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Novel Molecules as In Vivo Biological Probes

作为体内生物探针的新型分子

基本信息

批准号：
9330164
负责人：
THOMAS James WANDLESS
金额：
$ 32.1万
依托单位：
STANFORD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-01-01 至 2019-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9330164
关键词：
Affinity Alpha Cell Avena sativa Behavior Bilirubin Binding Binding Sites Biological Biological Models Cell physiology Cells Chimeric Proteins DNA Dependence Development Dose Engineering Ensure Equilibrium Eukaryotic Cell Family Fluorescence Genes Genetic Genetic Transcription Goals Human Engineering Ligand Binding Ligands Light Malaria Mammalian Cell Mammals Mediating Methodology Methods Modeling Optical Methods Parasites Permeability Pharmaceutical Preparations Plasmodium falciparum Proteins Quality Control RNA Interference Research Research Personnel Resolution Specificity System Tacrolimus Binding Protein 1A Tacrolimus Binding Proteins Techniques Technology Tertiary Protein Structure Testing Toxoplasma gondii Work Yeasts cofactor design falls human disease improved in vivo interest knock-down mRNA Precursor misfolded protein multicatalytic endopeptidase complex mutant novel overexpression programs protein aminoacid sequence protein degradation protein function public health relevance small molecule

项目摘要

DESCRIPTION: 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 or non-toxic light. The goal is to engineer small protein domains called destabilizing domains that are rapidly degraded when expressed in mammalian cells. The instability of destabilizing domains is faithfully conferred to other proteins fused to these small domains, allowing researchers to predictably control the levels of any protein-of-interest. One specific aim of this research program will result in new destabilizing domains that are intrinsically fluorescent, allowing researchers to quantify protein levels using optical methods. A second aim of this research is to produce destabilizing domains that are regulated by blue light rather than small molecules, thus enabling the control of protein stability with high spatial resolution. A third aim of these studies will provide a family of destabilizing domains whose cellular levels can be raised by treatment with one ligand or whose cellular levels can be made to fall by treatment with a different ligand. In this case a single destabilizin domain could be used to test the effects of overexpression with one ligand or strong knock-down with a different ligand in an isogenic background. The fourth aim of this research program is to produce destabilizing domains for use in Apicomplexan parasites such as Plasmodium falciparum and Toxoplasma gondii. 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 human diseases.

描述：本研究计划的广泛、长期目标是开发通用技术，在蛋白质分子水平上有条件地调节蛋白质功能，而不是通过靶向编码感兴趣蛋白质的DNA或mRNA前体。该技术对目标蛋白具有高度特异性，并利用细胞渗透性小分子或无毒光提供对蛋白质功能的快速可调控制。目标是设计一种被称为不稳定结构域的小蛋白质结构域，这种结构域在哺乳动物细胞中表达时会迅速降解。不稳定结构域的不稳定性忠实地传递给其他蛋白质