus 2019, mutations of the SARS-CoV-2 have been spreading widely over the world since the fall of 2020, several new variants have appeared. The primary variants of concern showed common mutations of the SARS-CoV-2 Spike protein, mostly on the S1 unit, resulting in a higher transmissibility rate compared to the wild type of the SARS-CoV-2. The SARS-CoV-2 variants of the global concern include the Alpha (B. show resistance to neutralization by some anti-NTD and anti-RBD monoclonal antibodies that may have increased the infectivity or reduced the potential for plasma neutralization, making these lineages spread faster than other variants (6), (7), (8), (9), (10) . In the SARS-CoV-2 Spike RBD, mutations emerge on the angiotensin-converting enzyme 2 (ACE2) binding surface's periphery, implying that the virus accumulates mutations there to weaken antibody recognition while retaining ACE2 binding (11) , (12) , (13) . The cell surface Glucose Regulated Protein 78 (GRP78), also known as BiP or HSPA5, is the primary chaperone of the endoplasmic reticulum (ER), which can help with protein folding, protein synthesis, and maturation, and plays an essential role in the regulation of ER stress signaling (14) , (15) , (16) , (17) , (18) . GRP78 plays a key role as a multiple-function cell surface receptor interacting with many proteins (19) , (20) , (21) , (22) . In addition to its role in the cell proliferation, invasion, and metastasis of cancer cells, GRP78 is sensitive to virus recognition through its substrate-binding domain (SBD) and participates in the assembly of its envelope protein (23), (24) , (25) , (26) . The beta substrate-binding domain (β-SBD) of the GRP78 has been determined to be the docking location for the C480-C488 region within the SARS-CoV-2 Spike RBD (27) , (28) . The GRP78 is predicted to bind to the SARS-CoV-2 Spike nearby the ACE2 host cell receptor (29) , (30) . The proposed theory is supported by the work of Carlos et al. and Elfiky et al. which found GRP78 binding to ACE2 require the SBD binding domain. They proposed that the substratebinding domain is significant for attachment (31) , (32) . The current work aims to calculate the free binding energy of GRP78 and ACE2 with the mutated SARS-CoV-2 Spike RBD Delta lineages (B1.617.1) and (B.1.617.2) to understand the strains' increased transmissibility and to investigate potential antiviral inhibitors. The research proposes that GRP78 establishes an alternative cell entry pathway for the mutated SARS-CoV-2 Spike when the ACE2 pathway is being blocked by the neutralized antibodies. The RBD in the wild-type and the mutant types of the ACE2-SARS-CoV-2 Spike RBD system was simulated and, with the presence of these mutant variants of the viral Spike, the GRP78 binding with the ACE2-SARS-CoV-2 Spike RBD complex (ACE2-RBD) was modeled. J o u r n a l P r e -p r o o f The SARS-CoV-2 Spike protein mutations (L452R, E484K) found in the SARS-CoV-2 Delta (B1.617.1) lineage and the SARS-CoV-2 Spike protein mutations (K417N, L452R, E484K) found in the SARS-CoV-2 Delta plus (B1.617.2) lineage (33) were prepared using the PyMOL molecular graphics system (34) . The GROMACS-2019 software package (35) and CHARMM36 force field (36) were used to perform the Molecular Dynamic Simulation (MDS) on the wild-type SARS-CoV-2 Spike RBD, as well as the mutated Delta and Delta plus RBD. CHARMM-GUI generated protein topology and parameter files (37) , (38) , (39) . The system was solvated with TIP3P water (40) and the complexes were neutralized by using the Monte-Carlo ion-placing method to add sufficient amounts of K+ and Cl− ions (40) . The system was energy-minimized for 5000 steps with the steepest descent algorithm (41) and equilibrated for 125 ps at a constant number of molecules, volume, and temperature (NVT) before running the simulation. The MDS was then performed for 50 ns at constant temperature (310 K), pressure (1 atm), and the number of molecules (NPT ensemble). The Root Mean Square Deviation (RMSD) of the protein atom backbone and the radius of gyration (Rg) were calculated (42) . The average Root Mean Square Fluctuation (RMSF) was plotted as a function of residue number. The ClusPro server is a tool for protein-protein docking (43) , (44) . The ClusPro 2.0 webserver was used to dock GRP78 (PDB ID: 5E84: A) to the SARS-CoV-2 three types complexes: the wild-type (PDB ID: 6M0J) Spike RBD-ACE2 complex, hereinafter referred to as (WT ACE2-RBD), the Delta mutated RBD-ACE2 complex, hereinafter referred to as (Delta Mut J o u r n a l P r e -p r o o f ACE2-RBD), and the Delta plus mutated RBD-ACE2 complex, hereinafter referred to as (Delta plus Mut ACE2-RBD). The binding energy of the complexes was calculated using MM/GBSA of the HawkDock server (45) , (46) . The PDBePISA server of the Protein Data Bank in Europe was used to analyze the interacting interfaces (47). The Structurally