刺梨是贵州重点发展的山地特色水果资源。项目前期研究发现，嗜酸乳杆菌GIM1.208产β-葡萄糖苷酶(BGL）对刺梨重要黄酮苷元—槲皮素具有高效释放能力，但相关机制尚不清楚。据此，本项目通过基因克隆外源表达获得GIM1.208BGL重组酶，进而建立BGL催化反应动力学模型，探明BGL对刺梨不同槲皮素糖苷底物识别特异性；采用DLS、FS、CD、MALDI-TOF-MS及分子序列分析，明确BGL催化活性结构、酶与不同底物识别的结构模式及分子间作用差异化表征；同时通过同源建模及分子对接三维结构模型与二维图谱研究，进一步揭示BGL及酶-底物结合的活性中心、活性位点、关键氨基酸残基及非共价作用与催化活性的相关性，最终阐明GIM1.208BGL高效释放刺梨槲皮素的底物结合模式及分子作用机制。该研究对刺梨功能挖掘提升及益生菌功能开发具有重要意义，同时为BGL下游工程研究提供必要的理论依据和指导。

Rosa roxburghii Tratt is the focus in the development of fruit resources with mountainous region characteristic in Guizhou. Our preliminary study found that β-glucosidase produced by L.acidophilusGIM1.208 can effectively induce the release of quercetin--the main aglycones in Rosa roxburghii Tratt，but the mechanism remains unclear. In this program,the recombinase was attained by cloning and heterologous expression of BGL gene from GIM1.208, then its specific substrate was clarified by analyzing enzyme activity and establishing the kinetic model. Meanwhile,the molecular structure combining with different substrates and different features of molecular interaction were revealed by determining the amino acid sequence, the dynamic scattering spectrum, the fluorescence spectrum and the circular dichroism. Finally,through the study of BGL homology model,the docking method for target substrate molecular using three-dimensional structure model and two-dimensional mapping, the relationship between BGL and the structure of active sites, active site, amino-acid residues, and the non-covalent interaction of different complexes were further clarified. Consequently, it will be elucidated that the substrate binding model and Interaction mechanisms of releasing effectively quercetin by BGL from Rosa roxburghii Tratt. This program has great significance for improvement of the flavonoid bioactivity from Rosa roxburghii Tratt and application of the probiotics, and provides guidance for the BGL functional modification and its downstream engineering as well.