CN-122017224-A - Method for screening antibacterial compounds by high-flux targeted phytophthora elicitin based on DHE fluorescence intensity and application

Abstract

The invention discloses a method for screening antibacterial compounds based on high-flux targeted phytophthora elicitin with DHE fluorescence intensity and application thereof, belonging to the fields of biotechnology and agricultural disease control. According to the method, the active compound capable of specifically targeting elicitin-sterol binding sites is rapidly screened in high throughput by detecting the change of the fluorescence intensity of a reaction system by incubating the compound to be detected, elicitin protein and the fluorescence sterol DHE in a PBS buffer system and taking the fluorescence value after the combination of the DHE and elicitin as a control and the fluorescence intensity of the compound to be detected is reduced after the competitive combination of elicitin. The screened compound has a wide antibacterial spectrum and good inhibitory activity on various important plant pathogenic fungi, and shows great potential as a lead compound of a broad-spectrum plant source bactericide.

Inventors

• DOU DAOLONG
• PEI YONG
• YIN ZHIYUAN
• WANG HUI
• GUO YINING

Assignees

• 南京农业大学三亚研究院

Dates

Publication Date

20260512

Application Date

20260414

Claims (4)

1. 1. The application of an active compound of a specific targeted elicitin-sterol binding site in competitive inhibition of elicitin binding to sterols is characterized in that the active compound is at least one of glabridin, carvacrol, aloin, pentofyllin A, licoisoflavone A, emodin anthrone, scutellarin methyl ester, aloesin, 6-feruloyl spinosin, punica granatum, aloe-emodin-8-O-beta-D-glucoside, xanthohumol, huang Caogan B, (E/Z) -demethoxycurcumin, theaflavin-3-gallate, sophocarpidone G and 3' -nornobiletin.
2. 2. The use according to claim 1, wherein the active compound is glabridin, penthorum chinense pursh a or licoisoflavone a.
3. 3. Use of the active compounds glabridin, penthorside a or licoisoflavone a as claimed in claim 1 or 2 in any of the following: (1) Use in inhibiting the growth of oomycetes or fungi by competitively inhibiting elicitin-sterol binding; (2) The application in preparing pesticide products for inhibiting the growth of oomycetes or fungi; The oomycete is at least one selected from phytophthora capsici, phytophthora, downy mildew, phytophthora nicotianae, pythium ultimum, phytophthora sojae and phytophthora infestans, and the fungi is at least one selected from sclerotinia sclerotiorum, fusarium graminearum and gray mold.
4. 4. Use of the active compound glabridin according to claim 1 or 2, in any of the following: (1) Application in preventing and treating soybean root rot; (2) The application in preparing pesticide products for preventing and treating soybean root rot; (3) Use in promoting soybean growth; (4) Use in the preparation of a pesticide product for promoting soybean growth.

Description

Method for screening antibacterial compounds by high-flux targeted phytophthora elicitin based on DHE fluorescence intensity and application Technical Field The invention belongs to the field of biotechnology and agricultural disease control, and particularly relates to a method for screening antibacterial compounds based on high-flux targeted phytophthora elicitin with DHE fluorescence intensity and application thereof. Background Phytophthora (Phytophthora) is a very damaging plant pathogen, has approximately 200 species, and can infect various important grain and economic crops such as potatoes, soybeans, tomatoes, peppers, cucumbers and the like to cause destructive diseases. Phytophthora is similar to fungi in morphology, but belongs to the phylum oomycete of the kingdom of the tricholoma and the cell wall of the Phytophthora contains cellulose instead of chitin, and zoospores have characteristics of double flagella and the like which are obviously different from fungi. The taxonomic status determines that the traditional fungus bactericide is basically ineffective to phytophthora, and brings great challenges to disease prevention and control. More seriously, phytophthora has the characteristics of complex genetic diversity, strong genome plasticity, rapid variation in natural environment and the like, and is extremely easy to generate strains with drug resistance and capability of overcoming host resistance. Pathogenic bacteria generate oospores through sexual reproduction to overwinter, so that an initial infection source of diseases is increased, meanwhile, the zoospores are spread by means of rainwater or wind power, the spreading distance can reach more than 10 km, and the diseases are caused to spread rapidly. The rapid adaptation of phytophthora makes it possible to rapidly overcome the genetic resistance of chemical fungicides and hosts, resulting in limited effectiveness of existing control measures. Sterols act as essential lipids in eukaryotes, not only structural components of cell membranes, but also as signal molecules in plant-pathogen interactions to regulate the growth, development and reproduction of organisms. Oomycetes such as phytophthora are typical sterol auxotrophic pathogens, which cannot synthesize sterols themselves and must be obtained from host plants to maintain their vegetative growth, vegetative propagation and sexual propagation. Phytophthora acquires the host sterols by secreting a class of effector proteins called elicitin. Elicitin is a small molecular weight, cysteine-rich secreted protein whose hydrophobic cavity in its structure is capable of binding sterol molecules with high affinity, thereby helping phytophthora to "predate" sterols from the host cell membrane. Elicitin is used as a key effector protein for obtaining sterol by phytophthora, plays an irreplaceable role in the pathogenic process of pathogenic bacteria, and is an ideal target for developing novel anti-oomycete medicines. However, in the prior art, elicitin is known to bind sterols, but there is a lack of a method to screen for compounds targeting elicitin-sterol binding sites directly, rapidly, and with high throughput. Traditional screening methods typically rely on complex biological activity assays, such as pathogen growth inhibition assays or spore germination inhibition assays, which are cumbersome to operate, long-term, low-throughput, and difficult to distinguish whether a compound acts directly on the elicitin target or acts bacteriostasis through other non-specific pathways. Therefore, there is an urgent need in the art to develop a compound screening method based on elicitin target points, which is simple to operate, has high sensitivity, and is suitable for high-throughput screening, so as to provide an effective technical platform for developing novel anti-oomycete drugs. Disclosure of Invention The invention aims to provide a method for screening antibacterial compounds by high-flux targeted phytophthora elicitin based on DHE fluorescence intensity and application thereof, The aim of the invention can be achieved by the following technical scheme: In a first aspect, the invention claims the use of an active compound specifically targeted to the elicitin-sterol binding site, said active compound being at least one of glabridin, carvacrol, aloin, pentofyllin a, licoflavone a, emodin anthrone, scutellarin methyl ester, aloesin, 6-feruloyl spinosin, punica granatum, aloe-emodin-8-O-beta-D-glucoside, xanthohumol, penoxsulam Huang Caogan B, (E/Z) -demethoxycurcumin, theaflavin-3-gallate, sophoraflavanone G and 3' -normethyl pericarpium Citri Tangerinae, for competitively inhibiting elicitin binding to sterols. Preferably, the active compound is glabridin, penthorum chinense pursh glycoside A or licoisoflavone A. In a second aspect, the invention claims the use of glabridin, penthorum chinense pursh or licoisoflavone a as an active compound as described above in any of the following: (1) Use