Application of a small molecule covalent inhibitor in the preparation of drugs inhibiting s-adenosylmethionine decarboxylase and its screening method

Abstract

The invention belongs to the field of bio-medicine, and in particular relates to an application of a small-molecule covalent inhibitor in preparing a medicine for inhibiting S-adenosylmethionine decarboxylase and a screening method of the small-molecule covalent inhibitor. The structural formula of the small-molecule covalent inhibitor is as shown in Description, wherein R1 represents phenyl or aniline or benzamide or an alkyl group or furan or pyrrole or pyridine and a substituted group thereof. A method for screening the small-molecule covalent inhibitor of the S-adenosylmethionine decarboxylase comprises the following steps: deleting pyruvoyl 68 residue in an S-adenosylmethionine decarboxylase crystal structure, and conducting optimizing in Rosetta software so as to obtain an optimized protein crystal structure; and conducting docking calculation on the optimized protein crystal structure and a searched small-molecule structure by virtue of AutoDockVina software, and screening calculating results in accordance with a grading order so as to obtain the small-molecule covalent inhibitor.

Description

technical field

[0001] The invention belongs to the field of biomedicine, and in particular relates to the application of a small molecule covalent inhibitor in the preparation of drugs inhibiting S-adenosylmethionine decarboxylase and a screening method thereof. Background technique

[0002] For a long time, in clinical and basic research, small molecular compounds have been widely used as drugs or regulatory molecules to treat diseases or study biological systems. These small molecular compounds can bind to target proteins in a covalent or non-covalent manner. However, covalent compounds were usually avoided in past rational drug screening and design because early researchers worried that covalent compounds would have large "off-target" effects and cause large side effects. Because of this, the past computer-aided screening methods were basically optimized for the docking of non-covalent compounds for screening, design, optimization and evaluation of the binding of non-co...

Examples

Embodiment 1

[0041] 1) Based on the crystal structure of S-adenosylmethionine decarboxylase (AdoMetDC), the Pyr68 residue in the crystal structure of AdoMetDC with PDB ID 3DZ5 was deleted and optimized in Rosetta to obtain the optimized SCAR protein Crystal structure;

[0042] 2) The small molecule compound library used for docking comes from the docking database with ZINC (http: / / zinc.docking.org). The search conditions for small molecules are: ①SMILES expression: CONN, and select the substructure of "substructure" to search Molecular structure containing the above groups; ② molecular weight greater than 200 and less than 400; ③ xLogP ≤ 3.5; ④ rotatable bond greater than or equal to 3 and less than or equal to 9; ⑤ hydrogen bond donor greater than or equal to 2 and less than or equal to 10; ⑥ hydrogen bond The receptor is greater than or equal to 2 and less than or equal to 10. After the server returns the search results, the structure of the corresponding small molecule is obtained;

[...

Embodiment 2

[0047] Utilizing the method of the patent (ZL201410530672.3), we have carried out AdoMetDC activity evaluation, spectroscopic detection described step 4) the activity inhibitory effect of the small molecule covalent inhibitor of Example 1 on AdoMetDC, small molecules mixed with AdoMetDC in After co-incubating at 37 degrees for 30 minutes, add the substrate and react for 5 minutes before testing. DMSO is used as the solvent control, and AdoMetDC inhibitor MGBG (mitoguanidine hydrazone) is used as the positive control. The results are shown in figure 1 , this small molecule covalent inhibitor has better inhibitory ability than MGBG;

