Background

SARS-CoV-2 has become a menace to the humanity and it imposed unprecedented epidemic condition. Great efforts were carried out by the scientists to develop potent vaccines like Astrazeneca/Oxford [1], Johnson & Johnson [2], Moderna [3], Pfizer/BionTech [4], Sinopharm, Sinovac [5], and COVISHIELD [6], having the potential to curb human mortality. The virus (a positive sense RNA virus with a genome of ~ 30 kb) has several types of vertebrate hosts including humans and transmission occurs through direct contact or aerosols [7, 8]. Like all animal viruses, their proteins hijack the cellular machineries to complete life cycle. These proteins are of great interest to the scientists to develop specific drug(s) or vaccine schemes against them. Search and trial of potential inhibitory drugs such as Remdesivir, Lopinavir-Ritonaviris were on the way but they were proven ineffective to prevent patient death [9,10,11]. The present work is based on the fact that most of the viral non-structural proteins (NSPs) which include enzymes remain structurally and chemically conserved as they have to interact with human proteins to carry out same biochemical processes within cell. SARS-CoV-2 genome encodes 16 non-structural proteins (NSPs), involved in genome replication and transcription [12, 13]. Nsp1 is a transcriptional, translational inhibitor and evades host immune system [14,15,16]. Nsp2 is involved in viral replication, disrupts host cell environment and, along with Nsp3, form proteases [12, 13]. Nsp4 interacts with Nsp3 to mediate viral replication [12, 13]. Main protease(Mpro) or NSP5 is essential for viral replication [7, 8, 12, 13]. Nsp6 generate autophagosomes that assemble replicase proteins [12, 13]. Nsp7, Nsp8 and Nsp12 form RNA polymerase complex [17, 18]. NSP9 replicase is dimeric and involved in viral RNA synthesis [7, 8, 12, 13, 19]. Nsp10 stimulate Nsp14 and Nsp16 which are methyl transferases [14, 20]. The function of Nsp11 is yet to be deciphered [12, 13]. Nsp13 together with Nsp12 exert helicase activity and is involved in capping of viral RNA [21]. Nsp14 has exoribonuclease and N7-methyltransferase activity [22]. Coronavirus endoribonuclease (NSP15/EndoU) is a hexameric protein that preferentially recognizes and cleaves RNA [7, 8, 12, 13, 23] and EndoU also evades host mediated viral double-stranded RNA recognition. Nsp16 has methyltransferase activity and complexes with Nsp10 [7, 8, 12, 13, 24].

In the present study, 11 PDB entries (7K3N, 6WEY, 6M03, 7JLT, 6W4B, 6ZCT, 6M71, 7NIO, 5C8S, 6VWW and 7BQ7) [25,26,27,28,29,30,31,32,33,34,35] representing twelve non-structural proteins and their complexes of SARS-CoV-2, i.e., NSP1, NSP3,NSP5, NSP7-8 complex, NSP9, NSP10, NSP7-8–12 complex, NSP13, NSP14, NSP15 and NSP16-10 complex respectively have been analyzed using Drug ReposeER web server program (http://27.126.156.175/drreposed/) [36] for their possible binding sites [37] to all drugs available in drug bank. Only the NSPs having 3D structures available in PDB, have been considered in the study as tertiary structures have utmost requirement to find 3D drug binding interfaces. The drug binding interfaces showed congruence with the known drug binding motifs (Additional file 1: S1, Additional file 2: S2, Additional file 3: S4, Additional file 4: S4, Additional file 5: S5, Additional file 6: S6, Additional file 7: S7, Additional file 8: S8, Additional file 9: S9, Additional file 10: S10 and Additional file 11: S11) .

