AGING & NEURONAL DEATH--FIRST GENERATION OF NGF ANALOG

老化

• 批准号: 3121813
• 负责人: FRANK M LONGO
• 金额: $ 8.8万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-05-01 至 1994-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3121813
• 关键词: active sites; aging; bioassay; cell death; chickens; dorsal root; ganglions; growth factor receptors; high performance liquid chromatography; immunocytochemistry; inhibitor /antagonist; laboratory mouse; mass spectrometry; molecular site; neurons; neurotrophic factors; peptide analog; peptide chemical synthesis; peptide structure; protein purification; protein structure; receptor binding; stimulant /agonist; synthetic peptide; tissue /cell culture

One of the primary pathological processses occurring in the aging nervous system is neuronal death. For example, in Alzheimer's disease (AD) cholinergic neurons in the basal forebrain, which are thought to function in memory and learning, are lost. Neurotrophic factors are proteins which interact with cell surface receptors to promote the survival and growth of neurons. There is considerable evidence that neurotrophic factors such as nerve growth factor (NGF) can prevent neuronal death or damage and that they may be able to enhance the function of compromised neurons. Basal forebrain cholinergic neurons have NGF receptors and respond to NGF; therefore, NGF may be relevant to understanding and perhaps minimizing the loss of neurons in AD. A significant limitation to both understanding the physiological role of endogenous NGF in vivo and the potential therapeutic application of NGF is the lack of stable, low molecular weight agonists and antagonists of NGF which can effectively reach the central nervous system and penetrate the brain parenchyma. The long term objectives of this work are: i) to begin to define structural features of NGF (NGF's active site) which will allow it to interact with its receptor and ii) to develop a first generation of low molecular weight synthetic NGF peptide analogs which can mimic or inhibit the neurotrophic effect of NGF in vitro and eventually in vivo. The general strategy of this proposal is to map the active site(s) of NGF by synthesizing short peptides corresponding to potential active sites. These peptides will be assayed for their ability to block or mimic several measures of NGF bioactivity in vitro. By systematically varying which residues of NGF are includes in each peptide and by making peptides of different lengths, one can begin to map an active site. Active peptides will then be modified by conventional techniques to form a first generation of NGF analogs. This synthetic peptide strategy has been used many times to define active sites and produce active analogs of many proteins, most of which are considerably more complex than NGF. Indeed, our initial studies with NGF peptides suggest that this method will be successful with NGF as well. The specific aims during the proposed grant period will be: 1) To synthesize and purify NGF peptides and analogs derived from peptides which are active. 2) To test these peptides and analogs for their ability to mimic or inhibit three well-characterized activities of NGF in vitro. 3) To test the most potent peptides for their ability to compete with NGF's binding at its receptor.

衰老的主要病理过程之一 神经系统是神经元死亡。 例如，在老年痴呆症中， (AD)基底前脑中的胆碱能神经元被认为 记忆和学习功能丧失。 神经营养因子是 蛋白质与细胞表面受体相互作用， 神经元的存活和生长。 有相当多的证据表明 神经营养因子如神经生长因子（NGF）可以防止 神经元死亡或损伤，它们可能能够增强 受损神经元的功能。 基底前脑胆碱能神经元 有神经生长因子受体，并对神经生长因子作出反应;因此，神经生长因子可能与 了解并可能最大限度地减少AD中神经元的损失。 一 这对理解生理作用的重要限制 体内内源性神经生长因子及其潜在的治疗应用 缺乏稳定的低分子量的激动剂和拮抗剂， 神经生长因子能有效地到达中枢神经系统， 脑实质 这项工作的长期目标是：i） 开始定义NGF的结构特征（NGF的活性位点）， 将允许它与其受体相互作用，以及ii）开发第一个 低分子量合成的NGF肽类似物的产生 可以在体外模拟或抑制NGF的神经营养作用， 最终在体内。 本提案的总体战略是绘制 通过合成相应的短肽来确定NGF的活性位点 潜在的活性位点。 将测定这些肽的 能够阻断或模拟体外NGF生物活性的几种测量。 通过系统性地改变每一个细胞中包括哪些NGF残基， 通过制备不同长度的肽，人们可以开始 绘制一个活动站点。 活性肽然后将被修饰， 用常规技术制备第一代NGF类似物。 这 合成肽策略已多次用于定义活性肽， 位点并产生许多蛋白质的活性类似物，其中大多数是 比NGF复杂得多。 事实上，我们最初的研究 神经生长因子肽表明，这种方法将是成功的与神经生长因子， 好. 在建议的资助期内，我们的具体目标如下：1） 合成和纯化NGF肽和衍生自肽的类似物 它们是活跃的。2)为了测试这些肽和类似物的 模拟或抑制NGF的三种充分表征的活性的能力 体外3)为了测试最有效的肽， 与神经生长因子的受体竞争。