Target spot for virtually screening anti-monkey pox virus medicine and application of target spot

Abstract

The invention provides a target spot for virtually screening an anti-monkey pox virus drug and application of the target spot, and belongs to the technical field of virtual screening of drugs. The target spot of the invention is a protein compound formed by molecular docking of MPXV F10 and VACV NPH-I; the amino acid sequence of the MPXV F10 is as shown in SEQ ID NO. 1, and the amino acid sequence of the VACVNPH-I is as shown in SEQ ID NO. 2. According to the invention, the crystal structure of the MPXV virus core protein F10 is determined, and a plurality of anti-monkey pox virus compounds are screened by virtual screening aiming at a contact cavity between the MPXV F10 and VACV NPH-I.

Description

Technical Field

[0001] The present invention belongs to the technical field of virtual drug screening, and in particular relates to a target for virtual screening of anti-monkeypox virus drugs and an application thereof. Background Art

[0002] Monkeypox virus (MPXV) is a rare zoonosis that is transmitted through contact with sick humans or animals. Infection can cause symptoms such as fever, rash, and swollen lymph nodes, and in severe cases can even be life-threatening.

[0003] Traditional methods of preventing and treating monkeypox virus mainly rely on vaccination and drug treatment. However, the development and production of vaccines require a lot of time and resources. With the rapid development of science and technology, virtual screening technology has gradually become an important means in the field of drug research and development. Virtual screening is a compound screening computational method based on a small molecule database, which can quickly screen out active compounds with medicinal properties from tens to millions of molecules, greatly reducing the number of experimental screening compounds, shortening the research cycle, and reducing the cost of drug research and development. Through virtual screening, massive compound databases can be processed in a short time, greatly improving the efficiency of drug research and development and reducing experimental costs. Compared with traditional experimental screening methods, virtual screening can quickly predict important parameters such as the binding ability and affinity of compounds to targets, providing valuable clues and directions for subsequent experimental research.

[0004] Currently, virtual screening research on monkeypox virus is still in its infancy. Existing research still faces many challenges in terms of accurate analysis of monkeypox virus targets, optimization of virtual screening algorithms, and accuracy and reliability of screening results. For example, the structure and function of key proteins of monkeypox virus are not fully understood, which limits the accuracy of virtual screening.

[0005] Therefore, developing a virtual screening method based on special drug targets and targeting monkeypox virus is of great significance and urgent need to accelerate the research and development of drugs for the prevention and treatment of monkeypox virus. Summary of the invention

[0006] In view of this, the purpose of the present invention is to provide a target for virtual screening of anti-monkeypox virus drugs and its application, determine the protein crystal structure of MPXV virus core protein F10, and screen out a variety of anti-monkeypox virus compounds through virtual screening targeting the contact cavity between MPXV F10 and VACVNPH-I.

[0007] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0008] A target for virtual screening of anti-monkeypox virus drugs, wherein the target is a protein complex formed by molecular docking of MPXV F10 and VACV NPH-I; the amino acid sequence of the MPXV F10 is shown in SEQ ID NO.1, and the amino acid sequence of the VACVNPH-I is shown in SEQ ID NO.2.

[0009] Preferably, the steps of docking the MPXV F10 and VACV NPH-I molecules are as follows:

[0010] The MPXV F10 target protein structure was processed on the Maestro11.9 platform, including removal of water and ions, protonation, addition of missing atoms, completion of missing groups, protein energy minimization, and energy optimization. The force field was selected as OPLS3e. The Ni ions at the active site were retained during the ion removal.