Results and discussion

DrReposER predicted numerous potential 3D-drug binding motifs of both left (L) and right (R) superpositions for 7K3N, 6WEY, 6M03, 7JLT, 6W4B, 6ZCT, 6M71, 7NIO, 5C8S, 6VWW and 7BQ7 (Additional file 1: S1, Additional file 2: S2, Additional file 3: S4, Additional file 4: S4, Additional file 5: S5, Additional file 6: S6, Additional file 7: S7, Additional file 8: S8, Additional file 9: S9, Additional file 10: S10 and Additional file 11: S11). Known drugs that bind these motifs bind either human, bacterial or viral proteins. Results after analyzing the 3D structures of the target molecules and complexes were further filtered for anti-viral drugs. From the hit results, 14 anti-viral drugs i.e., Amphetamine (Drug bank ID-DB00182), Amprenavir (Drug bank ID-DB00701), Atazanavir (Drug bank ID-DB01072), Darunavir (Drug bank ID-DB01264), Grazoprevir (Drug bank ID-DB11575), Indinavir (Drug bank ID-DB00224), Lopinavir (Drug bank ID-DB01601), Nelfinavir (Drug bank ID-DB00220), Nevirapine (Drug bank ID-DB00238), Ribavirin (Drug bank ID-DB00811), Rimantadine (Drug bank ID-DB00478), Ritonavir (Drug bank ID-DB00503), Saquinavir (Drug bank ID-DB01232), and Tipranavir (Drug bank ID-DB00932) were selected for having unique 3D-drug binding motifs (Tables 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 and 11). The findings showed that several anti-viral drugs had binding interfaces on a single protein or protein complexes and moreover, each anti-viral drug had one to several binding motifs (Tables 12 and 13).

Table 1 Possible binding sites of NSP1 against known anti-viral drugs
Full size table
Table 2 Possible binding sites of NSP3 against known anti-viral drugs
Full size table
Table 3 Possible binding sites of NSP5 against known anti-viral drugs.
Full size table
Table 4 Possible binding sites of NSP7-NSP8 against known anti-viral drugs
Full size table
Table 5 Possible binding sites of NSP9 against known anti-viral drugs
Full size table
Table 6 Possible binding sites of NSP10 against known anti-viral drugs
Full size table
Table 7 Possible binding sites of NSP7-NSP8-NSP12 complex against known anti-viral drugs
Full size table
Table 8 Possible binding sites of NSP13 against known anti-viral drugs
Full size table
Table 9 Possible binding sites of NSP14 against known anti-viral drugs
Full size table
Table 10 Possible binding sites of NSP15 against known anti-viral drugs
Full size table
Table 11 Possible binding sites of NSP16-NSP10 complex against known anti-viral drugs
Full size table
Table 12 Comparison of drug binding motifs of analyzed NSPs for antiviral drugs
Full size table
Table 13 Comparison of NSPs binding of the drugs analyzed
Full size table

