Sequence Analysis of Glycosaminoglycans

糖胺聚糖的序列分析

• 批准号: 6435723
• 负责人: RAM SASISEKHARAN
• 金额: $ 30.06万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-02-01 至 2006-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6435723
• 关键词: Escherichia coli; Flavobacteriaceae; analytical method; bacterial genetics; bacterial proteins; carbohydrate sequence; catalyst; chemical kinetics; chemical structure function; enzyme activity; enzyme substrate; heparin lyase; hexosamines; high performance liquid chromatography; hyaluronidase; intermolecular interaction; matrix assisted laser desorption ionization; molecular site; mucopolysaccharides; nucleic acid sequence; polymerase chain reaction; protein purification; protein structure function; recombinant proteins; site directed mutagenesis; technology /technique development

DESCRIPTION (provided by applicant): Sequencing of DNA and proteins has heralded a biotechnology revolution. Our ability to determine the nucleic acid or polypeptide structure with a specific biological function has enabled us to probe biological phenomenon in a mechanistic, rigorous manner and facilitated the development of novel therapeutics. However, this approach has yet to be achieved with the third major class of biopolymer, viz., polysaccharide. Complex polysaccharides of the glycosaminoglycan (GAG) family are important modulators of numerous biological processes, from development to neovascularization to maintenance of the nervous system. However, expect for a few cases, it is still unknown how GAG structures impinge on function. Only with this knowledge will it be possible to utilize the information inherent in GAGs, either scientifically or therapeutically. To this end, the principal investigator has recently developed a powerful sequencing approach for a subset of GAGs (heparan sulfate-like glycosaminoglycans or HLGAGs). One of the primary experimental constraints used in this sequencing approach is the heparinases, a group of polysaccharide lyases from Flavobacterium heparinum that the principal investigator has cloned and characterized. In this grant proposal, he proposes to extend the repertoire of tools for use in the sequencing approach and to probe further the biological functions of GAGs. He proposes to do this in two ways: (1) Clone and biochemically characterize other HLGAG-degrading enzymes from F. heparinum, and (2 establish a complementary sequencing approach for chondroitin/dermatan sulfate GAGs using the chondroitinases from F. heparinum. In this manner, the principal investigator hopes to broaden the knowledge of complex polysaccharides and learn how structure translates to function.

描述（由申请人提供）：DNA 和蛋白质的测序预示着生物技术革命。我们测定核酸的能力 或具有特定生物学功能的多肽结构使我们能够 以机械、严谨的方式探索生物现象并促进 新疗法的开发。不过，这种方法还有待 这是通过第三类主要生物聚合物（即多糖）实现的。 糖胺聚糖 (GAG) 家族的复合多糖非常重要 从发育到发育的众多生物过程的调节剂 新生血管形成以维持神经系统。然而，期望 在少数情况下，GAG 结构如何影响功能仍不清楚。仅有的 有了这些知识，就可以利用其中固有的信息 GAG，无论是科学上的还是治疗上的。为此，校长 研究人员最近开发了一种针对子集的强大测序方法 GAG（硫酸乙酰肝素样糖胺聚糖或 HLGAG）。初级之一 该测序方法中使用的实验限制是肝素酶，一种 来自肝素黄杆菌的一组多糖裂解酶，其主要成分是 研究人员已对其进行克隆和表征。在这份拨款提案中，他建议 扩展用于测序方法的工具库并 进一步探讨GAGs的生物学功能。他建议分两步进行 方法：（1）其他HLGAG降解酶的克隆和生化表征 来自 F. heparinum，并且（2 建立一种互补测序方法 使用来自肝素肝素的软骨素酶制备软骨素/硫酸皮肤素 GAG。 首席研究员希望通过这种方式拓宽知识面 复杂的多糖并了解结构如何转化为功能。