[0011] Obtain the protein complex of MPXV F10 and VACV NPH-I: Select the 6RFL crystal structure, use Pymol2.1 to superimpose and retain the target protein to obtain the multimer; use the Minimization module to optimize to solve the unreasonable spatial structure contact existing in rigid docking; select OPLS3e as the force field for energy minimization, select water molecules as the solvation model, and use the steepest descent method and conjugate gradient method for optimization, with the maximum number of iterations being 5000;

[0012] The protein complex was processed using Protein Preparation Wizard, and the OPLS3 force field was used to add bond steps and hydrogen atoms to the protein structure; Prime was used to repair missing residues or atoms in the protein; PROPKA was used to check the protonation state of ionizable groups in the protein; hydrogen bonds were optimized by reorienting hydroxyl bonds, thiol groups, and amide groups;

[0013] The system was preprocessed, optimized, and minimized under the OPLS3e force field through constrained minimization, and the RMSD convergence value was Determine docking sites based on protein-protein interactions and use them as The center of mass of the box.

[0014] More preferably, the docking sites are GLU-80, ARG-14, VAL-117, LEU-82, PRO-81 and LEU-83 of the MPXV F10 protein.

[0015] The present invention also provides application of the target in screening anti-monkeypox virus drugs.

[0016] The present invention also provides a virtual screening method for anti-monkeypox virus drugs, which includes docking the compounds to be screened to the binding pockets on the target interaction interface, performing docking scoring and sorting screening, and screening out anti-monkeypox virus compounds.

[0017] Preferably, the compounds to be screened are derived from APExBIO and Selleck compound entity libraries, and the processing steps are as follows:

[0018] The modified Lipinski five-drug rule was used to eliminate the screened compounds, and the set conditions were: molecular weight <850; number of hydrogen bond donors <15; number of hydrogen bond acceptors <15; lipid-water coefficient <5; number of rotatable bonds ≤10;

[0019] The compound was protonated and converted into a minimized three-dimensional structure by energy minimization optimization using the LigPre module in the Maestro11.9 platform. In addition, tautomers were generated, and the maximum number of generated conformations was set to 32. Each ligand retained the specified chirality, and the force field was selected as OPLS3e.

[0020] Preferably, the compounds to be screened are prepared according to the default settings of the LigPre module for subsequent molecular docking.

[0021] Preferably, the processing and optimization of the virtual screening is performed by The Glide module in Maestro software was completed and the protein complex was imported to specify the appropriate position in the protein complex grid generation, and the docking sites GLU-80, ARG-14, VAL-117, LEU-82, PRO-81, and LEU-83 were determined and used as The center of mass of the box.

[0022] Preferably, the data set is screened by a standard docking method, and then the super-precision docking method is used to screen out the pre-ligands with higher scores determined by the standard docking method.

[0023] Preferably, the scoring and ranking screening includes: screening out the top-ranked compounds according to the energy scores of the docked data, and screening out anti-monkeypox virus compounds by combining the energy scores and the evaluation method of the key residues of the active site.

[0024] Compared with the prior art, the present invention has the following beneficial effects:

[0025] The present invention performs X-ray diffraction on the harvested high-quality F10 core protein crystals, obtains diffraction data, performs data analysis and processing, and finally obtains a high-resolution 3D protein structure.

[0026] Since monkeypox virus (MPXV) and vaccinia virus (VACV) are conservative, the present invention docked MPXV F10 with VACV NPH-I, and found that the binding sites of the two proteins formed a cavity, and some amino acids formed strong electrostatic interactions and hydrophobic interactions. These interactions will also play an important role in screening, and have good binding parameters at the NTP and RNA sites.

[0027] The present invention screened out several compounds by virtual screening for the site between MPXV F10 and VACV NPH-I. The above compounds have a good binding effect with the protein complex of F10 and NPH-I and a high degree of matching, and are potential active inhibitors. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 This is the information of pET-28a vector;

[0029] Figure 2 This is the SDS-PAGE electrophoresis result of MPXV F10 core protein;

[0030] Figure 3 This is a protein crystal image of MPXV F10 core protein;

[0031] Figure 4 This is the crystal X-ray diffraction pattern of MPXV F10 core protein

