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HUMAN TRANSCRIPTION FACTOR IMMUNOGENS: GENERATION OF A COMPLETE SET

人类转录因子免疫原：全套的生成

基本信息

批准号：
8755145
负责人：
Stephen Anderson
金额：
$ 85万
依托单位：
RUTGERS, THE STATE UNIV OF N.J.
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-09-23 至 2015-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8755145
关键词：
Address Affinity Affinity Chromatography Agreement Antibodies Antigens Archives Arizona Atlases Attention Baculoviruses Binding Binding Proteins Biological Cells Characteristics Chromatin Collection Complex Cytolysis DNA Databases Deposition Disease Documentation Electronics Embryo Ensure Epitopes Escherichia coli Exhibits Future Generations Genes Genetic Germ Cells Goals Human Human Biology Human Genome Project Informatics Insecta Instruction Laboratories Malignant Neoplasms Membrane Metals Mining Monoclonal Antibodies Occupations Plasmids Principal Investigator Procedures Process Production Proteins Proteome Protocols documentation Reagent Recombinant Proteins Recombinants Records Residual state Resources Running Sampling Schedule Scientist Sequence Analysis Shipping Ships Software Tools Structure System Technology Testing Universities Wheat Work Yeasts base chromatin immunoprecipitation cost effective design design and construction experience expression vector kidney cell meetings milligram polypeptide protein expression repository scale up screening success three dimensional structure transcription factor vector wasting

项目摘要

The goal of the proposed project is to assemble a collection of optimized immunogens based on the complete set of human transcription factors. These immunogens will be used by others to create renewable affinity reagents such as monoclonal antibodies. An initial, high priority appiication envisioned for such reagents is chromatin immunoprecipitation studies, and our aim is to design our immunogens so as to optimize them for this purpose. We will do this primarily using two strategies: (a) we will express transcription factor domains that flank the DNA recognition domain but do not overlap it in order to minimize inducing antibodies that might interfere with chromatin binding; and (b) wherever possible we will focus on transcription factor flanking domains that fold into complex three dimensional structures (native folds) and thus present highly specific 3D epitopes. We already have a protein production pipeline up and running that is producing > 1000 purified protein samples (including hundreds of human proteins) per year at the multi- milligram scale, so we are highly experienced in the relevant technologies, and meeting the goals of this FOA will require adding only a modest amount of extra capacity. We also have specialized software tools for the sophisticated analysis of human transcription factor sequences and the design of appropriate expression constructs. Our experience has shown that paying attention to the proper curation of genes and construct design at the beginning pays dividends downstream in terms of timely and cost-effective protein production because it minimizes wasting expensive wet lab resources on sub-par targets. Finally, we have a range of adaptable expression technologies that can be tailored to other classes of human protein targets (including secreted and membrane-bound proteins) in the future, so our approach is highly amenable to expressing additional sectors of the human proteome beyond transcription factors. RELEVANCE (See instmctions): The human genome project gave us a complete blueprint for the genetic instructions that control human biology. This proposal concerns a special set of human proteins known as "transcription factors", which are the master effector.proteins in the cell that have the job of interpreting these instructions and telling the cell what to do under all sets of conditions. In diseases such as cancer these instructions can become garbled, which is why it is important to understand how transcription factors work. This proposal concerns the creation of a specific collection of human transcription factor-derived proteins that will allow scientists to precisely interrogate which transcription factors address which sets of instructions under a variety of conditions, thus establishing the outlines of the master control circuits in human cells.

提出的项目的目标是组装一个优化的免疫原集合基于