OPIOID PEPTIDE PROCESSING ENZYMES

描述（申请人摘要）： 要求K 02奖的更新是为了使申请人免于 增加实务教学及行政职责 研究生产力。 在过去的4年里，PI能够 她将80%以上的时间用于研究活动， 这大大增强了我们的 实验，建议博士后研究员和研究生，并获得 通过合作和动手来开发新技术 实验 预计这一研究工作的加强将 在申请的续期期间继续保护 由K 02奖提供;没有它，教学责任可能是 急剧增加，最明显的是假设， 在一个主要的服务课程的董事。 该提案基于目前正在进行的两个项目，一个是关于 阿片肽前体的蛋白水解加工，以及 PC 2结合蛋白7 B2的作用机制。 四个具体目标 1）使用以下方法确定重组PC 1和PC 2的特异性： 重组正常和突变型脑啡肽原及相关荧光素 基板;这些实验将描绘基板的偏好， 两台PC 2)为了鉴定新的PC 1抑制剂，研究其生物化学和 PC 1和PC 2抑制剂的细胞生物学，并评估 体内脑啡肽原加工的靶向抑制剂。 这些实验 将补充第一个具体目的中关于酶的描述 特异性，并应产生合成酶的设计信息 抑制剂的 3)确定7 B2在proPC 2调节中的作用 神经内分泌细胞系的转化，重点关注 前肽及其与7 B2裂解/解离的关系 forms. 4)为了探讨7 B2和PC 2的分子间相互作用， 体外实验 综合起来，这些实验应该可以帮助我们 了解阿片类激素原转化酶的基本生物化学 肽前体以及转化酶活性的调节。

DESCRIPTION (Applicant's Abstract): This renewal of a K02 award is requested in order to free the applicant from substantive teaching and administrative responsibilities to increase research productivity. During the past 4 years the PI has been able to devote more than 80% of her time to research activities, a level of effort which has resulted in a significantly enhanced ability to perform experiments, advise postdoctoral fellows and graduate students, and acquire new technologies both by collaboration as well as by hands-on experimentation. This enhancement of research effort is expected to continue during the renewal period of the application with the protection afforded by the K02 award; without it, teaching responsibilities are likely to increase dramatically, most notably by the assumption of course directorship in a major service course. The proposal is based upon two currently ongoing projects, one on the proteolytic processing of opioid peptide precursors, and one on the mechanism of action of the PC2 binding protein 7B2. The four specific aims are 1) to define the specificity of recombinant PC1 and PC2 using recombinant normal and mutant proenkephalins and related fluorogenic substrates; these experiments will delineate substrate preferences of the two PCs. 2) To identify novel PC1 inhibitors, study the biochemistry and cell biology of PC1 and PC2 inhibitors, and assess potential effects of targeted inhibitors on proenkephalin processing in vivo. These experiments will complement those described in the first specific aim as to enzyme specificity and should yield information on the design of synthetic enzyme inhibitors. 3) To determine the role of 7B2 in the regulation of proPC2 conversion in neuroendocrine cel1 lines, with a focus on the fate of the propeptide and the relationship of its cleavage/dissociation to that of 7B2 forms. 4) To explore the molecular interaction of 7B2 and PC2 through in vitro experiments. Taken together, these experiments should help us to understand the basic biochemistry of the prohormone convertases on opioid peptide precursors as well as the regulation of convertase activity.