Defining Macromolecular Recognition in Serine Proteases

本研究的目的是了解丝氨酸蛋白酶的底物特异性与有效抑制剂的活性位点结合序列之间的关系。丝氨酸蛋白酶在生物学中无处不在，可能在许多致病状态中发挥作用，包括血栓性疾病、高血压、骨关节炎、慢性退行性疾病和癌症。一个荧光合成肽底物组合文库将用于阐明一系列癌症相关丝氨酸蛋白酶的首选底物序列。从这些研究中，深入了解蛋白酶特异性的结构决定因素以及特定蛋白酶的生物学靶点信息。利用噬菌体展示技术，结合重新设计二聚体蛋白酶抑制剂ecotin的活性位点结合域，将开发出有效的单个蛋白酶抑制剂。对涉及底物或抑制剂识别的扩展蛋白-蛋白相互作用的了解将进一步加深我们对蛋白酶及其抑制剂之间结构-功能关系的理解。癌症相关丝氨酸蛋白酶的体内功能将开始被阐明，具有潜在治疗价值的有效抑制剂将被开发出来。

The objective of the proposed research is to develop an understanding of the relationship between the substrate specificity of a serine protease and the active-site binding sequence of a potent inhibitor. Serine proteases are ubiquitous in biology and may play a role in a number of pathogenic states including thrombotic disorders, hypertension, osteoarthritis, chronic degenerative disorders and cancer. A combinatorial library of fluorogenic synthetic peptide substrates will be used to elucidate the preferred substrate sequence for a series of cancer-related serine proteases. From these studies, insights into the structural determinants of protease specificity will be obtained along with information about the biological targets of specific proteases. Potent inhibitors of individual proteases will be developed by combinatorially redesigning the active- site binding domain of the dimeric protease inhibitor ecotin using phage display technologies. A knowledge of the extended protein-protein interactions that are involved in substrate or inhibitor recognition will further our understanding of the structure-function relationship among proteases and their inhibitors. The in vivo functions of cancer-related serine proteases will begin to be elucidated and potent inhibitors with potential therapeutic value will be developed.