Small molecule peptide for inhibiting growth of staphylococcus, analogue of small molecule peptide and application of small molecule peptide

Abstract

The invention relates to the field of medicinal chemistry, and relates to a small molecule peptide for inhibiting growth of staphylococcus, and an analogue and application thereof. The method comprises the following steps: finding a rhodococcus genome from an NCBI (National Center of Biotechnology Information) microbial genome database, and predicting a non-ribosome peptide structure synthesized by a non-ribosome peptide synthetase (NRPS) sequence through software such as AntiSMASH and Prism; the method comprises the following steps: classifying all NRPS biosynthetic gene clusters (BGCs) through Bigscape to obtain a gene cluster family (GCFs); the method comprises the following steps: predicting the selectivity of a GCFs substrate and screening a polypeptide with antibacterial potential through Phammapper; a small molecule peptide (compound I) is obtained through chemical synthesis, and the small molecule peptide is found to have an inhibition effect on staphylococcus epidermidis and staphylococcus pseudointermedius; afterwards, alanine is used for replacing glutamine to increase hydrophobicity of glutamine (compound II), the antibacterial effect of the compound II is obviously enhanced, and the compound II has the potential of developing novel antibacterial drugs and biopesticides.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of pharmaceutical chemistry, and relates to a small molecule peptide for inhibiting growth of staphylococcus and an analog and application thereof. BACKGROUND

[0002] Microorganisms, especially bacteria and fungi, have played a crucial role in the discovery and development of many drugs used in human medicine. In recent years, there has been growing interest in exploring bacteria as a potential source of new drug candidates. Nonribosomal Peptide Synthetase (NRPS) is synthesized by specific nonribosomal polypeptide synthesis proteins, which can assemble a large number of structurally and functionally diverse peptides, many of which have bioactivity that can be used for therapeutic applications, and there are currently more than 20 drugs on the market, such as antibacterial agents (penicillin, vancomycin), antitumor compounds (bleomycin) and immunosuppressants (cyclosporine). Red globulin is a kind of actinobacteria, and the content of NRPS in its secondary metabolites is rich. However, it is very difficult to obtain colonies, cultivate colonies under conditions allowing production of compounds, and isolate enough natural products for characterization and bioactivity testing. Recent studies have shown that these difficulties can be completely avoided by predicting NRPS gene clusters and their structures through bioinformatics software (such as AntiSMASH, Prism), screening small molecule peptides with antibacterial potential through Bigscape and Phammapper, and then directly producing them through chemical synthesis, which can screen and extract biosynthetic gene clusters from large-scale genomic data, saving time and economic cost.

[0003] The present application collects red globulin genomes in the NCBI microbial genome database, predicts possible molecular structures of nonribosomal peptides synthesized by NRPS in the genomes using bioinformatics software, screens small molecule peptides with potential activity, and then obtains these small molecule peptides through chemical synthesis. One of the small molecule peptides was found to have an inhibitory effect on the growth of staphylococcus, and then it was modified to enhance hydrophobicity, and the inhibitory effect was found to be significantly enhanced. SUMMARY

[0004] To accelerate the mining of natural products and enrich the molecular library of natural products, the present application uses software to predict possible molecular structures of nonribosomal peptides synthesized by NRPS in red globulin, and then obtains these small molecule peptides through chemical synthesis. One of the small molecule peptides (compound one) was found to have an inhibitory effect on the growth of staphylococcus, and then alanine was used to replace glutamine to increase its hydrophobicity, obtaining an analog (compound two) with significantly enhanced inhibitory effect.

[0005] A small molecule peptide for inhibiting growth of staphylococcus, with a chemical formula of C 54 H96 N 10 O 18 S1 (compound one), a structural formula as follows:

[0006] Compound one The chemical formula of the analog is C 52 H 93 N9O 17 S1 (compound two), a structural formula as follows:

[0007] Compound two The small molecule peptide or the analog can be applied to preparing an anti-staphylococcus product, which is an antibacterial drug or a biological pesticide.

[0008] The small molecule peptide has inhibitory effect on the growth of S. epidermidis and S. pseudintermedius. The analog also has inhibitory effect, and the inhibitory effect is better.

[0009] The beneficial effects of the present application are at least: The present application obtains a small molecule peptide and an analog thereof by mining the genome of Rhodococcus in the NCBI microbial genome database. The antibacterial experiment on the two compounds shows that they have good inhibitory activity on S. pseudintermedius, S. epidermidis and the like. Therefore, the compounds have the potential to become a lead compound of antibacterial drugs, and have good development prospects. BRIEF DESCRIPTION OF DRAWINGS

[0010] Figure 1 is the positive ion mass spectrum of the antibacterial compound one of the present application; Figure 2 is the positive ion mass spectrum of the antibacterial compound two of the present application; Figure 3 is the HPLC spectrum of the antibacterial compound one of the present application; Figure 4 is the HPLC spectrum of the antibacterial compound two of the present application. DETAILED DESCRIPTION

[0011] The present application is further described in combination with the drawings and examples. The small molecule peptide of the present application can be directly prepared by full chemical synthesis of polypeptide technology according to its structural formula. The liquid nutrient broth medium components used in the examples are as follows: peptone 1%, beef extract 1%, NaCl 0.5%, glucose 1%, pH 7.0, and the percentage content is mass percentage content.