[0032] Figure 5 This is the crystal structure model of MPXV F10 core protein;

[0033] Figure 6 The molecular docking results of MPXV F10 core protein and VACV NPH-I protein;

[0034] Figure 7 is a virtual screening flow chart of the present invention;

[0035] Figure 8 The binding modes of different compounds with MPXV F10 core protein. DETAILED DESCRIPTION

[0036] The present invention provides a target for virtual screening of anti-monkeypox virus drugs, wherein the target is a protein complex formed by molecular docking of MPXV F10 and VACVNPH-I. In the present invention, the amino acid sequence of the MPXV F10 core protein is shown in SEQ ID NO.1; VACV NPH-I is the transcription termination factor NPH-I in the RNA polymerase complex of vaccinia virus, and its amino acid sequence is shown in SEQ ID NO.2.

[0037] The present invention also provides the application of the above target in screening anti-monkeypox virus drugs.

[0038] The present invention further provides a virtual screening method for anti-monkeypox virus drugs, which includes docking the compounds to be screened to the binding pockets on the target interaction interface, performing docking scoring and sorting screening, and screening out anti-monkeypox virus compounds.

[0039] The technical solutions provided by the present invention are described in detail below in conjunction with the embodiments, but they should not be construed as limiting the protection scope of the present invention.

[0040] Example 1

[0041] The MPXV F10 core protein was expressed and purified through molecular cloning, protein expression and purification and other related methods.

[0042] (1) Molecular cloning

[0043] The full-length gene of MPXV F10 protein was downloaded from NCBI (GenBank: WNN26160.1) and submitted to Sangon Biotech Co., Ltd. for E. coli codon optimization (as shown in SEQ ID NO.3), constructed into vector pET.28a, transformed into DH5a E. coli and sequenced. The vectors with completely correct sequencing results and no mutation sites were transformed into E. coli BL21 (DE3) cells for expression.

[0044] The specific amino acid sequence of MPXV F10 core protein is as follows:

[0045] MELVNIFLETDAGRVKFVIKNTDDVCASELINKFVELLSEYIHIDQSEFYLVVKDKDIFYFKCDRGSISIVNNEFYVFDEPLLFVKDFTKITGVEFIVTETMPCRIIPKNNHAVISVVTNHKFYNGLSL, as shown in SEQ ID NO.1.

[0046] pET-28a vector information Figure 1 shown.

[0047] The plasmid containing the MPXV F10 gene and the pET-28a vector were subjected to double restriction digestion (Ncol and Xhol) experiments, and then the two were connected at 16°C for 1h using a ligase, and then immediately incubated on ice with DH5a Escherichia coli competent cells for 30min, and then incubated on ice for 5min after a heat shock at 42°C for 90s. After the ice bath, 600μL of LB medium was added and incubated in a shaker at 37°C for 60min. After the incubation, it was spread on a solid LB medium containing 100μg / ml kanamycin and inverted in a 37°C incubator overnight. The next day, a single clone strain was picked for double restriction digestion verification, and the pET-28a containing the MPXV F10 sequence that was successfully cloned was sequenced, and the vector with completely correct sequencing results and no mutation sites was transformed.

[0048] This transformation used a heat shock method, specifically: 1 μL of the successfully sequenced vector was added to 25 μL of BL21 (DE3) Escherichia coli competent cells, immediately placed in an ice bath for 10 minutes, then heat shocked in a 42°C water bath for 90 seconds, then immediately placed on ice and continued to ice bath for 5 minutes, after the ice bath, 200 μL of LB medium was added, and incubated in a shaker at 37°C for 30 minutes. After the incubation, it was spread on solid LB medium containing 100 μg / ml kanamycin and inverted in a 37°C incubator overnight. On the second day, a monoclonal strain was picked and placed in 5 mL of liquid LB medium containing 100 μg / ml kanamycin and incubated at 37°C for 8 h. Subsequently, 600 μL of the bacterial solution was added to 400 μL of 50% glycerol and stored in a -20°C refrigerator for subsequent purification and large-scale amplification for protein expression. In addition, 200 uL of the bacterial solution was reserved as a sample before the trial expression induction, and 1 mM IPTG was added to the remaining bacterial solution and incubated at 37°C for 4 h. Subsequently, 200 μL of the bacterial solution was reserved as a sample after the trial expression induction.

