禾谷镰孢菌β2微管蛋白与苯并咪唑类杀菌剂互作研究

小麦赤霉病是小麦上的主要病害之一,在全世界范围内引起该病害的致病菌主要是禾谷镰孢菌Fusarium graminearum。目前,使用杀菌剂是生产上防治小麦赤霉病发生和危害的主要手段,常用的杀菌剂主要有苯并咪唑类杀菌剂(benzimidazoles)等,苯并咪唑类杀菌剂的作用靶标是β2微管蛋白。本研究旨在探究小麦赤霉病菌中β2微管蛋白与苯并咪唑类杀菌剂的互作机制,通过同源建模的方法获得了禾谷镰孢菌β2微管蛋白的三维结构,并在此基础上将β2微管蛋白与4种苯并咪唑类杀菌剂(多菌灵、苯菌灵、噻菌灵、甲基硫菌灵)进行分子对接。分子对接结果显示β2微管蛋白第198位苯丙氨酸和第236位缬氨酸与4种苯并咪唑类杀菌剂直接互作形成氢键,第50、134、165、167、198、200、236、237、239、240、250、253、257、314位氨基酸形成药剂结合口袋。通过比较β2微管蛋白与4种苯并咪唑类杀菌剂结合自由能,发现与其他3种杀菌剂相比,β2微管蛋白与多菌灵的结合自由能最小(-5.72 kcal/mol),说明其与多菌灵互作亲和力更强。采用菌丝生长速率法测定了禾谷镰孢菌对4种苯并咪唑类杀菌剂的EC₅₀值,禾谷镰孢菌对多菌灵、苯菌灵、噻菌灵、甲基硫菌灵的EC₅₀值分别为0.772、0.862、1.088、13.266 mg/L,该结果表明禾谷镰孢菌对多菌灵的敏感性强于其他3种杀菌剂,与分子对接结果相吻合。

Interactions between benzimidazole fungicides and β2-tubulin of Fusarium graminearum

Fusarium head blight (FHB), mainly caused by Fusarium graminearum, is a devastating epidemic disease on wheat. At present, the control of FHB mainly depends on the application of fungicides, such as benzimidazoles, whose targets are β2-tubulin. The aim of this research is to investigate the interaction between β2-tubulin protein of F. graminearum and benzimidazoles. The β2-tubulin structure was obtained by homology modeling method. Structures of β2-tubulin complexed with the small molecular inhibitor, carbendazim, benomyl, thiabendazole and thiophanate-methyl, were built respectively by molecular docking based on β2-tubulin structure. Molecular docking results showed that the amino acid Phe198 and Val236 in F. graminearum β2-tubulin can directly interact through hydrogen bonds with four benzimidazole molecules, and the amino acid residues at position 50, 134, 165, 167, 198, 200, 236, 237, 239, 240, 250, 253, 257, 314 of F. graminearum β2-tubulin can form a drug binding pocket. The binding energy of β2-tubulin/carbendazim is lowest (-5.72 kcal/mol) and the affinity is highest, as compared with the other three complexes. The sensitivity of F. graminearum to four benzimidazole fungicides was tested using mycelium growth rate method, and the results showed that the EC₅₀ value of F. graminearum to carbendazim, benomyl, thiabendazole and thiophanate-methyl was 0.772, 0.862, 1.088, and 13.266 mg/L, respectively, indicating that F. graminearum is more sensitive to carbendazim than to the other three fungicides, and this is consistent with the molecular docking results.