PROBING THE S' SUBSITES OF CALPAIN

描述（由申请人提供）：心脏病和中风是主要原因 美国的发病率和死亡率。越来越多的证据表明 心脏缺血（心脏病发作）或脑缺血 （中风）引发一系列生化事件，导致增加 细胞内Ca 2+浓度，这反过来激活钙蛋白酶。激活 钙蛋白酶降解结构蛋白，导致细胞死亡。钙蛋白酶是 因此被认为是一个有吸引力的治疗靶点， 心脏病和中风 长期目标是发现新的钙蛋白酶抑制剂作为治疗 心脏病和中风具体目标是确定 钙蛋白酶的S亚位点，目的是开发有效的和选择性的 酶的抑制剂。抑制剂结合的结构要求 钙蛋白酶的S亚基已被充分研究。反倒是 结构要求抑制剂结合的S？钙蛋白酶亚位点 只进行了少量的调查。这是一个巨大的差距， 文学，必须填补，因为知识的S？子网站 钙蛋白酶的特异性将提供有价值的信息，可以帮助 酶的选择性抑制剂的设计。抑制剂与天然和 非天然D-和L-氨基酸在P1？P2？抑制剂的位置 将被合成来探测S1的特异性S2？子站点 钙蛋白酶比较分子场分析（CoMFA）将用于生成 抑制剂的结合位点模型，该模型将用于设计 并在化学治疗前预测新型抑制剂的钙蛋白酶抑制效力 合成和酶学。将测试有效抑制剂的能力， 进入细胞并抑制细胞内钙蛋白酶。选择的潜能和细胞 将在大鼠离体心脏缺血模型中测试渗透性抑制剂 有心脏保护作用。

DESCRIPTION (provided by applicant): Heart disease and stroke are major causes of morbidity and mortality in the United States. Mounting evidence suggests that an episode of cardiac ischemia (heart attack) or cerebral ischemia (stroke) initiate a chain of biochemical events that result in increased intracellular Ca2+ concentration, which in turn activates calpain. Activated calpain degrades structural proteins resulting in cell death. Calpain is therefore considered as an attractive therapeutic target for intervention in heart attack and stroke. The long-term goal is to discover novel calpain inhibitors as treatment for heart attack and stroke. The specific aim is to determine the specificities of the S subsites of calpain with the objective of developing potent and selective inhibitors of the enzyme. The structural requirements for inhibitor binding to the S subsites of calpain has been well investigated. On the contrary, the structural requirements for inhibitor binding to the S? subsites of calpain have only been marginally investigated. This is a significant gap in the literature that must be filled because knowledge about the S? subsite specificities of calpain will provide valuable information that can aid in the design of selective inhibitors of the enzyme. Inhibitors with natural and unnatural D- and L-amino acids at the P1? and P2? positions of the inhibitors will be synthesized to probe the specificities of the S1? and S2? subsites of calpain. Comparative Molecular Field Analysis (CoMFA) will be used to generate a binding site model for the inhibitors and the model will be used to design and predict the calpain inhibitory potency of novel inhibitors before chemical synthesis and enzymology. Potent inhibitors will be tested for their ability to enter cell and inhibit intracellular calpain. Selected potent and cell permeable inhibitors will be tested in the rat isolated heart ischemia model for cardioprotective effect.