Uncovering the controlling mechanisms in heparan sulfate biosynthesis

说明(申请人提供)：硫酸乙酰肝素(HS)是一种重要的多糖，大量存在于细胞表面和细胞外基质中。HS参与多种生理和病理生理功能，包括凝血、炎症反应和胚胎发育。HS是一种高度硫酸盐化的多糖。肝素是HS的一种特殊形式，是一种常用的抗凝药物。HS独特的硫酸盐化模式被认为调节其功能特异性。HS的广泛生物学功能引起了人们对HS生物合成调控机制的极大兴趣。HS的生物合成涉及一系列专门的HS磺基转移酶、糖基转移酶和一种差向异构酶。使用HS生物合成酶，我们的实验室可以生产出一系列具有独特的硫酸盐化模式和功能的HS。我们的成功证明了研究特定硫酸盐糖生物合成控制的可行性。 序列。我们的假设是，HS的生物合成是由一系列上游修饰步骤控制的，包括N-硫化和C5-异构化，以及是否同时进行2-O-硫化。在这个方案中，我们将研究C5-差向异构酶和2-O-磺基转移酶控制特定糖序列的合成的机制，这些糖序列具有抗凝血活性和刺激生长因子和生长因子受体介导的细胞增殖的活性。我们将使用具有精确控制的N-硫化模式的结构定义的寡糖底物来证明C5-差向异构酶和2-O-磺基转移酶的底物专一性。最后，我们计划证明一种独特的生物合成途径，用于制备肝素中常见的高度硫酸盐化结构域。该项目的成功将引领一种全面的新方法来研究HS的生物合成，可能导致基于HS的新型疗法的开发。

DESCRIPTION (provided by applicant): Heparan sulfate (HS) is an essential glycan that is present in large quantities on the cell surface and in the extracellular matrix. HS participates ina variety of physiological and pathophysiological functions, including blood coagulation, inflammatory response and embrynic development. HS is a highly sulfated polysaccharide. Heparin, a special form of HS, is a commonly used anticoagulant drug. The uniquely distributed sulfation pattern of HS is believed to regulate its functional specificity. The wide range of biological functions of HS attracts considerable interest in understanding the mechanism for controlling the biosynthesis of HS. HS biosynthesis involves a series of specialized HS sulfotransferases, glycosyltransferases and an epimerase. Using HS biosynthetic enzymes, our labs can produce an array of HS with unique sulfation patterns and functions. Our success has proved the feasibility for studying the control of the biosynthesis of specific sulfated saccharide sequences. Our hypothesis is that the HS biosynthesis is controlled by a series of up- stream modification steps involving N-sulfation and C5-epimerization with or without concurrent 2-O-sulfation. In this proposal, we will investigate the mechanism used by C5-epimerase and 2-O-sulfotransferase to control the synthesis of specific saccharide sequences that exhibit the anticoagulant activity and the activity in stimulating growth factors and growth factor receptor mediated cell proliferation. We will use structurally defined oligosaccharide substrates with precisely controlled N-sulfation patterns to prove the substrate specificity of C5-epimerase and 2-O-sulfotransferase. Finally, we plan to prove a unique biosynthetic pathway for preparing a highly sulfated domain that is commonly found in heparin. The success of this project will lead a comprehensive new approach to investigate the biosynthesis of HS, potentially leading to the development of novel HS-based therapeutics.