Structural Characteristics for the Interaction of 1-Benzyl-2-Methylbenzimidazoles as Insect Growth Regulators and Juvenile Hormone Binding Protein

The authors previously reported that 2-methylbenzimidazole derivatives (MBIs) exhibit insect growth-regulating activity against the silkworm, Bombyx mori. However, despite their unique effects on juvenile hormone (JH)-related endocrine pathways, the precise mode of action of MBIs remained unclear. In the present study, the interactions between MBIs and the lepidopteran hemolymph JH-binding protein (JHBP), a key regulator of JH transport and activity, were investigated using multiple approaches, including in vitro binding affinity assays, X-ray crystallography, and molecular docking simulations. A series of MBIs bearing a 1-(4-alkoxybenzyl) group, which exhibited potent insect growth-regulating activity, showed high binding affinity and structural compatibility with the JH-binding pocket of JHBP. In contrast, 1-(4-alkylbenzyl) MBIs, which displayed weak or negligible insect growth-regulating activity, exhibited low affinity for JHBP. These findings suggest that the insect growth-regulating activity of MBIs is mediated through inhibition of JHBP function, likely by disrupting the precise regulation of JH concentration in the hemolymph during larval development and pupal metamorphosis.