新靶标发现是新农药创制研究的重要方向。水稻白叶枯病菌（Xanthomonas oryzae pv. oryzae, Xoo）严重危害我国水稻生产，防治十分困难。本课题组前期发现2个高效抗Xoo的喹唑啉化合物KN1和KN2，为阐明其作用机制，本申请拟基于活性的蛋白质组分析（activity-based protein profiling，ABPP）揭示活性化合物的分子靶标。具体为：①制备CC（click chemistry）-ABPP直接反应型和PAL（photo affinity labeling）-CC-ABPP光亲和反应型分子探针。②评估探针对Xoo的抑制活性，垂钓菌体内潜在靶标蛋白。③采用pull down技术分离、鉴定靶标蛋白，通过原核表达获得蛋白，开展蛋白与KN1、KN2及探针的体外相互作用等验证分子靶标。研究结果有望为抗水稻白叶枯病菌新杀菌剂先导结构发现提供重要科学基础。

Molecular target discovery is one of the most important researches in agrochemical innovations. Xanthomonas oryzae pv. oryzae (Xoo) seriously harms rice production in China and is very difficult to control. Previously, two quinazoline compounds KN1 and KN2 were found by our group with excellent antibacterial activity against Xoo. This grant aims to unravel the molecular targets of KN1 and KN2 using activity-based protein profiling (ABPP) technology. Firstly, two kinds of molecular probes, which are CC (click chemistry)-ABPP probe with direct reaction ability and PAL (photo affinity labeling)-CC-ABPP photoaffinity probe, will be synthesized and bioassayed against Xoo. Then the bioactive probes will be used for target identification in Xoo cells in vivo. Pull down experiments will then be performed to enrich target proteins, followed by on-bead digestion and LC-MS/MS analysis to give the potential target proteins. Prokaryotic expression will be carried out to obtain recombinant target proteins for further evaluation of interactions with KN1, KN2 and probes. The findings of this study are expected to provide insightful scientific data for innovation of novel antibacterial chemical entity against Xoo.