Structural and functional studies of enzymes involved in glycosaminoglycan synthesis and degradation

参与糖胺聚糖合成和降解的酶的结构和功能研究

• 批准号: 155375-2012
• 负责人: Cygler, Mirek
• 金额: $ 2.48万
• 依托单位: University of Saskatchewan
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2018
• 资助国家: 加拿大
• 起止时间: 2018-01-01 至 2019-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=648455
• 关键词: Structural; functional; studies; enzymes; involved

This proposal aims at a structural and functional characterization of enzymes involved in synthesis and degradation of glycosaminoglycans (GAGs). GAGs are negatively charged linear polysaccharides that are essential for providing resilience to connective tissues, storage of growth factors and other proteins, participate in tissue repair and signaling through receptors. The presence of GAGs is usually associated with higher organisms; however, they are not unique to eukaryotes. Several specialized microorganisms also produce unsulfated forms of these polymers. Mammalian GAGs undergo rapid turnover through action of hydrolases. Bacteria evolved enzymes that use a lytic mechanism to degrade GAGs, namely polysaccharide lyases. These enzymes help bacteria to gain access to mammalian cells by degrading the surface polysaccharides or to utilize GAGs as a food source.**Here we propose to expand our research into GAG processing enzymes and provide structural, functional and enzymological characterization of hyaluronan synthase, GAG sulfotransferases and polysaccharide lyases. All these proteins may also potential applications in synthesis of designer oligosaccharides for treatment of diseases and specific tools for characterization of GAGs.**The short-term objectives are:**1. To determine the structures and catalytic mechanisms of polysaccharide lyases from families PL12 and PL17 and establish their evolutionary relationship to other PL families. **2. To establish through structural investigations and site-directed mutagenesis the mechanism of a concerted alternate addition of a glucuronate and a glucosamine by two independent glycosyltransferase active sites within hyaluronan synthase. **3. To determine the structures of lumenal fragments of mammalian sulfotransferases and identify features which define their positional specificity for sulfate addition.**

该提案旨在对参与糖胺聚糖（GAG）合成和降解的酶进行结构和功能表征。GAG是带负电荷的线性多糖，其对于为结缔组织提供弹性、储存生长因子和其他蛋白质、参与组织修复和通过受体进行信号传导至关重要。GAG的存在通常与高等生物有关;然而，它们并不是真核生物所独有的。几种特殊的微生物也产生这些聚合物的非硫酸化形式。哺乳动物GAG通过水解酶的作用进行快速周转。细菌进化出使用裂解机制降解GAG的酶，即多糖裂解酶。这些酶通过降解表面多糖或利用GAG作为食物来源，帮助细菌进入哺乳动物细胞。在这里，我们建议扩大我们的研究GAG加工酶，并提供透明质酸合成酶，GAG磺基转移酶和多糖裂解酶的结构，功能和酶学特性。所有这些蛋白质也可能在合成用于治疗疾病的设计寡糖和用于表征GAG的特定工具中具有潜在的应用。短期目标是：**1。确定PL 12和PL 17家族多糖裂解酶的结构和催化机制，并建立它们与其他PL家族的进化关系。** 二.通过结构研究和定点诱变，确定透明质酸合酶内两个独立的糖基转移酶活性位点协同交替添加葡萄糖醛酸和葡萄糖胺的机制。** 三.确定哺乳动物磺基转移酶内腔片段的结构，并确定其对硫酸盐加成的位置特异性特征。