AN ASSESSMENT OF THE INTERACTION BETWEEN INSECT BRAIN PROTEIN AND NONSTRUCTURAL PROTEIN OF CORONAVIRUS USING IN-SILICO ANALYSES

an active role in the replication of the virus in the host. Molecular docking methodology plays an important role in predicting interaction between the insect proteins with non-structural protein (nsp) of coronavirus. The results predict good bonding affinity with possible interaction as hydrogen bond and salt bridge between antibacterial protein and nsp of CoViD-19 indicating as afuture alternative medications. GRAMM-X docking GAPDH


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necessary for viral replication whereas nsp7 forms a hexadecamer with nsp8 (8 subunits of each) that may participate in viral replication by acting as a primase. Alternatively, it may synthesize substantially longer products than oligonucleotide primers Or may be required to activate the RNA-synthesizing activity of Polymerase (Uniprot, Decemebr 2019). A biomolecule targeting these two proteins may help in inhibiting their replication mechanism which in turn may stop viral infections.
Insect proteins isolated from brain tissue lysate of cockroach have indicated control on the growth of drug-resistant bacteria like MRSA and MRSS. They are also found to show the effect on pathogens like S.typhi, P.aeruginosa, K.pneumoniae, etc (Sagar and Jayaprada, 2015). Interaction of this protein with nsp4 and nsp7 can give a ray of hope for controlling replication of SARS-CoV-2 in host cells. Protein docking is the task of calculating the 3D structure of a protein complex from its unbound or model-built subunits. Although proteins are intrinsically flexible, many protein docking algorithms begin by assuming that the proteins are rigid and they use geometric hashing (Bachar et al., 1993) or fast Fourier transform (FFT) correlation techniques (Katchalski-Katzir et al., 1992) to find a relatively small number of putative docking orientations which may be refined and re-scored using more sophisticated techniques. In recent years, several protein docking programs have been made available with web servers like ClusPro (Comeau et al., 2004), GRAMM-X (Tovchigrechko and Vakser, 2006) and ZDOCK (Chen et al., 2003). Docking reports obtained from these servers are always in .pdb format which can beanalyzed in detail using different servers like PDBsum, pyMol (Laskowski et al., 2018), LIGPLOT (McDonals and Thornton, 1994), HBPLUS (Wallace and Laskowski, 1995), etc.

Material and Method:-
Protein modeling: 7 Antibacterial proteins were isolated from the brain tissue lysate of cockroach (Siddharth and Jayaprada, 2015). Of these, Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and Transferrin were considered for docking study with nsp4 and nsp7. For molecular docking amino acid sequence was procured from UniProt (Uniprot-Consortium, 2011) database (accession no. 343965965 and 372292427) and considered for structural homology modeling in SWISS MODEL online portal (Aartjan et al.,2010). The structure of nsp4 and nsp7 was directly procured from the SWISS MODEL repository database of SARS-CoV-2 (Swiss Model repository, 2019).

Protein-Protein docking:
The protein structures were uploaded in GRAMM-X and allowed for Docking. The results of protein interaction obtained between virus protein nsp7, nsp4 with antibacterial protein transferrin and GAPDH respectively were analyzed using EMBL-EBI's PDBsum sever.

Binding energy prediction:
HAWKDOCK server was used to identify the best binding pose for protein-protein in docking based on binding free energy score. The more the negative value more is the interaction and complex suitable to bind. The .pdb model of GRAMM-X docked protein-protein interaction was uploaded for the calculation of binding energy based on Molecular Mechanics/Generalized Born Surface Area (MM/GBSA).

Results:-
Single structure model of nsp4 and three models of nsp7 were obtained which were labelled as nsp7.1,nsp7.       3 -4680.22 The binding energy defines the affinities and stability of the bond between protein and ligand. Complexes providing the lowest binding energy per area square of interaction tends to show maximum binding efficiency. With the above data, we can conclude that transferrin shows its best interaction with the third model of nsp7 and GAPDH shows its best interaction with the first model of nsp7. Interaction of GAPDH and Transferrin with nsp4 is also significant.

Discussion:-
The increasing havoc of human infection by a coronavirus has become a hot subject of research for scientists worldwide. Best control measures that have been concluded by WHO are to take precautions for the spread of pandemic like covering your mouth while coughing and sneezing, sanitization of hand and infected areas, etc. However, this doesn't help in curing the infected patients. Human infected by SAR-CoV-2 needs to be registered with strong antiviral chemicals. Coronavirus attaches host by spike protein which gets cleaved by host protease allowing the entry of virus by endocytosis or by direct fusion of the virus with host membrane. The genomic of the virus thus entered in host cell gets translated and proteases of itself cleaves this protein leading to the development of multiple nsp (Fehr and Perlman, 2015). Zinc inhibits RNA synthesis of coronavirus. A more than 50% reduction of overall RNA-synthesis was observed at a Zn2+ concentration of 50 µM, while less than 5% activity remained at a Zn2+ concentration of 500 µM (Aartjan et al.,2010). 3CLP is an essential viral protein for the viral replication cycle, and as a result, becomes an attractive target for anti-SARS drug development (Yang et

Conclusion:-
GRAMM-X docking shows promising interaction of antibacterial protein GAPDH with nsp4 and all three models of nsp7 of SAR-CoV-2. Transferrin an iron carrier protein, also showed interactions with nsp4 and all models of nsp7. Docking results showed no disulfide bonds. The obtained salt bridge and hydrogen bond interactions give us scope to consider the antibacterial protein as posible sources of drugs that can controlling replication of coronavirus. Acknowledgement:-

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We thank SWISS MODEL portal for easy provision of protein structure database of SAR-CoV-2 which helped us with easy docking with our protein.