[0049] All samples were centrifuged at 12,000 rpm for 1 min, and 30 μL H2O and 10 μL loading were added after discarding the supernatant. The samples were boiled at 100°C for 10 min and then analyzed by SDS-PAGE. Subsequent large-scale protein expression was performed on strains with obvious changes before and after induction.

[0050] (2) Protein expression

[0051] The E. coli containing the target protein was first cultured in 5 mL LB containing 100 μg / mL kanamycin, cultured overnight at 37°C, and transferred to 1 L liquid culture medium the next day for culture at 37°C. When the OD value at 600 nm was 0.8, 1 mM IPTG was then added to induce protein expression at 16°C overnight. Finally, it was centrifuged in batches at 4,000 rpm for 25 min, the supernatant was discarded, and the bacterial precipitate was suspended in buffer A (50 mM Tris-HCl, pH 7.5, 0.5 M NaCl, 20 mM imidazole, 5% glycerol).

[0052] (3) Protein purification

[0053] The E. coli containing the target protein was broken in turn by high-pressure homogenization and ultrasonic cell disruption machine, and centrifuged in a refrigerated centrifuge at 4°C, 15,000rpm for 1h. The supernatant was collected and incubated with Ni medium for affinity chromatography purification. After eluting with buffer B (50mM Tris-HCl, pH 7.5, 0.5M NaCl, 250mM imidazole, 5% glycerol), ion exchange chromatography was performed. The chromatography column was balanced in advance with buffer C (50mM Tris-HCl, pH 7.5, 0.05M NaCl, 5% glycerol) and D (50mM Tris-HCl, pH 7.5, 1M NaCl, 5% glycerol), and the protein-containing solution in the previous step was reduced to less than 100Mm NaCl and concentrated to 5mL, and then filtered through HiTrap TM Q HP, use NaCl concentration gradient to elute the target protein, and finally the target protein is eluted at 250mM NaCl salt concentration.

[0054] Then, gel filtration chromatography was performed to concentrate the target protein collected in the previous step to 500 μL and injected into Superdex TM 200 Increase 10 / 300GL column, and elute the target protein with buffer E (25mM Tris-HCl, pH 7.5, 150mM NaCl, 5mM DTT, 5% glycerol).

[0055] Samples were kept in each step and tested by SDS-PAGE electrophoresis to obtain the target protein with high purity and good homogeneity. Figure 2 The concentration was adjusted to 2.5 mg / mL and 5 mg / mL using a 10 kDa concentrator for crystallography experiments.

[0056] Example 2

[0057] Crystallography experiments

[0058] Finally, 5 mg / mL MPXV F10 core protein was grown by sitting drop method in a crystal incubator at 16°C. Protein crystals were obtained in a pool solution (0.2 M ammonium sulfate, 0.1 M BIS-TRIS pH 5.5, 25% w / v Polyethylene glycol 3350), as Figure 3 These crystals were stored in a reservoir solution containing 20% ​​glycerol and snap-frozen in liquid nitrogen before being shipped to the Shanghai Light Source to obtain diffraction data.

[0059] Example 3

[0060] MPXV F10 diffraction data and optimization

[0061] X-ray diffraction data were collected at the BL02U1 beamline of the Shanghai Synchrotron Radiation Facility (SSRF). Figure 4 ), and all diffraction data were processed using HKL3000 software. PDB 6RFG was used as a molecular replacement model, and the structure of the MPXV F10 virus core protein was analyzed and modified using coot and PHENIX. As shown in Table 1. After the structure was analyzed, Pymol2.1 software was used to draw the following Figure 5 shown.

