权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

PROTEIN INTERACTION CORE

蛋白质相互作用核心

基本信息

批准号：
8389590
负责人：
William Jarvis Britt
金额：
$ 20.02万
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
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/8389590
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 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：在后基因组时代，系统分析揭示了基因产物之间的互连网络。这种网络预测相互作用的蛋白质和将蛋白质分组成簇之间的关系，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将有助于更好地了解神经系统疾病并确定治疗药物的新靶点。几个大型项目已使用高通量酵母二杂种 (Y2H) 分析来识别生物体开放阅读框 (ORES) 的相互作用产物。功能上重要的蛋白质相互作用网络已被定义，并且对于秀丽隐杆线虫而言，研究表明必需蛋白质经常彼此相互作用^^¿^^。类似的分析对于发现中枢神经系统发育障碍的常见机制很有用，正如最近的一项研究所说明的那样，该研究通过 Y2H 和网络分析检查了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致病机制方面的力量是显着的。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。多个基因或遗传位点也与自闭症等复杂疾病有关。连锁分析表明，许多基因位点与自闭症有关，可以想象，与自闭症相关的基因可以相互连接，对相似的细胞通路产生影响 (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，-80% 的疾病相关 SNP 会破坏蛋白质结构的稳定性，从而可能改变与其他蛋白质的相互作用 (2,38,41)。努力了解蛋白质相互作用如何介导复杂的生物过程是 NINDS 支持的 UAB 研究人员研究目标的主要组成部分。核心 C 将促进这些领域的进展。我们建议维护一个蛋白质相互作用核心，该核心可在原核和真核表达系统中提供蛋白质表达，适用于蛋白质体外纯化和体内表达、体外和体内表达的载体设计和选择、蛋白质的生化和成像表征相互作用，包括 FRET 和分裂荧光素酶/GFP、酵母双杂交筛选系统以及用于串联亲和纯化 (TAP) 和质谱的构建体的生成。核心实验室的目标是加速生物学上重要的蛋白质相互作用的识别和表征，而无需在 NINDS 支持的各个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合适的筛选系统，生成用于筛选测定的表达载体构建体，并在筛选程序中提供故障排除和技术援助。这核心实验室还将促进试剂交换，更重要的是，扩大信息和试剂共享资源（www.neurosciencecore.uab.edu/coreclinks.htm），这将允许个人实验室与 UAB 的其他 NINDS 支持的研究人员分享蛋白质表达系统和相互作用蛋白质筛选系统的相关经验。最后，核心实验室将限制这些方法开发中的冗余，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 支持的 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 C) 与 NINDS 支持的 UAB 研究人员相互作用的几个示例。