PROTEIN INTERACTION CORE

蛋白质相互作用核心

基本信息

批准号：
8788956
负责人：
William Jarvis Britt
金额：
$ 16.87万
依托单位：
UNIVERSITY OF ALABAMA AT BIRMINGHAM
依托单位国家：
美国
项目类别：
财政年份：
2005
资助国家：
美国
起止时间：
2005-07-01 至 2016-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8788956
关键词：
ATP-Binding Cassette Transporters Adenovirus Vector Adenoviruses Affinity Affinity Chromatography Alleles Alzheimer&apos s Disease Antibodies Area Autistic Disorder Baculovirus Expression System Base Sequence Behavior Disorders Binding Biochemical Biochemical Genetics Biological Assay Biological Process Brain C-terminal CMV promoter Caenorhabditis elegans Calcium Calmodulin Cataloging Catalogs Cell Line Cell Nucleus Cell physiology Cells Central Nervous System Diseases Cerebellar Ataxia Chimeric Proteins Cloning Collaborations Complement Complementary DNA Complex Computer software Consultations Core Facility DNA Sequencing Facility Databases Dementia Detection Development Disease Electroporation Elements Epitopes Fluorescence Resonance Energy Transfer Fluorochrome Fusion Protein Expression Galactosidase Gene Delivery Gene Expression Gene Expression Regulation Gene Transfer Generations Genes Genetic Genetic study Genomics Glioblastoma Goals Housing Human Human Resources Image Immune response In Vitro Individual Institution Instruction Integral Membrane Protein Interphase Cell Laboratories Laboratory Personnel Lead Lentivirus Vector Libraries Life Liposomes Luciferases Maintenance Maps Marketing Mass Spectrum Analysis Mediating Metabolic Diseases Methodology Methods Microscopy Migration Assay Modification National Institute of Neurological Disorders and Stroke Nerve Degeneration Nervous system structure Neurodegenerative Disorders Neurons Neurosciences Normal Cell Online Systems Open Reading Frames Organism Parkinson Disease Pathway interactions Physiological Plasmids Post-Translational Protein Processing Precipitation Preparation Production Promega Protein Fragment Proteins Protocols documentation Publishing Puromycin RNA Interference Reagent Research Research Personnel Resistance Resource Sharing Resources Retroviridae Role Screening procedure Secondary to Services Signal Transduction Site Streptavidin Students Subfamily lentivirinae System Systems Analysis TEV protease Technology Tetanus Helper Peptide Tetracycline Control Tetracyclines Therapeutic Agents Transfection Two-Hybrid System Techniques Ubiquitin Update Vaccinia Vaccinium Variant Venus Viral Viral Genes Virus Yeasts adeno-associated viral vector base biological systems body system cDNA Library cellular imaging cognitive function cryogenics design design and construction disease phenotype enhanced green fluorescent protein experience experimental analysis expression cloning expression vector genetic linkage analysis high throughput screening in vivo interest nervous system disorder network models programs promoter protein expression protein function protein protein interaction protein purification protein structure receptor coupling reconstitution research study screening sensor small hairpin RNA success transcription factor vector web site yeast two hybrid system