Annotated Therapeutic Peptides Database (SATPdb) was used to find biologically active peptides (48) . The peptides were docked to the Delta mutant RBD and the Delta plus mutated RBD using the ClusPro 2.0 platform. The complexes' binding energy was determined using the HawkDock server's MM/GBSA. The interactions were analyzed using the PDBePISA platform. Molecular dynamics simulation was used to assess the stability of the produced complexes. In the current work, the ACE2-SARS-CoV-2 Spike RBD complex was docked using the RBD in the wild-type and the mutated Delta lineages. In addition, the mutant variants of the viral Figure 3 illustrates the complexes formed. The binding energies between GRP78 and ACE2 with RBD within the three complexes were calculated using MM/GBSA. The interacting interfaces were analyzed using the PDBePISA server by predicting the hydrogen bonds and salt bridges and Table 1 lists the interactions formed. Table 2 . GRP78 binds to the Delta and Delta plus mutant of the RBD at a completely different site from ACE2 but does not bind to ACE2 as displayed in Figure 3 . This means that the waning immunity or the neutralized antibodies are blocking the main ACE2 pathway, which in turn would not prevent the interaction between GRP78 and the Delta Mut RBD and Delta plus Mut J o u r n a l P r e -p r o o f SARS-CoV-2 has changed through time, with mutations that make a significant difference soon becoming dominant lineages. Neutralization of mutated viruses is reduced compared with the wild-type strains (51) , (52) . Recent studies focused on protection against SARS-CoV-2 variants of concern using alternative epitopes (53) , (54) The molecular docking procedure creates instantaneous interactions between peptides and proteins, which might be unstable (55) . Molecular dynamic simulations can provide information about the stability of the formed complexes' molecular interactions. The Delta Mut RBD and Delta plus Mut RBD in complex with peptides satpdb10029 and satpdb10668 were used to perform the molecular dynamics simulations. Using the RMSD approach, the stability of the complexes was evaluated for the backbone atoms concerning the original starting structures (56) . Additionally, the stability of those complexes was assessed further using the Rg plot (56) . A plot of the RMSD and Rg values over the simulation time scale is displayed in Figure 5 . The Delta Mut RBD-satpdb10029 complex did not stabilize over the simulation time, whereas the Delta Mut RBD-satpdb10668 complex was unstable between 10 and 20 nm before becoming stable. The Delta plus Mut RBD-satpdb10029 and Delta plus Mut RBD-satpdb10668 complexes were roughly stable over the simulation time. The findings suggest peptide satpdb10668 as a potential inhibitor of the mutated SARS-CoV-2 Delta and Delta plus lineages. The coronavirus disease of 2019 (COVID-19) remains a worldwide health concern due to its high infection rate. Genetic mutations of the SARS-CoV-2 Spike protein keep emerging with time and the variants keep spreading rapidly. The Delta variants and their sub-lineages are reported to have increased infection transmissibility and build waning immunity or resistance. In this study, we analyzed the binding energy of the mutated SARS-CoV-2 Spike protein of the Delta Mut RBD and Delta plus Mut RBD with GRP78, which formed an alternative pathway for the cell entry Centers for Disease Control and Preventions C. Variants of the Virus Department of Health & Human Services Covid-19: New coronavirus variant is identified in UK Implications of the Emergence of a New Variant of SARS-CoV-2, VUI-202012/01 Clinical Characterization and Genomic Analysis of Samples from COVID-19 Breakthrough Infections during the Second Wave among the Various States of India Potential Therapeutic Targets and Vaccine Development for SARS-CoV-2/COVID-19 Pandemic Management: A Review on the Recent Update. 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Neuron What is the probability of a chance prediction of a protein structure with an rmsd of 6 Å? The authors wish to acknowledge the Department of Biophysics, Faculty of Science, Cairo University for facilitating the work. J o u r n a l P r e -p r o o f Table 1 . The binding energy of GRP78 and ACE2 with RBD was calculated using MM/GBSA, and the interactions are analyzed by using PDBePISA.WT ACE2-RBD-GRP78 complex ACE2-RBD interactions (-87. 5 GLN24 ASP30 HIS34 GLU35 GLU37 ASP38 TYR41 GLN42 TYR83 LYS353 LYS417 GLY446 TYR449 ASN487 TYR489 GLN493 GLY496 GLN498 THR500 GLY502 TYR505 --ASP56 GLN57 ASN82 LYS152 GLU155 YR160 LYS552 ASP556 ASN559 GLU599 GLU560 GLU562 SER563 TYR566 ASP574 GLU576 GLU602 ARG346 ARG355 ARG357 ASP389 THR393 ASN394 ASP428 ARG466 SER469 THR470 LEU517 LEU518 HIS519 LYS46 ARG49 GLU51 GLU155 LYS552 ASP556 GLU560 LU562 ASP574 GLU576 ARG346 ARG357 LYS386 ASP389 ARG466 GLU516 HIS519 J o u r n a l P r e -p r o o f