Amphetamine (DB00182) targeted only a single binding interface on Nsp5 (6M03) (Tables 3, 12, 13). Amprenavir (DB00701) targeted four binding motifs on Nsp3 (6WEY), three motifs onNsp1 (7K3N), Nsp7-8-12 complex (6M71), Nsp13 (7NIO) and Nsp14 (5C8S), and two binding motifs on Nsp7-8 complex (7JLT), Nsp15 (6VWW) and Nsp16-10 complex (7BQ7) (Tables 2, 1, 7, 8, 9, 4, 10, 11, 12, Figs. 1, 23, 4, 5, 6, 7, 8, 9, 10 and 11). Atazanavir (DB01072) targeted three motifs on Nsp16-10 complex (7BQ7), two motifs on Nsp10 (6ZCT) and single motif each on Nsp1, Nsp7-8-12, Nsp13, Nsp14 and Nsp15 (Tables 11, 6, 12). Darunavir (DB01264) is the most promising drug as it targeted the greatest number of binding motifs and targeted every molecule except Nsp9. It targeted ten motifs on Nsp1 (7K3N), seven motifs on Nsp14 (5C8S), six motifs on Nsp3 (6WEY), five motifs on Nsp15 (6VWW) and Nsp16-10 complex (7BQ7), four motifs on Nsp7-8-12 complex (6M71), three motifs on Nsp10 (6ZCT), two motifs each on Nsp5 (6M03) and Nsp13 (7NIO), respectively and a single motif on Nsp7-8 complex (Tables 1, 9, 2, 10, 11, 7, 6, 3, 8, 4, 12, Figs. 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 and 11). Grazoprevir (DB11575) targeted two motifs, on Nsp10 (6ZCT) and two on Nsp16-10 complex (7BQ7) and single motif each on Nsp9 and Nsp14 (Tables 6, 11, 5, 9, 12). Indinavir (DB00224) significantly targeted three motifs, each on Nsp13 (7NIO) and Nsp15 (6VWW) (Tables 8, 10, 12). Lopinavir significantly targeted three motifs on Nsp15 and 2 motifs each on Nsp13 and Nsp14 (Tables 10, 8, 9). Nelfinavir targeted two interfaces on Nsp1 and Nsp7-8–12 complexes (Tables 1, 7). On the other hand, Nevirapine targeted only a single motif on Nsp5 (Table 3). Rimantadine (DB00478) significantly targeted five binding interfaces on Nsp14 (5C8S), three binding motifs each on Nsp5 (6M03) and Nsp9 (6W4B), and two motifs on Nsp3 (6WEY), Nsp13 (7NIO), Nsp16-10 (7BQ7) and a single motif on Nsp1, Nsp7-8 and Nsp7-8-12 complex (Tables 9, 3, 5, 2, 8, 11, 1, 4, 7, 12, Figs. 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 and 11). Ritonavir targeted two motifs on Nsp16-10 complex (Table 11). Saquinavir (DB01232) targeted four motifs on Nsp16-10 complex (7BQ7), three interfaces each on Nsp7-8–12 (6M71) and Nsp15 (6VWW), two motifs on Nsp1 and Nsp14 (5C8S) and a single motif on Nsp3, Nsp7-8, Nsp10 and Nsp13 (Tables 11, 7, 10, 1, 9, 3, 4, 6, 8, Figs. 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 and 11). Finally, Tipranavir (DB00932) targeted two binding motifs; each on Nsp3, Nsp7-8–12 complex and Nsp14 (Tables 3, 7, 9), whereas single binding interface each on Nsp1, Nsp5, Nsp9, Nsp13, Nsp15 and Nsp16-10 (Table 12).

Fig. 1
figure 1

3D-binding interfaces of NSP1with Amprenavir, Darunavir, Rimantadine &Saquinavir. a–c Binding motifs of Amprenavir. d All the binding motifs of Amprenavir. e–n Binding interfaces of Darunavir. o All the binding motifs of Darunavir. p, q Rimantadine binding motif. r, s Saquinavir binding motifs and t All the motifs on NSP1. Numbers indicate the motif forming amino acids. Three letter codes of amino acids have been mentioned

Full size image
Fig. 2
figure 2

3D-binding interfaces of NSP3 with Amprenavir, Darunavir, Rimantadine &Saquinavir. a–d Binding motifs of Amprenavir. e All the binding motifs of Amprenavir together. f–k Binding interfaces of Darunavir. L Combined binding motifs of Darunavir. m, n Rimantadine binding motifs. o All motifs of RIM. p, q Saquinavir binding motif. Numbers indicate the motif forming amino acids. Three letter codes of amino acids have been mentioned

Full size image
Fig. 3
figure 3

3D-binding interfaces of NSP5 with Darunavir&Rimantadine. a–c Binding motifs of Rimantadine. d All the binding motifs of RIM on NSP5. e, f Binding interfaces of Darunavir. g All the binding motifs of Darunavir. Numbers indicate the motif forming amino acids. Three letter codes of amino acids have been mentioned

Full size image
Fig. 4
figure 4

3D-binding interfaces of NSP7-8 complex with Amprenavir, Darunavir, Rimantadine &Saquinavir. a, b. Binding motifs of Amprenavir. c All the binding motifs of Amprenavir together. d, e Binding interfaces of Darunavir. f, g Rimantadine binding motifs. h, i Saquinavir binding motif. Numbers indicate the motif forming amino acids. Three letter codes of amino acids have been mentioned