项目摘要

DESCRIPTION: In a post-genomic era, systems analyses have revealed interconnected networks between gene products. Such networks predict the relationship between interacting proteins and group proteins into clusters to facilitate definition of protein function. Understanding networks of protein-protein interactions in the brain will undoubtedly lead to a better understanding of neurological diseases and identify new targets for therapeutic agents. Several large scale projects have used high throughput yeast two hybrid (Y2H) analyses to identify interacting products of open reading frames (ORES) of an organism. Functionally important protein interaction networks were defined and, for C elegans, it was shown that essential proteins often interact with one another ^^¿^^. Similar analyses are useful for discovering common mechanisms of disorders of CNS development as illustrated by a recent study that examined a protein interaction network of cerebellar ataxias by Y2H and networking analyses ^^. The power of the Y2H approach in elucidating pathogenic mechanisms of neurodegenerative disorders is remarkable. Many human neurodegenerative, developmental, and cognitive function disorders have been associated with multiple alleles. Multiple genes or genetic loci have also been implicated in complex disorders such as autism ^¿. Linkage analysis suggests that many genetic loci are associated with autism and it is conceivable that the genes associated with autism could interconnect, having effects on similar cellular pathways (13-14,19). In addition, studies of diseases associated with a single gene have also benefited from studies of protein interactions. It is estimated that -80% of disease-associated SNPs destabilize a protein's structure and thereby may alter interactions with other proteins (2,38,41). Efforts to understand how protein interactions mediate complex biological processes are maior components of the research goals of NINDS supported investigators at UAB. Core C will facilitate progress in these areas. We propose to maintain a Protein Interaction Core that offers expression of proteins in both prokaryotic and eukaryotic expression systems suitable for protein purification in-vitro and expression in-vivo, vector design and selection for in-vitro and in-vivo expression, biochemical and imaging characterization of protein interactions including FRET and split luciferase/GFP, yeast two-hybrid screening systems and generation of constructs for tandem affinity purification (TAP) and mass spectroscopy. The goal of the Core Laboratory is to accelerate identification and characterization of biologically important protein interactions without the need to establish these methodologies in individual NINDS supported laboratories. The Core Laboratory personnel will assist investigators in selection of appropriate screening systems, generate expression vector constructs for screening assays, and provide troubleshooting and technical assistance in the screening procedures. The Core Laboratory will also facilitate the exchange of reagents and more importantly, expand an information and reagent sharing resource (www.neurosciencecore.uab.edu/coreclinks.htm ) that will permit individual laboratories to share relevant experience with protein expression systems and interacting protein screening systems with other NINDS supported investigators at UAB. Finally, the core laboratory will limit redundancy in development of these methodologies and permit more efficient use of the resources provided by NINDS to investigate protein function in complex biologic systems that are the focus of study by NINDS supported investigators at UAB. Table 1 provides several examples of the interaction of the Protein Interaction Core Core C) with NINDS supported UAB investigators.

描述：在基因组后时代，系统分析揭示了基因产品之间的互连网络。这样的网络预测了相互作用的蛋白质与组蛋白之间的关系到簇中，以促进蛋白质功能的定义。了解大脑中蛋白质 - 蛋白质相互作用的网络无疑会更好地理解神经系统疾病并确定治疗剂的新靶标。几个大型项目使用了高通量酵母两种混合（Y2H）分析来识别有机体开放阅读框（矿石）的相互作用产品。定义了功能上重要的蛋白质相互作用网络，对于秀丽隐杆线虫，结果表明，必需蛋白通常相互作用^^。^^。类似的分析对于发现CNS发育疾病的常见机制很有用，如最近的一项研究所示，该研究检查了通过Y2H和网络分析的小脑共济失调的蛋白质相互作用网络^^。 Y2H方法在阐明神经退行性疾病的致病机制方面的功能是显着的。许多人类神经退行性，发育和认知功能障碍与多个等位基因有关。在自闭症等复杂疾病中，也暗示了多个基因或遗传位置。链接分析表明，许多遗传局部与自闭症有关，可以想象与自闭症相关的基因可能会互连，对相似的细胞途径产生影响（13-14,19）。此外，与单个基因相关的疾病的研究也受益于蛋白质相互作用的研究。据估计，与疾病相关的SNP中的-80％破坏了蛋白质的结构，因此可能会改变与其他蛋白质的相互作用（2,38,41）。了解蛋白质相互作用如何介导复杂的生物学过程的努力是NINDS研究目标的Maior组成部分。核心C将促进这些领域的进步。我们建议维持蛋白质相互作用核心，该核心在原核和真核表达系统中提供蛋白质的表达包括FRET和分裂荧光素酶/GFP的相互作用，酵母的两杂化筛选系统以及串联亲和纯度纯化（TAP）和质谱的结构的产生。核心实验室的目的是加速和表征生物学上重要的蛋白质相互作用，而无需在各个ninds支持实验室中建立这些方法。核心实验室人员将协助研究人员选择适当的筛选系统，生成表达矢量构造以进行筛选测定，并在筛选程序中提供故障排除和技术帮助。核心实验室还将促进试剂的交换，更重要的是，扩展信息和试剂共享资源（www.neurosciencecore.uab.edu/coreclinks.htm）实验室与蛋白质表达系统共享相关经验，并与其他NINDS支持UAB的研究人员相互作用的蛋白质筛选系统。最后，核心实验室将限制这些方法开发的冗余，并允许更有效地利用NINDS提供的资源来研究复杂生物系统中的蛋白质功能，这是NINDS支持UAB研究人员的研究重点。表1提供了蛋白质相互作用核心核心与NINDS支持的UAB研究者相互作用的几个示例。