Collaborative Project: Receptors for Juvenile Hormones and JH Mimics

拟议的研究是一个跨学科的延续 在结构、动力学和生理学方面的合作 结合和降解保幼激素（JH）的蛋白质， 模拟物（JHM）。 这项合作研究的最初目标 发展化学和生物化学技术， 保幼激素（JH）受体的分子生物学， 猫 这些初步目标已经实现， 在这一更新请求中，提出了进一步的目标：第一， 研究了JH和JHM结合蛋白（JHBP和JHM）的结构， RP）在Manduca幼虫表皮细胞胞质溶胶和细胞核中的含量， JH酯酶（JHE）和环氧化物水解酶（JHEH）至（a） 放射性配体合成，（B）JHBP/RP和JHE/EH纯化 和测序，（c）活性位点标记和测序，（d） 抗体产生，（e）基因分离和测序，和（f） 蛋白质的过度生产;第二，检查 配体和大分子，包括（a）时间和位置 （B）激素和蛋白质在JHBP/RP、JHE/EH之间的运动 细胞区室，和（c）蛋白质-配体，蛋白质-蛋白质，和 蛋白质-DNA相互作用 结构和动力学的关键 目标将是开发一个杆状病毒表达系统 在墨西哥的蛋白质生产过剩。 第三个目标是 将结构和动力学与生理学联系起来， 表征，（a）JHBP/RP和JHE/EH的出现， 生命周期，和（B）JHBP/RP和 昆虫中的JHE/EH，关于配体识别和蛋白质 相似之处 保幼激素受体蛋白的鉴定 (JH)和JH模拟（JHM）和他们的途径的知识， 降解是理解分子基础的关键 用于昆虫基因表达的激素调节。 这 研究是一个跨学科的合作，结合 分子生物学和生理表现方面的专业知识 昆虫（L. M. Riddiford），生化和 JH代谢的分子研究（B. D.吊床）和合成 以及使用高比活性标记的JH和JHM（G. D. Prestwich）。 该提案是在NSF倡议下提交的 生命过程的化学

The proposed research is a continuation of an interdisciplinary collaboration on the structure, dynamics, and physiology of proteins which bind and degrade juvenile hormone (JH) and JH mimics (JHM). The original goals of this collaborative research were to develop chemical and biochemical techniques to approach the molecular biology of juvenile hormone (JH) reception and catabolism. These initial objectives have been acheived, and in this renewal request, further goals are proposed: First, to study the structure of the JH and JHM binding proteins (JHBP an RP) in Manduca larval epidermal cell cytosol and nuclei, and of JH esterases (JHE) and epoxide hydrolases (JHEH) through (a) synthesis of radioligands, (b) JHBP/RP and JHE/EH purification and sequencing, (c) active site labeling and sequencing, (d) antibody production, (e) gene isolation and sequencing, and (f) overproduction of proteins; second, to examine the dynamics of the ligands and macromolecules, including (a) timing and location of JHBP/RP, JHE/EH, (b) movement of hormones and proteins between cell compartments, and (c) protein-ligand, protein-protein, and protein-DNA interactions. A key to the structure and dynamics goals will be the development of a baculovirus expression system in Bombyx for protein overproduction. The third goal is to correlate structure and dynamics with physiology by characterizing, (a) the appearance of JHBP/RP and JHE/EH in the life cycle, and (b) comparative biochemistry of JHBP/RP and JHE/EH in insects, with respect to ligand recognition and protein similarities. The characterization of receptor proteins for juvenile hormone (JH) and JH mimics (JHM) and knowledge of the pathways of their degradation are major keys to understanding the molecular basis for hormonal regulation of gene expression in insects. This research is an interdisciplinary collaboration which combines expertise in molecular biology and physiological manifestations of gene expression in insects (L. M. Riddiford), biochemical and molecular studies of JH metabolism (B. D. Hammock), and synthesis and use of high specific activity labeled JHs and JHMs (G. D. Prestwich). This proposal was submitted under the NSF initiative in the Chemistry of Life Processes.