Full size image
Fig. 5
figure 5

3D-binding interfaces of NSP9 with Rimantadine. a–c Three binding motifs of Rimantadine. d All the binding motifs of RIM together. Numbers indicate the motif forming amino acids. Three letter codes of amino acids have been mentioned

Full size image
Fig. 6
figure 6

3D-binding interfaces of NSP10with Darunavir&Saquinavir. a–c. Binding motifs of Darunavir. d Combined binding motifs of Darunavir. e, f Saquinavir binding motif. Numbers indicate the motif forming amino acids. Three letter codes of amino acids have been mentioned

Full size image
Fig. 7
figure 7

3D-binding interfaces of NSP7-8–12 complex with Amprenavir, Darunavir, Rimantadine &Saquinavir. a–c Binding motifs of Amprenavir. d All the binding motifs of Amprenavir together. e–h Binding interfaces of Darunavir. i Combined binding motifs of Darunavir. j, k Rimantadine binding motifs. l–n Saquinavir binding motifs. o Combined motifs of ROC. Numbers indicate the motif forming amino acids. Three letter codes of amino acids have been mentioned

Full size image
Fig. 8
figure 8

3D-binding interfaces of NSP13 with Amprenavir, Darunavir, Rimantadine &Saquinavir. a–c Binding motifs of Amprenavir. d All the binding motifs of Amprenavir together. e, f. Binding interfaces of Darunavir. g Combined binding motifs of Darunavir. h, i. Rimantadine binding motifs. j All motifs of RIM. k, l Saquinavir binding motif. Numbers indicate the motif forming amino acids. Three letter codes of amino acids have been mentioned

Full size image
Fig. 9
figure 9

3D-binding interfaces of NSP14 with Amprenavir, Darunavir, Rimantadine &Saquinavir. a–c Binding motifs of Amprenavir. d All the binding motifs of Amprenavir together. e–k.Binding interfaces of Darunavir. l Combined binding motifs of Darunavir. m–q Rimantadine binding motifs. r All motifs of RIM. s, t Saquinavir binding motifs. u All the Saquinavir motifs together. Numbers indicate the motif forming amino acids. Three letter codes of amino acids have been mentioned

Full size image
Fig. 10
figure 10

3D-binding interfaces of NSP15 with Amprenavir, Darunavir&Saquinavir. a, b Binding motifs of Amprenavir. c All the binding motifs of Amprenavir together. d–h Binding interfaces of Darunavir. i Combined binding motifs of Darunavir. j–l Saquinavir binding motifs. m All the ROC binding interfaces. Numbers indicate the motif forming amino acids. Three letter codes of amino acids have been mentioned

Full size image
Fig. 11
figure 11

3D-binding interfaces of NSP16-10 complex with Amprenavir, Darunavir, Rimantadine &Saquinavir. a, b Binding motifs of Amprenavir. c All the binding motifs of Amprenavir together. d–h Binding interfaces of Darunavir. i Combined binding motifs of Darunavir. j, k Rimantadine binding motifs. l All motifs of RIM. m–p Saquinavir binding motifs. q All the Saquinavir binding interfaces. Numbers indicate the motif forming amino acids. Three letter codes of amino acids have been mentioned

Full size image

All the binding results were further compiled and analyzed. Results revealed that Darunavir (DB01264) had 45 unique binding sites and targeted 10 SARS-CoV-2 PDB entries or 10 NSPs (Tables 12, 13). The Lowest Root Mean Square Deviation (RMSD) value of Darunavir among all the target molecules was 0.54 Å for Nsp16-10 complex and maximum number of residues involved in interaction was 27 (Tables 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 and 11). Significant binding interfaces were again targeted by Amprenavir (DB00701) and Saquinavir (DB01232) with 22 and 18 (Tables 12, 13), respectively. The two drugs had eight and nine binding partners, respectively (Tables 12, 13). The lowest RMSDs for them were 0.54 Å and 0.52 Å and maximum residues involved in drug-target binding were 28 and 31, respectively (Tables 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 and 11). Additionally, Rimantadine (DB00478) had 20 drug binding motifs that targeted nine binding partners (Tables 12, 13) with the lowest RMSD value of 0.67 Å and maximum number of residues involved in binding were 10 (Tables 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 and 11). Again, Tipranavir (DB00932) and Indinavir (DB00224) both showed 12 binding motifs for nine and eight binding partners, respectively (Tables 12, 13). Lowest RMSD values for these two drugs were 0.53 Å and 0.72 Å and maximum number of residues involved in binding were 27 and 24, respectively (Tables 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 and 11).

