Novel Molecules as In Vivo Biological Probes

作为体内生物探针的新型分子

• 批准号: 7983424
• 负责人: THOMAS James WANDLESS
• 金额: $ 31.73万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-01-01 至 2014-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7983424
• 关键词: Affinity; Behavior; Biological; Biological Models; Cell physiology; Cells; Complement; Cultured Cells; DNA; Dependence; Development; Disease model; Dose; Engineering; Ensure; Equilibrium; Eukaryotic Cell; Funding; Genes; Genetic; Goals; Hour; Human; Life; Ligand Binding; Ligands; Mammalian Cell; Mammals; Mass Spectrum Analysis; Methodology; Methods; Modeling; Mus; Pharmaceutical Preparations; Proteins; RNA Interference; Reagent; Research; Research Personnel; Screening procedure; Small Interfering RNA; Specificity; Speed; System; Tacrolimus Binding Proteins; Techniques; Technology; Tertiary Protein Structure; Testing; Tetracyclines; Withdrawal; base; design; human disease; improved; in vivo; interest; knock-down; mRNA Precursor; mutant; novel; programs; prospective; protein degradation; protein function; protein misfolding; public health relevance; small molecule; transgene expression

DESCRIPTION (provided by applicant): The broad, long-term objective of this research program is to develop general technology to conditionally regulate protein function at the level of the protein molecules rather than by targeting the DNA or mRNA precursors that encode a protein-of-interest. This technology is highly specific for the targeted protein and provides rapid and tunable control of protein function using cell-permeable small molecules. The goal is to engineer small protein domains called destabilizing domains that are rapidly and conditionally degraded when expressed in mammalian cells. The instability of a destabilizing domain is faithfully conferred to other proteins fused to these small domains. A cell-permeable ligand that binds tightly to the destabilizing domains regulates their stability. The genetic fusion of the destabilizing domain to the gene-of-interest ensures specificity, and the attendant small-molecule control confers speed, reversibility and dose-dependence to this method. The reagents developed to date involve domains that are unstable in the absence of ligand and stabilized when ligand binds. The specific aims of this proposal are designed to deliver complementary new domains that are destabilized rather than stabilized by the addition of the cell-permeable ligand. A mechanistic understanding of how these domains are recognized and degraded in mammalian cells will make this technology more useful to users. These studies may also reveal general mechanisms that cells use to recognize and degrade unfolded or misfolded proteins, and these mechanisms are likely relevant to important human diseases. PUBLIC HEALTH RELEVANCE: The goal of this research program is to develop novel methods to rapidly and reversibly regulate the stability of specific proteins in eukaryotic cells and living mammals. This methodology would enable the development of many new models for human diseases, and these model systems are enormously helpful in the ongoing search for new and improved drugs to treat human diseases.

描述（由申请人提供）：该研究计划的广泛，长期目标是开发一般技术，以有条件地调节蛋白质分子水平的蛋白质功能，而不是针对编码蛋白质蛋白质的DNA或mRNA前体。该技术对靶向蛋白质高度特异，并使用可渗透细胞的小分子对蛋白质功能进行快速可调的控制。目的是设计称为不稳定结构域的小蛋白质结构域，这些结构域在哺乳动物细胞中表达时会迅速而有条件地降解。破坏稳定域的不稳定性忠实地赋予了与这些小领域融合的其他蛋白质。与不稳定结构域紧密结合的可渗透性配体调节其稳定性。不稳定结构域与利益基因的遗传融合确保了特异性，而随之而来的小分子控制则赋予了速度，可逆性和剂量依赖于此方法。迄今为止开发的试剂涉及在没有配体的情况下不稳定并在配体结合时稳定的域。该提案的具体目的旨在提供互补的新域，这些新域是不稳定的，而不是通过添加细胞可渗透的配体稳定。对哺乳动物细胞中如何识别和降解这些域的机械理解将使该技术对用户更有用。这些研究还可能揭示了细胞用来识别和降解未折叠或错误折叠蛋白的一般机制，这些机制可能与重要的人类疾病有关。 公共卫生相关性：该研究计划的目的是开发新的方法，以快速和可逆地调节真核细胞和活哺乳动物中特定蛋白质的稳定性。这种方法将使许多新的人类疾病模型的发展发展，这些模型系统对正在进行的寻找新的和改进的药物来治疗人类疾病非常有帮助。