STRUCTURE & FUNCTION OF NERVE GROWTH FACTOR

神经生长因子(NGF)是分化和维持的关键 感觉神经元、交感神经元和中枢神经元的蛋白质。信息 关于活性β亚基的蛋白质结构是可用的 以及7S寡聚体形式的蛋白质，在发育中的作用 神经生长因子在神经系统中的作用，其质膜的结构 受体、受体的细胞动力学和运动 在绑定之后，以及所需信令的某些方面 用于反应细胞的分化。有关的信息 β-NGF分子的结构-功能关系是 对于理解这类神经营养蛋白是必不可少的。 该项目的总体目标是确定结构 活性β-NGF亚单位的功能 与细胞受体的相互作用和产生的细胞内 使用重组DNA和免疫化学方法发出信号。 分子工程学。一个目标是通过以下方法获得β-NGF变体 分子中四个选定区域的氨基酸替换 并将这些结构变化与生物特性相关联。 在之前的赠款期间，具有生物活性的NGF 利用克隆的重组蛋白在大肠杆菌系统中成功表达 小鼠c DNA。关键氨基酸的定点突变 现在将允许确定单个残留物在 生物功能。β-神经生长因子拮抗剂和部分激动剂 也可用于生物学研究。免疫化学。 另一个目标是确定NGF分子的表面表位 通过产生特定位点参与受体识别 针对合成的NGF寡肽的抗体 蛋白。与特定序列相对应的寡肽 目前正在合成特定部位的β-NGF分子 会产生针对它们的抗体，抗体会 被用来确定表面部分的功能作用 β-NGF分子。 这些突变的NGF蛋白和位点特异性抗体将是 大鼠嗜铬细胞瘤细胞突起生长的研究 和鸡背根感觉神经元，在受体结合和 PC12细胞的细胞转运、细胞信号和基因 在形成过程中，对β-NGF自我二聚化的调节 7S齐聚物，并分析其构象 重组蛋白。这些研究的结果将推动我们的 的基本结构要求的知识 神经生长因子与神经元靶点的相互作用及其分子机制 以及神经元分化和存活的细胞方面。 将生产出新的NGF类似物，这些类似物将在 发展研究。这些信息最终将导致 更好地了解神经发育问题和相关问题 阿尔茨海默氏症和亨廷顿氏症等疾病。

Nerve Growth Factor (NGF) is a key differentiation and maintenance protein for sensory, sympathetic, and central neurons. Information is available about the protein structure of the active beta-subunit and the 7S oligomeric form of the protein, the developmental role of NGF in the nervous system, the structure of its plasma membrane receptor, the cellular dynamics and movement of the receptor subsequent to binding, and some aspects of the signalling required for differentiation of responsive cells. Information about the structure-function relationships of the beta-NGF molecule is essential for understanding such neuronotrophic proteins. The overall goal of this project is to determine structural features of the active beta-NGF subunit that are responsible for interaction with the cellular receptor and resultant intracellular signalling, using recombinant DNA and immunochemical approaches. Molecular engineering. One aim is to obtain beta-NGF variants with amino acid replacements in four selected regions of the molecule and correlate these structural changes with biological properties. In the previous grant period biologically active NGF has been successfully expressed in an E. coli system utilizing the cloned moused cDNA. Site specific mutagenesis of critical amino acids will now allow determination of the role of individual residues in biological function. Antagonists and partial agonists of beta-NGF may also be obtained for biological studies. Immunochemistry. Another aim is to determine surface epitopes of the NGF molecule involved in receptor recognition by producing site-specific antibodies directed against synthetic oligopeptides of the NGF protein. Oligopeptides corresponding to particular sequences in the beta-NGF molecule are being synthesized, site specific antibodies toward them will be produced, and the antibodies will be used to ascertain the functional role of surface portions of the beta-NGF molecule. These mutant NGF proteins and site specific antibodies will be studied by neurite outgrowth in rat Pheochromocytoma (PC12) cells and chick dorsal root sensory neurons, in receptor binding and cellular trafficking in PC12 cells, in cellular signalling and gene regulation, in the self-dimerization of beta-NGF, in the formation of the 7S oligomer, and analysis of the conformation of the recombinant protein. The results of these studies will further our knowledge of the fundamental structural requirements for the interaction of NGF with neuronal targets and elucidate molecular and cellular aspects of neuronal differentiation and survival. Novel analogues of NGF will be produced that should be useful in developmental studies. this information will eventually lead to better understanding of neurodevelopmental problems and related diseased such as Alzheimer's and Huntington's diseases.