Synthesis, molecular docking simulation, and antifungal activities of novel strobilurin derivatives containing covalent reactive warheads

Abstract

The prolonged overuse of strobilurin fungicides has exacerbated resistance issues, diminishing control efficacy. Despite extensive modification of strobilurin core scaffolds, fungicide resistances persist unresolved. To discover strobilurin fungicides with novel modes of action, we performed covalent drug design targeting the Cys39 residue of cytochrome b in Colletotrichum gloeosporioides. Specifically, covalent reactive warheads (acrylamide and chloroacetamide) were strategically incorporated into methoxyimino acetate and methoxyacrylate scaffolds through rational molecular design. This approach yielded two series of 36 novel strobilurin derivatives, with some exhibiting significantly higher antifungal activity against C. gloeosporioides. Compounds 6e, 6h, 6i, 6j, 6s, and 14c exhibited EC₅₀ values (42.0, 41.0, 18.5, 25.9, 43.8, and 19.8 μg/mL, respectively) exceeding that of kresoxim-methyl (EC₅₀ > 100 μg/mL), with 6i and 14c showing particularly potent activity. Morphological analysis, electrical conductivity assays and intracellular content leakage measurements were conducted to confirm compound 6i disrupted cellular membrane integrity. Further non-covalent and covalent molecular docking results indicated compounds 6i and 14c bound effectively to cyt b and positioned their functional groups in a favorable geometry for covalent reaction with Cys39. This study offers a potential solution for structural modification of resistant pesticides.