Molecular Recognition Studies of the Interleukin-7 Pathway

IL-7 通路的分子识别研究

• 批准号: 8010931
• 负责人: SCOTT T WALSH
• 金额: $ 29.4万
• 依托单位: UNIV OF MARYLAND, COLLEGE PARK
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-01-15 至 2012-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8010931
• 关键词: Address; Affinity; Autoimmune Process; B-Lymphocytes; Bacteriophages; Binding; Biological; Biological Assay; Budgets; Calorimetry; Cell Energetics; Cell Proliferation; Cell Surface Receptors; Cell surface; Cells; Complex; Computers; Contract Services; Coronary Arteriosclerosis; Crystallography; Data; Data Analyses; Deficiency Diseases; Development; Direct Costs; Engineering; Enzyme-Linked Immunosorbent Assay; Epitopes; Event; Experimental Designs; Extracellular Matrix; Family; Funding; Generations; Goals; Heterodimerization; Homeostasis; Human; IL7R gene; Insecta; Interleukin 2 Receptor Gamma; Interleukin 7 Receptor; Interleukin-7; Kinetics; Label; Left; Malignant Neoplasms; Measurement; Medicine; Methods; Modeling; Mus; Mutagenesis; Mutate; NMR Spectroscopy; Ohio; Pathogenesis; Pathway interactions; Peptide N-glycohydrolase F; Peptides; Phage Display; Point Mutation; Principal Investigator; Process Measure; Property; Proteins; Published Comment; Reagent; Reporting; Research; Research Personnel; Research Proposals; Scanning; Severe Combined Immunodeficiency; Signal Transduction; Site; Structure; Sulfhydryl Compounds; Surface Plasmon Resonance; T-Lymphocyte; Techniques; Thermodynamics; Titrations; Universities; Variant; Work; college; cytokine; design; fast protein liquid chromatography; glycosylation; instrument; instrumentation; molecular recognition; mutant; neutravidin; novel; novel therapeutics; programs; public health relevance; receptor; research study; response; three dimensional structure; trend

DESCRIPTION (provided by applicant): Interleukin-7 (IL-7) is an essential pleiotrophic cytokine regulating numerous immunological events. Normal IL-7 signaling responses trigger the development, proliferation, and homeostasis of T and B cells as well as remodeling of the extracellular matrix. IL-7 activates its signaling cascade by interacting with its own cell surface receptor, IL-7R alpha, and the common gamma chain receptor through a cytokine-induced receptor heterodimerization mechanism. Under- and over-stimulation of the IL-7 pathway has been implicated in the pathogenesis of a form of severe combined immunodeficiency, autoimmune conditions, coronary artery disease, and several cancers. Structural, energetic, and cell biological studies of IL-7-induced receptor heterodimerization require a careful analysis of two distinct events: the association of IL-7 to IL-7R alpha and of IL-7:IL-7R alpha complex to the common gamma chain receptor. This research proposal will focus on the initiation step, the binding interaction of IL-7 with its alpha receptor. Results show that glycosylation of the IL- 7R alpha confers optimal binding affinity to IL-7. The importance of glycosylation of the IL-7R alpha appears to be a novel binding recognition mechanism for the common gamma chain receptor family. The thermodynamic and kinetic properties of the wild-type IL-7:IL-7R alpha interaction will be ascertained using a variety of biophysical methods including isothermal titration calorimetry and surface plasmon resonance. The functional epitopes on both IL-7 and IL-7R alpha will be determined using site-directed scanning mutagenesis, biophysical methods, and cell proliferation assays. Three-dimensional structures of free IL-7, free unglycosylated and glycosylated IL-7R alpha, and complexes of IL-7 bound to unglycosylated and glycosylated IL-7R alpha will be determined using x-ray crystallography or NMR spectroscopy. Our understanding of the binding determinants for the IL-7-receptor interaction will be used to engineer novel peptide and protein antagonists using rational protein design and phage display techniques. The long-term goal of this research involves development of a structural energetic model to explain the normal and aberrant IL-7 signaling mechanisms and the generation of new therapeutic peptides/proteins to treat the many deficiencies and diseases associated with impaired IL-7 signaling. Public Health Relevance Statement: This research proposal will focus on the structural, energetic, and cell biological studies of the interaction between human interleukin-7 and its receptor, interleukin-7 receptor alpha. Our understanding of the binding determinants for the IL-7-receptor interaction will be used to engineer novel peptide and protein antagonists using rational protein design and phage display techniques. The long-term goal of this research involves development of a structural energetic model to explain the normal and aberrant IL-7 signaling mechanisms and the generation of new therapeutic peptides/proteins to treat the many deficiencies and diseases associated with impaired human IL-7 signaling.

描述（由申请人提供）：白细胞介素-7（IL-7）是一种调节多种免疫事件的必需多效性细胞因子。正常的IL-7信号传导应答触发T和B细胞的发育、增殖和稳态以及细胞外基质的重塑。IL-7通过与其自身的细胞表面受体IL-7 R α和共同γ链受体相互作用，通过精氨酸诱导的受体异源二聚化机制激活其信号传导级联。IL-7通路的刺激不足和过度与严重联合免疫缺陷、自身免疫性疾病、冠状动脉疾病和几种癌症的发病机制有关。IL-7诱导的受体异二聚化的结构、能量和细胞生物学研究需要仔细分析两个不同的事件：IL-7与IL-7 R α的结合和IL-7：IL-7 R α复合物与常见γ链受体的结合。这项研究计划将集中在起始步骤，IL-7与其α受体的结合相互作用。结果显示IL-7 Ra的糖基化赋予对IL-7的最佳结合亲和力。IL-7 R α糖基化的重要性似乎是常见γ链受体家族的新型结合识别机制。野生型IL-7：IL-7 R α相互作用的热力学和动力学性质将使用多种生物物理方法确定，包括等温滴定量热法和表面等离子体共振。将使用定点扫描诱变、生物物理方法和细胞增殖测定来确定IL-7和IL-7 R α上的功能性表位。游离IL-7、游离非糖基化和糖基化IL-7 R α以及IL-7与非糖基化和糖基化IL-7 R α结合的复合物的三维结构将使用X射线晶体学或NMR光谱法测定。我们对IL-7-受体相互作用的结合决定簇的理解将用于使用合理的蛋白质设计和噬菌体展示技术来设计新的肽和蛋白质拮抗剂。这项研究的长期目标包括开发结构能量模型，以解释正常和异常的IL-7信号传导机制，并产生新的治疗肽/蛋白质，以治疗与IL-7信号传导受损相关的许多缺陷和疾病。 公共卫生相关性声明：这项研究计划将集中在人白细胞介素-7及其受体白细胞介素-7受体α之间相互作用的结构，能量和细胞生物学研究。我们对IL-7-受体相互作用的结合决定簇的理解将用于使用合理的蛋白质设计和噬菌体展示技术来设计新的肽和蛋白质拮抗剂。这项研究的长期目标包括开发结构能量模型，以解释正常和异常的IL-7信号传导机制，并产生新的治疗肽/蛋白质，以治疗与人类IL-7信号传导受损相关的许多缺陷和疾病。