Structural analysis of BNIP3 transmembrane interactions

BNIP3跨膜相互作用的结构分析

基本信息

批准号：
7471442
负责人：
Kevin R MacKenzie
金额：
$ 24.19万
依托单位：
RICE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2004
资助国家：
美国
起止时间：
2004-08-01 至 2010-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7471442
关键词：
Address Adenoviruses Apoptosis Apoptosis Regulator Apoptotic Binding Biochemical Biological Biological Assay C-terminal Caenorhabditis elegans Cell Death Cellular biology Complex Condition Data Detergents Development Dimerization Drug Delivery Systems Exhibits Goals Homo Homodimerization Homologous Gene Hypoxia Ischemia Lateral Length Malignant Neoplasms Mammals Measures Mediating Membrane Membrane Proteins Micelles Mitochondria Mutagenesis Mutation NMR Spectroscopy Neurodegenerative Disorders Organ Orthologous Gene Prevention Principal Investigator Proteins Range Regulation Research Research Personnel Role Role playing therapy Signal Transduction Solutions Specificity Structure Tail Transmembrane Domain Work Yeasts analytical ultracentrifugation base dimer fly genetic regulatory protein in vivo insight man member mutant novel programs protein folding protein function protein protein interaction protein structure research study

项目摘要

DESCRIPTION (provided by applicant): The long range goals of this research are to determine how the transmembrane domain sequences of "BH3-only"apoptotic regulatory proteins result in structural and functional interactions that give rise to programmed cell death. More than a dozen members of the 'BH3-only' subclass of the Bcl-2 superfamily provide cell- and signal-specific inputs to mitochondria-mediated apoptosis in mammals, but the biochemical and structural basis for this control is poorly understood. This project focuses on BNIP3 (Bcl-2/adenovirus E1B 19-kDa protein-interacting protein 3), a "BH3-only" protein that is functionally conserved from C. elegans to man. The transmembrane (TM) domain of BNIP3 is implicated in the pro-apoptotic function of the protein, in homodimerization, and in interactions with the anti-apoptotic protein Bcl-2. This research will determine the sequence specificity and structural basis for these TM domain interactions and will determine how these lateral associations within membranes contribute to the regulation of mitochondria-mediated apoptosis. Elucidating the mechanism of action of these proteins will have implications for our understanding of development, cancer, and neurodegenerative diseases. In the next five years, this proposed research will determine the structural basis for homo- and hetero-oligomerization of the hydrophobic C-terminal transmembrane (TM) domain of the mammalian "BH3-only" protein BNIP3 and its orthologs in C. elegans and D. melanogaster. Saturation mutagenesis experiments will define the sequence requirements for these associations in biological membranes and in detergents, and the structures of BNIP3 homodimers and heteromeric complexes will be determined using solution NMR spectroscopy. Interpretation of the mutagenesis data in the context of structures will elucidate the physical basis for the stability and specificity of protein-protein interactions inside membranes. Measuring the apoptogenic effects of wild type and mutant full-length BNIP3 constructs in vivo will determine the functional role(s) played by TM-TM interactions. The results will reveal fundamental rules of membrane protein folding, stimulate new types of experiments by cell biology researchers, and may identify novel drug targets for prevention of apoptosis-related cell death and organ damage such as hypoxia-induced apoptosis following ischemia.

描述（由申请人提供）：本研究的长期目标是确定“仅 BH3”凋亡调节蛋白的跨膜结构域序列如何导致结构和功能相互作用，从而引起程序性细胞死亡。 Bcl-2 超家族“仅 BH3”亚类的十多个成员为哺乳动物中线粒体介导的细胞凋亡提供细胞和信号特异性输入，但这种控制的生化和结构基础知之甚少。该项目重点关注 BNIP3（Bcl-2/腺病毒 E1B 19-kDa 蛋白相互作用蛋白 3），这是一种从线虫到人类功能保守的“仅 BH3”蛋白。 BNIP3 的跨膜 (TM) 结构域与该蛋白的促凋亡功能、同源二聚化以及与抗凋亡蛋白 Bcl-2 的相互作用有关。这项研究将确定这些 TM 结构域相互作用的序列特异性和结构基础，并将确定膜内的这些横向关联如何促进线粒体介导的细胞凋亡的调节。阐明这些蛋白质的作用机制将对我们理解发育、癌症和神经退行性疾病产生影响。在接下来的五年中，这项拟议的研究将确定哺乳动物“仅 BH3”蛋白 BNIP3 及其在秀丽隐杆线虫和D.黑腹果蝇。饱和诱变实验将确定生物膜和洗涤剂中这些关联的序列要求，并且 BNIP3 同二聚体和异聚复合物的结构将使用溶液 NMR 光谱法确定。在结构背景下解释诱变数据将阐明膜内蛋白质-蛋白质相互作用的稳定性和特异性的物理基础。测量体内野生型和突变型全长 BNIP3 构建体的凋亡效应将确定 TM-TM 相互作用所发挥的功能作用。这些结果将揭示膜蛋白折叠的基本规则，刺激细胞生物学研究人员开展新型实验，并可能确定新的药物靶点，以预防与细胞凋亡相关的细胞死亡和器官损伤，例如缺血后缺氧诱导的细胞凋亡。