Recombinant antibodies for cytoplasmic, nuclear and transmembrane proteins

细胞质、核和跨膜蛋白的重组抗体

• 批准号: 8796585
• 负责人: DONALD B ARNOLD
• 金额: $ 32.93万
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-30 至 2018-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8796585
• 关键词: AMPA Receptors; Affinity; Animals; Binding; Brain; Cell Nucleus; Cells; Cytoplasmic Protein; Cytoskeletal Proteins; Cytoskeleton; DLG4 gene; Epitopes; Event; Feedback; Fibronectins; Genes; Genome; Grant; In Vitro; Integral Membrane Protein; Kinesin; Label; Laboratories; Life; Location; Messenger RNA; Methods; Mitochondria; Molecular; Molecular Motors; Motor; Nervous system structure; Neurons; Nuclear Protein; Nuclear Proteins; Organism; Peptides; Protein Region; Proteins; Recombinant Antibody; Regulation; Reporting; Structure; Synapses; System; Techniques; Time; Transcription Factor AP-1; Transcriptional Regulation; Work; base; density; design; extracellular; gephyrin; in vivo; insight; novel; postsynaptic; protein transport; public health relevance; receptor; reconstitution; research study; trafficking; transcription factor

DESCRIPTION (provided by applicant): The purpose of the experiments proposed in this grant is to generate recombinant antibody-like proteins (intrabodies) that can label specific proteins of all types, including cytoplasmic and nuclear proteins, and extracellular epitopes on transmembrane proteins. Recently we have shown the utility of the intrabody approach by generating probes that recognize the postsynaptic proteins Gephyrin and PSD95. These probes work efficiently and can be used to visualize endogenous proteins in living organisms. However, the design of these intrabodies places constraints on the targets against which they can be made. In particular, target proteins must be fixed to the cytoskeleton of the cell. Here we propose to develop a new strategy for generating intrabodies that does not place any requirements on the type of target protein. To establish the efficacy of this strategy we will generate intrabodies against the motor protein Kinesin 1, the transcription factor AP-1 and an extracellular epitope of the AMPA receptor GluA1.

描述（由申请人提供）：本拨款中提出的实验目的是产生重组抗体样蛋白（体内），可以标记所有类型的特定蛋白，包括细胞质和核蛋白，以及跨膜蛋白上的细胞外表位。最近，我们通过生成识别突触后蛋白Gephyrin和PSD95的探针，展示了体内方法的实用性。这些探针工作效率高，可用于可视化生物体内的内源性蛋白质。然而，这些内体的设计限制了它们可以针对的目标。特别是，靶蛋白必须固定在细胞骨架上。在这里，我们建议开发一种新的策略来产生不需要任何类型的靶蛋白的体内。为了确定这一策略的有效性，我们将产生针对运动蛋白激酶1、转录因子AP-1和AMPA受体GluA1的细胞外表位的体内抗体。