Uncovering the controlling mechanisms in heparan sulfate biosynthesis

描述（由申请方提供）：硫酸乙酰肝素（HS）是一种大量存在于细胞表面和细胞外基质中的必需聚糖。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是高度硫酸化的多糖。肝素是HS的一种特殊形式，是常用的抗凝药物。HS的独特分布的硫酸化模式被认为调节其功能特异性。HS广泛的生物学功能吸引了人们对理解HS生物合成控制机制的极大兴趣。HS生物合成涉及一系列专门的HS磺基转移酶、糖基转移酶和差向异构酶。使用HS生物合成酶，我们的实验室可以生产一系列具有独特硫酸化模式和功能的HS。我们的成功为研究特定硫酸化糖的生物合成控制提供了可行性 序列的我们的假设是HS生物合成受一系列上游修饰步骤控制，包括N-硫酸化和C5-差向异构化，同时存在或不存在2-O-硫酸化。在本研究中，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合成的机制，这些糖序列具有抗凝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-硫酸化模式的结构确定的寡糖底物来证明C5-差向异构酶和2-O-磺基转移酶的底物特异性。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化结构域。该项目的成功将导致一种全面的新方法来研究HS的生物合成，可能导致开发新的基于HS的治疗方法。

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.