Likelihood of new allosteric sites for glycolytic enzymes, phosphofructokinase (PFK), glyceraldehyde-3-phosphate dehydrogenase (GADPH) and pyruvate kinase (PK) was evaluated for bacterial, parasitic and human species. Allosteric effect of a ligand binding at a site was revealed on the basis of low-frequency normal modes via Cα-harmonic residue network model. In bacterial PFK, perturbation of the proposed allosteric site outperformed the known allosteric one, producing a high amount of stabilization or reduced dynamics, on all catalytic regions. Another proposed allosteric spot at the dimer interface in parasitic PFK exhibited major stabilization effect on catalytic regions. In parasitic GADPH, the most desired allosteric response was observed upon perturbation of its tunnel region which incorporated key residues for functional regulation. Proposed allosteric site in bacterial PK produced a satisfactory allosteric response on all catalytic regions, whereas in human and parasitic PKs, a partial inhibition was observed. Residue network model based solely on contact topology identified the ‘hub residues’ with high betweenness tracing plausible allosteric communication pathways between distant functional sites. For both bacterial PFK and PK, proposed sites accommodated hub residues twice as much as the known allosteric one. Tunnel region in parasitic GADPH with the strongest allosteric effect among species, incorporated the highest number of hub residues. These results clearly suggest one-to-one correspondence between the degree of allosteric effect and the number of hub residues in that perturbation site, which increases the likelihood of its allosteric nature.
The novel coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) still has serious negative effects on health, social life, and economics. Recently, vaccines from various companies have been urgently approved to control SARS-CoV-2 infections. However, any specific antiviral drug has not been confirmed so far for regular treatment. An important target is the main protease (Mpro), which plays a major role in replication of the virus. In this study, Gaussian and residue network models are employed to reveal two distinct potential allosteric sites on Mpro that can be evaluated as drug targets besides the active site. Then, FDA-approved drugs are docked to three distinct sites with flexible docking using AutoDock Vina to identify potential drug candidates. 14 best molecule hits for the active site of Mpro are determined. 6 of these also exhibit high docking scores for the potential allosteric regions. Full-atom molecular dynamics simulations with MM-GBSA method indicate that compounds docked to active and potential allosteric sites form stable interactions with high binding free energy (∆Gbind) values. ∆Gbind values reach -52.06 kcal/mol for the active site, -51.08 kcal/mol for the potential allosteric site 1, and -42.93 kcal/mol for the potential allosteric site 2. Energy decomposition calculations per residue elucidate key binding residues stabilizing the ligands that can further serve to design pharmacophores. This systematic and efficient computational analysis successfully determines ivermectine, diosmin and selinexor currently subjected to clinical trials, and further proposes bromocriptine, elbasvir as Mpro inhibitor candidates to be evaluated against SARS-CoV-2 infection