Results showed that Darunavir, Amprenavir, Rimantadine, Saquinavir, Tipranavir and Indinavir were more effective in targeting the twelve SARS-CoV-2 proteins and their complexes (Tables 12, 13). Darunavir is a nonpeptidic benzenesulfonamide inhibitor that targets active site of HIV-1 protease [38, 39]. Amprenavir is a hydroxyethylamine sulfonamide derivative that inhibits HIV-1 protease [40, 41]. Rimantadine is an alkylamine that specifically targets Influenza A virus M2 protein [42,43,44]. Saquinavir is a L-asparagine derivative that acts as HIV-1 protease inhibitor [45, 46]. Tipranavir is a sulfonamide that acts as HIV-1 protease inhibitor [47]. Moreover, Indinavir is a piperazinecarboxamide having HIV-1 protease inhibitory activity [48, 49]. The drug binding interfaces determined in the present study is very much significant as the analysis considered previously known potent binding information between specific drugs and target proteins that were again supported by very low RMSD values of the motifs such as 0.54 Å for both Darunavir and Amprenavir, 0.52 Å for Saquinavir and 0.67 Å for Rimantadine (Tables 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 and 11). RMSD values well below 1.0 was indicative of presence of similar drug binding structures or motifs as in active site of HIV-1 protease or M2 of Influenza A and these results emphasized that the selected drugs would effectively target those similar interfaces found on different NSPs of SARS-Cov2 to inhibit them. Furthermore, considering the produced results, it has been proposed that combination of Darunavir, Amprenavir and Rimantadine could effectively target and inhibit all the NSPs that were studied. Darunavir targeted all NSPs except Nsp9, whereas Amprenavir targeted all except Nsp5, Nsp9 and Nsp10 and interestingly Rimantadine complementarily and significantly targeted Nsp5 and Nsp9, which are two key enzymes (Tables 12, 13). However, it has been reported that Darunavir was unable to protect HIV patients from SARS-Cov2 infection who were under Darunavir treatment [50]. Though, the claim has to be experimentally proven. In such cases, if Darunavir fails to prevent infection, then another potent inhibitor Saquinavir, having similar target profiles, could be used in combination along with Amprenavir and Rimantadine, in replacement of Darunavir (Tables 12, 13, 14).

Table 14 Active site residues of the analyzed SARS-CoV2 enzymes and the inhibitory drug binding motifs
Full size table

