Molecular and structural biostudies on streptococcalinyasion/infection mechanisms to host cells through degradation of extracellular matrices glyoceaminoglycans

一些致病微生物产生多糖降解酶（裂解酶和水解酶）侵入哺乳动物和/或植物细胞。哺乳动物糖胺聚糖构成细胞表面基质的一部分，是微生物酶的典型靶标。不饱和葡萄糖醛酸水解酶（UGL）在遗传上首次在芽孢杆菌菌株GL1中被发现，它催化通过降解基质的多糖裂解酶（如透明质酸和软骨素裂解酶）反应产生的寡糖水解释放不饱和糖醛酸，表明这些酶在微生物感染中起着毒力因子的作用。本研究分析了糖胺聚糖降解酶——裂解酶和水解酶的结构和功能关系。基于酶-底物复合物的x射线晶体学和位点定向诱变，阐明了透明质酸裂解酶同系物（黄原胶裂解酶）的催化反应和底物识别机制。一个单独的酪氨酸…More残基在亚位+1处提取醛酸残基的c5 -质子，并将质子提供给糖苷键进行裂解。与具有保持或倒置催化机制的一般糖苷水解酶不同，UGL独特地触发了不饱和脲酸酯残基中的乙烯醚基团的水解，而不是糖苷键的水解。最近的全基因组序列分析表明，从细菌到真菌（超过70种）的大量微生物在其基因组中具有UGL同源基因。在碳水化合物活性酶（CAZy）数据库中，UGL及其同源物组成了一个新的家族GH-88。UGL的微生物生产者包括致病菌，如梭状芽胞杆菌、链球菌和弧菌。该酶同源基因在链球菌、无乳链球菌、肺炎链球菌、化脓性链球菌和猪链球菌中均有发现。与UGL芽孢杆菌不同，链球菌UGL作用于不饱和软骨素双糖，在GalNAc的C4上有一个硫酸盐基团。少

Some pathogenic microorganisms produce polysaccharide-degrading enzymes (lyases and hydrolases) to invade mammalian and/or plant cells. Mammalian glycosaminoglycans that form part of cell surface matrix are typical targets for microbial enzymes. Unsaturated glucuronyl hydrolase (UGL), which was genetically first identified in Bacillus sp. strain GL1, catalyzes the hydrolytic release of an unsaturated uronic acid from oligosaccharides produced through the reaction of the matrix-degrading polysaccharide lyases (e.g., hyaluronate and chondroitin lyases), suggesting that these enzymes function as a virulent factor in microbial infection. In this study, structure and function relationship of glycosaminoglycan-degrading enzymes, lyase and hydrolase, was analyzed.Based on X-ray crystallography of enzyme-substrate complexes and site-directed mutagenesis, mechanisms for catalytic reaction and substrate recognition of hyaluronate lyase homologue (xanthan lyase) were clarified. A single tyrosine … More residue abstracts C5-proton of the uronate residue at subsite +1 and donates the proton to glycosidic bond to be cleaved.In contrast with general glycoside hydrolases with the retention or inversion catalytic mechanism of an anomeric configuration, UGL uniquely triggers hydrolysis of vinyl ether groups in the unsaturated uronate residue but not of the glycosidic bond.Recent complete genome sequence analyses indicate that a large number of microorganisms ranging from bacteria to fungi (over 70 species) have a UGL homologous gene in their genome. In the Carbohydrate-Active enZyme (CAZy) database, UGL and its homologues form a new family, GH-88. Microbial producers of UGL include pathogenic bacteria such as clostridia, streptococci, and vibrios. The enzyme homologous gene has been found especially in streptococci, Streptococcus agalactiae, Streptococcus pneumoniae, Streptococcus pyogenes, and Streptococcus suis. Unlike the bacillus UGL, streptococcal UGL acts on unsaturated chondroitin disaccharide with a sulfate group at C4 of GalNAc. Less