[0062] Table 1 Diffraction data collection and correction

[0063]

[0064]

[0065] In Table 1:

[0066] a: Values ​​in brackets are statistics of the highest resolution shell;

[0067] b: R merge =ΣhklΣj|Ihkl-Ihkl(J)| / ΣhklΣj|Ihkl(J)|, Ihkl(J) and Ihkl represent the Jth and average reflection intensity hkl respectively;

[0068] b: R work =Σhkl||Fobs|-|Fcalc|| / |Fobs|, Fobs and Fcalc represent the observed and calculated structure factors, respectively, R free is the R factor calculated using 10% reflection as the test set;

[0069] d: The above values ​​are modified by PHENIX.

[0070] Example 4

[0071] Preparation of compound library

[0072] The compound library for this docking mainly comes from APExBIO and Selleck compound entity libraries (15325, https: / / www.apexbio.cn / screening-library.html, https: / / www.selleck.cn / ), and all compounds are protonated and energy minimized using the LigPre module in the Maestro11.9 platform. The force field selected is OPLS3e, which can perform constrained minimization.

[0073] Preparation of protein structures

[0074] The protein structure (MPXV F10 structure was obtained by analysis, and the VACVNPH-1 structure was obtained from PDB 6FRL) was processed on the Maestro11.9 platform, including removal of water and ions (retaining Ni ions in the active site), protonation, addition of missing atoms, completion of missing groups, protein energy minimization, and energy optimization. The force field was selected as OPLS3e.

[0075] Example 5

[0076] Molecular docking and virtual screening

[0077] (1) Virtual screening processing and optimization are performed by The Glide module in Maestro software was used for protein preparation. Protein Preparation Wizard module was used for protein preparation, optimization and minimization of receptors (constrained minimization using OPLS3e force field). All compounds were prepared according to the default settings of LigPre module.

[0078] When screening in the Glide module, the prepared receptors are imported to specify the appropriate positions in the receptor grid generation. The docking sites (GLU-80, ARG-14, VAL-117, LEU-82, PRO-81, LEU-83) are determined based on protein-protein interactions and used as The center of mass of the box.

[0079] First, the ligands were re-docked to confirm the feasibility of the selected docking method. Then, the data set was screened by SP (standard docking method) docking. The SP docking template is suitable for screening compounds in large quantities. Finally, the XP (super precision docking method) docking template was used to screen out the pre-ligands with higher scores determined by the SP method. The XP method is designed to provide a better correlation between good poses and scores.

[0080] In the screening process, in order to retain as many docking molecules as possible, the molecular docking binding energy scoring threshold is set to -6kcal / mol. If this condition is met, it is considered that the molecule has good affinity with the protein. In the actual screening process, although we set the threshold to -6kcal / mol, there are actually many molecules with binding energies less than -6kcal / mol, and then the potential anti-monkeypox virus compounds are further screened in combination with the reference key sites of the positive molecules. The purpose of this experiment is to screen proteins and protein inhibitors. Therefore, in order to obtain the structure of the MPXV F10 protein complex, the crystal structure (6RFL) crystal is selected and superimposed using Pymol2.1 to retain the target protein to obtain a polymer. Afterwards, the Minimization module is used for optimization to solve the unreasonable spatial structure contacts that may exist in rigid docking. The force field selected for energy minimization is OPLS3E, and the solvation model selects water molecules. The optimization method is divided into two-step steepest descent method (Steepest Descent) and conjugate gradient method (Conjugate Gradient), and the maximum number of iterations is 5000. The resulting complex model will be used to analyze the interaction interface between MPXV F10 and VAVC NPH-I.

