Revealing anti-viral potential of Bio-active therapeutics targeting SARS-CoV2- polymerase (RdRp) in combating COVID-19: Molecular Investigation on Indian traditional medicines

Spread of severe acute respiratory syndrome coronavirus (SARS-CoV-2) made a historic transition between December 2019 to March 2020. In the present scenario SARS-CoV-2 as becomes a major burden on public health and economic stability of societies around the globe. From the substantial evidences gained from the pandemic of SARS-CoV-2 and MERS-CoV (Middle East respiratory syndrome coronavirus), scientists and clinicians strongly believes that these pathogenic viruses share common homology of some biologically active enzymes which includes RNA-dependent RNA polymerase (RdRP), 3-chymotrypsin-like protease (3CL pro ), papain-like protease (PL pro ) etc. RdRP relatively grabs higher level of clinical importance in comparison with other enzyme target. Indian system of traditional medicine pioneering the therapy towards infectious disease since several centuries. In view of this potential therapeutic leads from some of the Indian medicines along with standard drug favipiravir subjected to docking investigation targeting SARS-CoV-2- RNA dependent RNA polymerase (RdRp). Residual proximity analysis reveals 18 out of 28 compounds reveals potential binding affinity of about 100% with the target amino acid residue (618 ASP, 760 ASP,761 ASP), 7 out of 28 reveals 75% binding efficacy and 3 out of 28 reveals 25% binding efficacy with that of the target residue. Hence further clinical validation may be warranted with proper in-vitro and in-vivo studies prior to the clinical recommendation in treating COVID-19 patient’s.

some of the Indian medicines along with standard drug favipiravir subjected to docking investigation targeting SARS-CoV-2-RNA dependent RNA polymerase (RdRp). Residual proximity analysis reveals 18 out of 28 compounds reveals potential binding affinity of about 100% with the target amino acid residue (618 ASP, 760 ASP,761 ASP), 7 out of 28 reveals 75% binding efficacy and 3 out of 28 reveals 25% binding efficacy with that of the target residue. Hence further clinical validation may be warranted with proper in-vitro and in-vivo studies prior to the clinical recommendation in treating COVID-19 patient's.

1.Introduction
Pandemic spread of COVID-19 caused by severe acute respiratory syndrome coronavirus (SARS-CoV-2) has now shifted across the boundaries, authoritative data issued by WHO on 29 March 2020 intensifies the severity of viral invasion that prevails in more than 190 countries.
Fatality index attaining greater hikes on hourly basis, according to recent demography global mortality due to COVID-19 was found to be 29,957 deaths 1 . Countries like Italy (92,472 cases), Spain (72,248 cases), Germany (52,547) and Iran (35,408 cases) seems severely affected with striking invasion of disease.
As per the epidemiological investigations is concern the spectrum of COVID-19 in India majorly due to imported infection rather than the acquired local transmission. India reported with 3 positive cases on 1 March 2020, whereas it has reached 979 on 29 March 2020 these data's alarms the need of preventive measure that has to be focused with higher priority.
China become a global role model in advocating the traditional chinese medicine (TCM) along with conventional therapy for managing COVID-19 which was evidenced by initiation of several multi centric randomized trials for validating the safety and efficacy of TCM in patients reported with COVID-19 [2][3][4][5] . National health commission of china framed a policy guideline for ensures the efficacy of TCM in treated patients 6,7 . There are several enzymes involved in the pathogenesis of SARS-CoV-2 of which the enzyme RNA dependent RNA polymerase (RdRP) that exist only on the viral genome relatively grabs higher level of therapeutic importance due to its versatile action in mediating nonstructural protein (nsp 12) essential for viral replication 14,15 . Drugs that inhibits this RdRP activity may have expected to halt the replication of viral genome and thereby control disease progression 16,17 .Recent clinical outcomes reveals improved efficacy of favipiravir (RdRP) inhibitor with estimated recovery rate of 71.43% in treated cases 18 .
Still now there is no proper documented evidence supporting the efficacy of prescribed indian medicines on targeting SARS-CoV-2-RNA dependent RNA polymerase (RdRp). Hence present study aimed at exploring RdRp enzyme inhibition potential of both the formulations (Nilavembu Kudineer and Kaba sura kudineer) using predictive molecular docking assay.

2.1.Protein-ligand docking
Molecular docking investigation was performed using Auto Dock version 4 which predicts interaction binding affinity between selected therapeutic lead with that of the protein target (SARS-CoV-2 virus spike RNA dependent RNA polymerase-RdRp).

2.2.Protein preparation
Three dimensional (3D) structure of SARS-CoV-2 virus spike RNA dependent RNA polymerase with protein data bank (PDB)-6NUR ( Figure 1) retrieved from Research Collaboratory for Structural Bioinformatics (RCSB). Protein structure were cleaned by removing the existing lead components, water molecules cleaved, Gasteiger charges computed with inclusion of polar hydrogens, merging of non-polar and rotatable bonds were defined using Auto Dock 4 19 .

2.3.Active site prediciton on the target protein
Active amino acids involved in mediating the enzyme activity was predicted using ramachandran plot indicating localization of the residues on the A chain of the target enzyme. Prediction by MolProbity server and also through literature survey. As shown in Figure 2.

2.5.Docking simulations
3D componential structure of lead molecules and protein were docked using AutoDock analytical tool version 4. Affinity (grid) maps of ×× Å grid points and 0.375 Å spacing were generated using the Autogrid program. AutoDock parameter set-and distance-dependent dielectric functions were used in the calculation of the van der Waals and the electrostatic terms, respectively. Docking simulations were performed using the programmed algorithm inbuilt with pre automation in the software 20. Initial position, orientation, and torsions of the ligand molecules were set randomly. All rotatable torsions were released during docking. Each docking experiment was derived from 2 different runs that were set to terminate after a maximum of 250000 energy evaluations. The population size was set to 150. During the search, a translational step of 0.2 Å, and quaternion and torsion steps of 5 were applied.

3.1.Molecular docking analysis
Docking becomes a reliable drug discovery tools for optimizing the potential lead molecules against selective target. Docking score implicates the binding affinity between the lead and target

4.Conclusion
Emerging SARS-CoV-2 infection rates urge the need of immunonutrient that has a tendency to