[0012] In this embodiment, the compound one is: Compound one was prepared by solid-phase chemical synthesis method by Genview Biotech (Shanghai) Co., Ltd., and the purity was greater than 95%. High performance liquid chromatography (HPLC) and electrospray ionization mass spectrometry (ESI-MS) were used to detect the purity and accurate molecular weight of the antibacterial peptide, respectively.

[0013] Compound two in this embodiment is: Compound two was prepared by solid-phase chemical synthesis method by Genview Biotech (Shanghai) Co., Ltd., and the purity was greater than 95%. High performance liquid chromatography (HPLC) and electrospray ionization mass spectrometry (ESI-MS) were used to detect the purity and accurate molecular weight of the antibacterial peptide, respectively.

[0014] (I) Structure analysis of the compound The antibacterial compound of the present application is a white solid, easily soluble in dimethyl sulfoxide, methanol, and slightly soluble in water.

[0015] Compound one: ESI-MS m / e 1191.7 ([M+H] + ).

[0016] 1 H NMR (400 MHz, DMSO) δ 8.20 – 7.69 (m, 9H), 7.26 (s, 1H), 6.78 (s,1H), 4.46 (q, J = 7.1 Hz, 1H), 4.27 (ddd, J = 30.9, 12.6, 5.9 Hz, 7H), 4.15 –4.07 (m, 1H), 3.65 – 3.48 (m, 8H), 2.31 – 2.21 (m, 2H), 2.17 – 2.09 (m, 4H),2.03 – 1.89 (m, 2H), 1.78 (dt, J = 14.1, 7.0 Hz, 2H), 1.59 (dq, J = 13.6, 6.1Hz, 3H), 1.51 – 1.42 (m, 8H), 1.24 (s, 22H), 0.92 – 0.73 (m, 21H).

[0017] Compound two: ESI-MS m / e 1132.55 ([M+H]+).

[0018] 1H NMR (400 MHz, DMSO) δ 8.22 - 7.70 (m, 9H), 4.38 - 4.14 (m, 8H), 3.65 - 3.51 (m, 6H), 2.83 - 2.72 (m, 2H), 2.45 - 2.31 (m, 2H), 2.25 (dt, J = 9.8, 7.2 Hz, 2H), 2.13 (dd, J = 8.1, 6.4 Hz, 2H), 1.66 - 1.55 (m, 3H), 1.47 (t, J = 7.2 Hz, 8H), 1.31 - 1.21 (m, 25H), 0.85 (ddd, J = 17.4, 6.5, 2.5 Hz, 21H).

[0019] Antibacterial activity detection 1. Test bacteria Staphylococcus pseudintermedius; Staphylococcus epidermidis.

[0020] 2. Determination of antibacterial activity The bacteria were cultured in liquid medium at 30 ℃ on a shaker to obtain a bacterial solution, which was then diluted to a concentration of about 10 6 CFU / ml. 90 μl of liquid nutrient broth medium and 10 μl of compound were added to a 96-well plate to give a concentration of 32-128 μg / ml. After incubation in a 30 ℃ incubator for 16 h, the optical density (OD 600) at 600 nm was recorded using a multifunctional enzyme marker, and the minimum inhibitory concentration (MIC) was calculated. DMSO was used as a negative control, and each group was repeated three times. The MIC was defined as the minimum compound concentration that completely inhibited the growth of the culture, i.e., the inhibition rate was >90%.

[0021] Table 1. Antibacterial activity of compounds Staphylococcus epidermidis Pseudointermedius Compound one 128 μg / ml 128 μg / ml Compound two 48 μg / ml 48 μg / ml Experimental conclusion: The antibacterial compounds of the present application have good inhibitory activity against Staphylococcus pseudintermedius and Staphylococcus epidermidis.

[0022] The content of the present application is not limited to the specific embodiments listed, and any equivalent transformation of the technical solutions of the present application made by a person of ordinary skill in the art by reading the specification of the present application is covered by the claims of the present application.

Claims

1. A small molecule peptide that inhibits growth of Staphylococcus, characterized in that: The small molecule peptide has a chemical formula of C 54 H 96 N 10 O 18 S1, and a structural formula of 。 2. The analog of the small modular peptide of claim 1, characterized in that: The chemical formula of the analog is C 52 H 93 N9O 17 S1, and the structural formula is: 。 3. Use of the small molecule peptide of claim 1 or the analogue of claim 2 in the preparation of an anti-staphylococcal growth product.

4. Use according to claim 3, characterized in that, The staphylococcus includes Staphylococcus epidermidis, Staphylococcus intermedius.

5. Use according to claim 3, characterized in that, The product is a medicine or a biopesticide.

6. A bacteriostatic agent characterized in that, The small molecule peptide of claim 1 and / or the analogue of claim 2 are contained.

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