Discovery of novel inhibitors targeting the PH domain of PcORP1: Virtual screening, structural optimization,...

The oxysterol-binding protein (OSBP)-related protein 1 (ORP1) serves as a critical target for oomycete inhibitors, given that OSBPI fungicides demonstrate exceptionally high biological efficacy through their specific binding to the ORD domain of ORP1. Our previous research has demonstrated that the PH domain of ORP1 in Phytophthora species plays a crucial role in its biological functions. However, the potential of this domain as a viable target for fungicides remains to be elucidated. In this study, we describe the discovery of the PH domain inhibitor, compound X4. Initially, a hit compound VS-07, with good fungicidal activity against oomycetes, was identified through docking-based virtual screening. Subsequently, 27 analogs were designed and synthesized by structurally optimizing of VS-07. Among these, compound X4 demonstrated exceptional inhibitory activities against six oomycetes, with EC₅₀ values ranging from 1 to 4 μg/mL, and effectively inhibited the PH domain of PcORP1. Molecular docking revealed that compound X4 formed more hydrogen bond interactions with Trp51, Ser91, and Thr93 of the PH domain. Preliminary mechanistic studies indicated that compound X4 affected lipid transport and metabolism by targeting the PH domain of PcORP1, resulting in mycelium hyperplasia and cell membrane ruffling in P. capsici. Moreover, compound X4 provided over 90 % protection against P. capsici in detached leaf assays at 100 and 200 μg/mL. This study not only validates the targetability of the PH domain of ORP1, but also identifies compound X4 as a potential inhibitor targeting this domain, offering a potential fungicide for oomycete management.