Among the twelve proteins studied, eight were key enzymes involved in viral replication, transcription and life cycle processes. Hence, the study was further extended to provide insight whether the binding motifs of the selected drugs were significant in inhibiting these enzymes possibly by intercepting active sites of those enzymes. Active sites of enzymes are surface regions that are highly conserved and involved in catalysis or substrate binding. In this study, active sites of SARS-CoV-2 enzymes were predicted by a web server, GASS-WEB (http://gass.unifei.edu.br/) that uses Genetic Active Site Search based on genetic algorithms [51]. Active site residues and the drug binding interfaces of the four drugs viz. Amprenavir (478), Darunavir (017), Rimantadine (RIM) and Saquinavir (ROC) were presented in surface topography presentations of each of the enzymes and were analyzed for their inhibitory association. Results revealed that active site residues of the papain- like protease NSP3 were in close association with drug binding motifs of Amprenavir (270D, 252G, 253 V, 335I, 300 V, 304 V, 287L), Darunavir (252G, 227I, 253 V, 335I, 286 V, 297L, 287L), Rimantadine (337G, 333A, 315S, 281 V) and Saquinavir (252G, 253 V, 335I) (Fig. 12, Table 14). Active sites of protease NSP5 were closely apposed to Darunavir (133 N, 194A, 195G, 200I, 109G, 293P) and Rimantadine (254S, 255A, 251G) binding residues (Fig. 13, Table 14). NSP9 active sites were exclusively targeted by Rimantadine (108 V, 109A, 111 V, 106S, 105G) (Fig. 14, Table 14). RNA polymerase NSP12 active sites were targeted by Amprenavir (166 V, 760D, 203G, 204 V, 201I), Darunavir (53 V, 106I, 119I, 203G, 204 V, 201I), Rimantadine (774G, 771A, 772S) and Saquinavir (623D, 817 T, 820 V, 203G, 204 V, 201I) (Fig. 15, Table 14). The helicase NSP13 active residues were targeted by Amprenavir (195I, 151I, 226 V, 258I), Darunavir (195I, 226 V, 258I), Rimantadine (1A, 3G, 523S, 527G) and Saquinavir (258I) (Fig. 16, Table 14). Exoribonuclease NSP14 active sites were closely apposed to Amprenavir (31I, 14I, 87I, 412P), Darunavir (389 V, 26A, 78R, 390D, 108 V, 152L, 118 V, 120 V), Rimantadine (32A, 34G, 35G, 33S) and Saquinavir (31I, 14I, 84R) binding residues (Fig. 17, Table 14). On the other hand, endonuclease NSP15 active sites were targeted by Amprenavir (276 V, 156 V), Darunavir (80I, 23 V, 212I, 156 V, 3L, 86I), and Saquinavir (119P, 80I, 156 V) (Fig. 18, Table 14). Finally, methyltransferase NSP16 active site residues were targeted by Amprenavir (71A, 70G), Darunavir (21 V, 22D, 26A, 71A, 290I, 121A, 200S), Rimantadine (32A, 33S, 34G, 199A, 197 V, 200S) and Saquinavir (71A, 70G) (Fig. 19, Table 14). Close association of drug binding motifs with the active sites indicated that these would interfere with catalytic activity and substrate binding of the enzymes.

Fig. 12
figure 12

Active site residues & drug binding motifs of NSP3. a, b Two different surfaces showing drug binding motifs in close association with active site residues of the enzyme. Here Anprenavir, Darunavir and Saquinavir targeted active site residue VAL253 in a pocket. 478-Amprenavir; 017-Darunavir; RIM Rimantadine, ROC Saquinavir

Full size image
Fig. 13
figure 13

Active site residues & drug binding motifs of NSP5. a Position of active site residues within and near a pocket. b 017 & RIM targeted residues closely associated with that pocket which accounts for inhibition of active site functioning. 478-Amprenavir; 017-Darunavir; RIM Rimantadine, ROC Saquinavir

Full size image
Fig. 14
figure 14

Active site residues & drug binding motifs of NSP9. a Surface view showing position of active site residues. b Only Rimantadine showed numerous inhibitory binding. 105G and 106S active residues were targeted by RIM. RIM Rimantadine

Full size image
Fig. 15
figure 15

Active site residues & drug binding motifs of NSP12. a Position of active site residues. b, c Different surfaces showing 478, 017, RIM and ROC binding interfaces or residues. 478-Amprenavir; 017-Darunavir; RIM Rimantadine, ROC Saquinavir

Full size image
Fig. 16
figure 16

Active site residues & drug binding motifs of NSP13. a, b Position of active site residues and drug binding motifs in surface presentation. 478-Amprenavir; 017-Darunavir; RIM Rimantadine, ROC Saquinavir

Full size image
Fig. 17
figure 17

Active site residues & drug binding motifs of NSP14. a, b Position of active site residues and drug binding interfaces in surface presentation. 478-Amprenavir; 017-Darunavir; RIM Rimantadine, ROC Saquinavir