Embodiment 3

[0049] The inhibitory effect of the small molecule in Example 1 on AdoMetDC was verified by detecting the consumption of the substrate AdoMet by using high performance liquid chromatography (HPLC). The instrument used for HPLC is Waters e2695, the column is Waters C18 reverse phase column (5um, 4.6x250mm), the column temperature is 30 degrees, the mobile phase is 0.01mol / L ammonium formate (pH 3.5):methanol=97:3, and the flow rate is 0.5mL / min , detection wavelength 254nm. Before loading the sample, add 9 times the volume of methanol to stop the reaction, centrifuge at 12000g for 10 minutes to remove the denatured protein, then take the supernatant sample for analysis, the results are shown in figure 2 ,Depend on figure 2 It can be seen that after mixing and incubating the small molecule and AdoMetDC at 37 degrees for 30 minutes, the reaction was terminated after adding the substrate for 0 minutes, 5 minutes or 10 minutes, and the consumption of the substrate was detected. ...

technical field

[0001] The invention belongs to the field of biomedicine, and in particular relates to the application of a small molecule covalent inhibitor in the preparation of drugs inhibiting S-adenosylmethionine decarboxylase and a screening method thereof. Background technique

[0002] For a long time, in clinical and basic research, small molecular compounds have been widely used as drugs or regulatory molecules to treat diseases or study biological systems. These small molecular compounds can bind to target proteins in a covalent or non-covalent manner. However, covalent compounds were usually avoided in past rational drug screening and design because early researchers worried that covalent compounds would have large "off-target" effects and cause large side effects. Because of this, the past computer-aided screening methods were basically optimized for the docking of non-covalent compounds for screening, design, optimization and evaluation of the binding of non-co...

Examples

Embodiment 1

[0041] 1) Based on the crystal structure of S-adenosylmethionine decarboxylase (AdoMetDC), the Pyr68 residue in the crystal structure of AdoMetDC with PDB ID 3DZ5 was deleted and optimized in Rosetta to obtain the optimized SCAR protein Crystal structure;

[0042] 2) The small molecule compound library used for docking comes from the docking database with ZINC (http: / / zinc.docking.org). The search conditions for small molecules are: ①SMILES expression: CONN, and select the substructure of "substructure" to search Molecular structure containing the above groups; ② molecular weight greater than 200 and less than 400; ③ xLogP ≤ 3.5; ④ rotatable bond greater than or equal to 3 and less than or equal to 9; ⑤ hydrogen bond donor greater than or equal to 2 and less than or equal to 10; ⑥ hydrogen bond The receptor is greater than or equal to 2 and less than or equal to 10. After the server returns the search results, the structure of the corresponding small molecule is obtained;

[...

Embodiment 2

[0047] Utilizing the method of the patent (ZL201410530672.3), we have carried out AdoMetDC activity evaluation, spectroscopic detection described step 4) the activity inhibitory effect of the small molecule covalent inhibitor of Example 1 on AdoMetDC, small molecules mixed with AdoMetDC in After co-incubating at 37 degrees for 30 minutes, add the substrate and react for 5 minutes before testing. DMSO is used as the solvent control, and AdoMetDC inhibitor MGBG (mitoguanidine hydrazone) is used as the positive control. The results are shown in figure 1 , this small molecule covalent inhibitor has better inhibitory ability than MGBG;

Embodiment 3

[0049] The inhibitory effect of the small molecule in Example 1 on AdoMetDC was verified by detecting the consumption of the substrate AdoMet by using high performance liquid chromatography (HPLC). The instrument used for HPLC is Waters e2695, the column is Waters C18 reverse phase column (5um, 4.6x250mm), the column temperature is 30 degrees, the mobile phase is 0.01mol / L ammonium formate (pH 3.5):methanol=97:3, and the flow rate is 0.5mL / min , detection wavelength 254nm. Before loading the sample, add 9 times the volume of methanol to stop the reaction, centrifuge at 12000g for 10 minutes to remove the denatured protein, then take the supernatant sample for analysis, the results are shown in figure 2 ,Depend on figure 2 It can be seen that after mixing and incubating the small molecule and AdoMetDC at 37 degrees for 30 minutes, the reaction was terminated after adding the substrate for 0 minutes, 5 minutes or 10 minutes, and the consumption of the substrate was detected. ...

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