[0081] (2) Docking results

[0082] The binding sites of the two proteins form a cavity. ARG-30 and GLU-22 of the VACV NPH-I protein are inserted into the MPXVF10 protein cavity, forming strong electrostatic interactions (ionic bonds) with GLU-80 and ARG-14 of the MPXV F10 protein cavity, which plays an important role in the binding of the two proteins. Moreover, the hydrophobic residues (PRO-20, MET-24, VAL-21) of the VACV NPH-I can also form good hydrophobic interactions with VAL-117, LEU-82, PRO-81, and LEU-83 of MPXV F10. These interactions play an important role in stabilizing the ligand, which further indicates that these amino acids will play an important role in the screening process, such as Figure 6 shown.

[0083] (3) Filter results

[0084] According to the energy scores of the docked data, the top 225 compounds were screened out, and 28 compounds were finally obtained by combining the energy scores and the evaluation method of the key residues of the active site, such as Figure 7According to the contact between the top-ranked molecules and the protein active sites, it is judged whether the compound can effectively occupy the key residues of the MPXV F10 and VAVC NPH-I binding sites to inhibit the binding of the two proteins to further evaluate and screen out potential compounds. This screening will effectively reduce the problem of possible off-target effects of the compounds, improve screening accuracy, and reduce experimental costs.

[0085] The specific screening results are shown in Table 2.

[0086] Table 2 Compound screening of MPXV F10 core protein inhibitors

[0087]

[0088]

[0089] According to the screening results, six compounds (Chicoric acid, Pseudouridine, VER155008, Nec-29, Iohexol, OroxinB) were selected for display. The compounds have a good binding effect with the MPXV F10 protein and a high degree of match (the binding energy is less than -6kcal / mol). The complex formed by the compound and the protein after docking was visualized using Pymol2.1 software to obtain the binding mode of the compound and the protein. According to the binding mode, the amino acid residues that bind to the compound and the protein pocket can be clearly seen. For example: The active site of Chicoric acid (GLU-80, THR-119, LYS-54, GLU-2, LYS-16) forms multiple hydrogen bond interactions, and the hydrogen bond distance is short and the binding ability is strong, which is conducive to the small molecule being firmly anchored in the site of the MPXVF10 protein. In addition, the benzene ring of the compound can also form a strong hydrophobic interaction with PHE-84, which also contributes to the stability of small molecules. Figure 8 (a) As shown. Pseudouridine compounds can form strong hydrogen bond interactions with ASP-79, GLU-80, LEU-82, and GLU-2 at the sites of MPXVF10 protein. The hydrogen bond distance is short and the binding ability is strong, which plays an important role in anchoring small molecules in the protein cavity, such as Figure 8 (b) As shown. The amino acids that have hydrogen bond interactions between VER155008 and MPXVF10 protein mainly include GLU-80, LYS-54, PHE-78, etc. These hydrogen bonds can effectively promote the formation of stable complexes between small molecules and proteins, such as Figure 8As shown in (c). Nec-29 has a good match with the MPXV F10 target site and can form multiple hydrogen bond interactions with the amino acids (GLU-80, VAL-4) of the protein site. It also has strong hydrophobic interactions with VAL-117, PHE-7, and PHE-84, which plays an important role in anchoring small molecules in the protein pocket. It also shows that the small molecule binds well to the protein pocket, as shown in Figure 8 (d) As shown. Iohexol can form a good match with the MPXVF10 protein pocket, such as forming multiple hydrogen bond interactions with LEU-82, VAL-4, LYS-54, and PHE-78, effectively promoting the formation of a stable complex between small molecules and proteins, such as Figure 8 (e) OroxinB can form hydrogen bond interactions with GLU-2, VAL-4, and LYS-54, and form strong hydrophobic interactions with LEU-82, PHE-84, VAL-18, and VAL-77. These interactions can effectively promote the formation of a stable complex between small molecules and MPXVF10 protein, such as Figure 8 In summary, these compounds have good performance in docking scoring and binding mode with the MPXVF10 target protein, can form stable complexes with the protein, and have high similarity in binding mode, and are potential active inhibitors.

