Brognaro Hevila, Almeida Vitor Medeiros, de Araujo Evandro Ares, Piyadov Vasily, Santos Maria Auxiliadora Morim, Marana Sandro Roberto, Polikarpov Igor
Instituto de Física de São Carlos, Universidade de São Paulo, Avenida Trabalhador São Carlense 400, São Carlos, SP, 13566-590, Brazil.
Instituto de Química, Universidade de São Paulo, Avenida Prof. Lineu Prestes, 748, Bloco 10, Sala 1054, São Paulo, SP, 05508-900, Brazil.
Mol Biotechnol. 2016 Dec;58(12):777-788. doi: 10.1007/s12033-016-9977-3.
The marine bacteria Saccharophagus degradans (also known as Microbulbifer degradans), are rod-shaped and gram-negative motile γ-proteobacteria, capable of both degrading a variety of complex polysaccharides and fermenting monosaccharides into ethanol. In order to obtain insights into structure-function relationships of the enzymes, involved in these biochemical processes, we characterized a S. degradans β-glycosidase from glycoside hydrolase family 1 (SdBgl1B). SdBgl1B has the optimum pH of 6.0 and a melting temperature T of approximately 50 °C. The enzyme has high specificity toward short D-glucose saccharides with β-linkages with the following preferences β-1,3 > β-1,4 ≫ β-1,6. The enzyme kinetic parameters, obtained using artificial substrates p-β-NPGlu and p-β-NPFuc and also the disaccharides cellobiose, gentiobiose and laminaribiose, revealed SdBgl1B preference for p-β-NPGlu and laminaribiose, which indicates its affinity for glucose and also preference for β-1,3 linkages. To better understand structural basis of the enzyme activity its 3D model was built and analysed. The 3D model fits well into the experimentally retrieved low-resolution SAXS-based envelope of the enzyme, confirming monomeric state of SdBgl1B in solution.
海洋细菌嗜糖降解菌（也称为降解微球杆菌）是杆状革兰氏阴性运动性γ-变形菌，既能降解多种复杂多糖，又能将单糖发酵成乙醇。为了深入了解参与这些生化过程的酶的结构-功能关系，我们对糖苷水解酶家族1中的嗜糖降解菌β-糖苷酶（SdBgl1B）进行了表征。SdBgl1B的最适pH为6.0，解链温度T约为50℃。该酶对具有β-连接的短D-葡萄糖寡糖具有高度特异性，偏好顺序为β-1,3＞β-1,4≫β-1,6。使用人工底物对-β-NPGlu和对-β-NPFuc以及二糖纤维二糖、龙胆二糖和昆布二糖获得的酶动力学参数表明，SdBgl1B偏好对-β-NPGlu和昆布二糖，这表明其对葡萄糖的亲和力以及对β-1,3连接的偏好。为了更好地理解酶活性的结构基础，构建并分析了其三维模型。该三维模型与通过实验获得的基于小角X射线散射的酶的低分辨率包络线拟合良好，证实了SdBgl1B在溶液中的单体状态。