Full size image
Fig. 18
figure 18

Active site residues & drug binding motifs of NSP15. a–c Different surface projections of NSP15 showing positions of active residues and drug binding motifs. 478-Amprenavir; 017-Darunavir; RIM Rimantadine, ROC Saquinavir

Full size image
Fig. 19
figure 19

Active site residues & drug binding motifs of NSP16. a–c Different surface projections showing inhibitory association of drug binding motifs with active site residues of the enzyme. 478-Amprenavir; 017-Darunavir; RIM Rimantadine, ROC Saquinavir

Full size image

Previously, several drug repurposing analysis were performed by several groups to find potential drug inhibitors like sirolimus, dactinomycin, mercaptopurine, melatonin, toremifene, emodin, zotatifin, ternatin-4, hydroxychloroquine, clemastine, Atazanavir, remdesivir, efavirenz, Ritonavir, dolutegravir, carfilzomib, cyclosporine A, azithromycin, favipiravir, Ribavirin, galidesivir and many others against SARS-CoV-2 proteins but their efficacy is questionable in treating and curing COVID-19 patients [52,53,54,55,56,57].

Conclusion

The findings strongly suggested that among the fourteen anti-viral drugs predicted and analyzed, six drugs significantly targeted twelve SARS-Cov2 non structural proteins and specifically the key enzymes. Considering the binding parameters it can be concluded that combination of Darunavir (DB01264), Amprenavir(DB00701) and Rimantadine(DB00478) or Saquinavir (DB01232), Amprenavir (DB00701) and Rimantadine (DB00478) or all the four drugs together can potentially bind and inhibit the cellular activities of these proteins that are essential for viral replication and life cycle. Using anti-viral drug has great advantage in that these have specific target and less or no similar binding partners like Rimantadine had no other binding partners other than SARS-Cov-2 NSPs (Tables 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 and 11). Finally, these predicted drug combinations must be clinically tested to save thousands of lives in the vicinity of limited effectiveness of developed vaccines [58, 59].

Methods

Key resources table

Resource

Source

Identifier

Analyzed data

 SARS-CoV-2 NSP1 3D-structure

[25]

PDB ID: 7K3N

 SARS-CoV-2 NSP3 3D-structure

[26]

PDB ID: 6WEY

 SARS-CoV-2 NSP5 3D-structure

[27]

PDB ID: 6M03

 SARS-CoV-2 NSP7-8 complex 3D-structure

[28]

PDB ID: 7JLT

 SARS-CoV-2 NSP9 3D-structure

[29]

PDB ID: 6W4B

 SARS-CoV-2 NSP10 3D-structure

[30]

PDB ID: 6ZCT

 SARS-CoV-2 NSP7-8-12 complex 3D-structure

[31]

PDB ID: 6M71

 SARS-CoV-2 NSP13 3D-structure

[32]

PDB ID: 7NIO

 SARS-CoV-2 NSP14 3D-structure

[33]

PDB ID: 5C8S

 SARS-CoV-2 NSP15 3D-structure

[34]

PDB ID: 6VWW

 SARS-CoV-2 NSP16-10 complex 3D-structure

[35]

PDB ID: 7BQ7

Web server

 DrReposER

[37]

http://27.126.156.175/drreposed/

 GASS-WEB

[51]

http://gass.unifei.edu.br/

DrReposERhas been used to find binding interfaces or 3D-motifs of target proteins (PDB ID: 7K3N, 6WEY, 6M03, 7JLT, 6W4B, 6ZCT, 6M71, 7NIO, 5C8S, 6VWW and 7BQ7) for all possible drugs. The program uses SPRITE and ASSAM web servers to find amino acid side chains. Drug ReposER compares structurally similar side chain arrangements from PDB repository and assign hit results for different drug targets in the query PDB ID [37].

GASS-WEB has been used to predict active sites of SARS-CoV-2 enzymes (NSP3, NSP5, NSP9, NSP12, NSP13, NSP14, NSP15 and NSP16) considered in this study. It uses genetic algorithms to find active sites of enzymes that are meant for catalytic activity or substrate binding [51].