[0090] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A method for virtual screening of targets for anti-monkeypox virus drugs, characterized in that: The target is a protein complex formed by molecular docking of MPXV F10 and VACVNPH-I; the amino acid sequence of the MPXVF10 is shown in SEQ ID NO.1, and the amino acid sequence of the VACVNPH-I is shown in SEQ ID NO.

2.

2. The target according to claim 1, characterized in that The steps of MPXVF10 and VACVNPH-I molecular docking are as follows: The target protein structure of MPXVF10 was processed on the Maestro11.9 platform, including removal of water and ions, protonation, addition of missing atoms, completion of missing groups, protein energy minimization, and energy optimization. The force field was selected as OPLS3e. The Ni ions at the active site were retained during the ion removal. Obtain the protein complex of MPXVF10 and VACVNPH-I: Select the 6RFL crystal structure, use Pymol2.1 to superimpose and retain the target protein to obtain the multimer; use the Minimization module to optimize to solve the unreasonable spatial structure contact existing in rigid docking; select OPLS3e as the force field for energy minimization, select water molecules as the solvation model, and use the steepest descent method and conjugate gradient method for optimization, with the maximum number of iterations being 5000; The protein complex was processed using Protein Preparation Wizard, and bond steps and hydrogen atoms were added to the protein structure using the OPLS3 force field; Prime was used to repair missing residues or atoms in the protein; PROPKA was used to check the protonation state of ionizable groups in the protein; hydrogen bonds were optimized by reorienting hydroxyl bonds, thiol groups, and amide groups; The system was preprocessed, optimized, and minimized under the OPLS3e force field through constrained minimization, and the RMSD convergence value was Determine docking sites based on protein-protein interactions and use them as The center of mass of the box.

3. The target according to claim 2, characterized in that The docking sites are GLU-80, ARG-14, VAL-117, LEU-82, PRO-81 and LEU-83 of the MPXV F10 protein.

4. Use of the target according to any one of claims 1 to 3 in screening anti-monkeypox virus drugs.

5. A virtual screening method for anti-monkeypox virus drugs, characterized in that: The compounds to be screened are docked to the binding pocket on the target interaction interface according to any one of claims 1 to 3, and docking scoring and sorting screening are performed to screen out anti-monkeypox virus compounds.

6. The virtual screening method according to claim 5, characterized in that: The compounds to be screened are derived from APExBIO and Selleck compound entity libraries, and the following steps are used for subsequent molecular docking: The modified Lipinski five-rule drug-like rule was used to eliminate the compounds to be screened, and the conditions were set as follows: molecular weight <850, number of hydrogen bond donors <15, number of hydrogen bond acceptors <15, lipid-water coefficient <5, number of rotatable bonds ≤10; The compound was protonated and converted into a minimized three-dimensional structure by energy minimization optimization using the LigPre module in the Maestro11.9 platform. In addition, tautomers were generated, and the maximum number of generated conformations was set to 32. Each ligand retained the specified chirality, and the force field was selected as OPLS3e.

7. The virtual screening method according to claim 5, characterized in that: The virtual screening process and optimization are performed by The Glide module in Maestro software was completed and the protein complex was imported to specify the appropriate position in the protein complex grid generation, and the docking sites GLU-80, ARG-14, VAL-117, LEU-82, PRO-81, and LEU-83 were determined and used as The center of mass of the box.

8. The virtual screening method according to claim 5, characterized in that: The data set was screened using the standard docking method, and the super-precision docking method was then used to screen out the pre-ligands with higher scores determined by the standard docking method.

9. The virtual screening method according to claim 5, characterized in that: The scoring and ranking screening includes: screening out the top-ranked compounds according to the energy scores of the docked data, and screening out anti-monkeypox virus compounds by combining the energy scores and the evaluation method of the key residues of the active site.

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