Chemoenzymatic Synthesis of Glycosaminoglycan Oligos

描述（由申请人提供）：糖胺聚糖（GAG）是一种结构复杂、高度硫酸化、多分散的线性多糖家族。肝素、硫酸乙酰肝素、硫酸软骨素、硫酸皮肤素和透明质酸都是这个家族的成员。肝素是研究最广泛的GAG，是丝氨酸蛋白酶抑制剂的主要激活剂，最近肝素和其他GAG已被证明在细胞生长和细胞-细胞相互作用的调节中是重要的。全世界每年从动物组织中生产近300公吨肝素，用作抗凝剂。肝素与抗凝血酶III结合，使其成为凝血丝氨酸蛋白酶的强效抑制剂。与抗凝血酶III结合的五糖序列已在>60个合成步骤中以< 0.25%的产率化学合成。尽管这种具有挑战性的合成，这种五糖目前正用于治疗。很少有报道的寡糖组成的其他糖胺聚糖的合成。提出了利用多糖裂解酶和糖苷酶化学酶促合成GAG寡糖的方法。这些类别的酶首先在主要研究者和合作研究者的实验室中分离和表征。将评价这些合成寡糖与许多GAG结合蛋白的相互作用。 目前的建议有四个具体目标。这些具体目标包括：1 .寡糖将使用多糖裂解酶制备，用于在针对GAG寡糖合成的各种糖基化反应中作为底物进行评价。2.使用标准UDP单糖供体和新型部分保护的UDP单糖供体，将使用合成酶来延伸GAG寡糖受体。3. RTILS将用作多糖裂解酶制备的寡糖的化学和酶促糖基化的溶剂。4.将使用多种GAG结合蛋白评价合成寡糖的相互作用。 合成GAG寡糖是为了评价与蛋白质结合伴侣进行选择性、高亲和力相互作用所需的关键结构元件。三维结构可以识别潜在的键合网络，但仅凭这些信息无法明确预测主要的结合力。只有类似物的合成才能提供生物学理解的结构见解，从而导致新的治疗药物。

DESCRIPTION (provided by applicant): Glycosaminoglycans (GAGs) are a family of structurally complex, highly sulfated, polydisperse, linear polysaccharides. Heparin, heparan sulfate, chondroitin sulfates, dermatan sulfate, and hyaluronic acid are all members of this family. Heparin, the most widely studied GAG, is a major activator of serine protease inhibitors and more recently heparin and other GAGs have been shown to be important in the regulation of cell growth and cell-cell interaction. Nearly 300 metric tons of heparin are produced worldwide each year from animal tissue and used as an anticoagulant. Heparin binds to antithrombin III making it a potent inhibitor of coagulation serine proteases. The pentasaccharide sequence binding to antithrombin III, has been chemically synthesized in >60 synthetic steps and in < 0.25% yield. Despite this challenging synthesis, this pentasaccharide is currently being used therapeutically. There have been few reports of the synthesis of the oligosaccharide comprising of the other GAGs. The chemoenzymatic synthesis of GAG oligosaccharides is proposed using polysaccharide lyases and synthases. These classes of enzymes were first isolated and characterized in the laboratories of the principal investigator and co-investigator. The interaction of these synthetic oligosaccharides to a number of GAG-binding proteins will be evaluated. There are four specific aims in the current proposal. These specific aims include: 1 .Oligosaccharides will be prepared using polysaccharide lyases for evaluation as substrates in a variety of glycosylation reactions directed at the synthesis of GAG oligosaccharides. 2. Synthases will be used to extend GAG oligosaccharide acceptors using both standard UDP monosaccharide donors and novel, partially protected UDP monosaccharide donors. 3. RTILS will be used as solvents for the chemical and enzymatic glycosylation of polysaccharide lyase-prepared oligosaccharides. 4. The interaction of the synthesized oligosaccharides will be evaluated using a variety of GAG-binding proteins. GAG oligosaccharides are being synthesized to evaluate critical structural elements required for selective, high affinity interaction with protein binding partners. Three-dimensional structures identify potential bonding networks, but this information alone cannot predict definitively the major binding forces. Only the synthesis of analogs can afford the structural insights for biological understanding leading to new therapeutic agents.