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Analysis and engineering of MD-2 and related proteins from common allergens

常见过敏原 MD-2 和相关蛋白的分析和改造

基本信息

批准号：
8228053
负责人：
David M. Kranz
金额：
$ 23.01万
依托单位：
UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-02-15 至 2014-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8228053
关键词：
Adverse reactions Affect Affinity Allergens Allergic Reaction Anti-Inflammatory Agents Anti-inflammatory Antigens Asthma B-Lymphocytes Bacterial exotoxin Behavior Binding Biological Assay CD14 gene Cell Wall Cells Cessation of life Cloning Collection Complex Data Disease Dominant-Negative Mutation Endotoxins Engineering Exhibits Frequencies Homologous Gene Homologous Protein Hypersensitivity IgE Immune Immune response Inflammatory Kinetics Lead Libraries Ligands Lipid Binding Lipids Lipopolysaccharides Lung Lung diseases Lymphocyte Antigen 96 Mediating Modeling Molecular Oryctolagus cuniculus Patients Pharmaceutical Preparations Pneumonia Positioning Attribute Protein Analysis Protein Engineering Proteins Reaction Recruitment Activity Septic Shock Shock Source Surface System T-Cell Receptor T-Lymphocyte Technology Testing Therapeutic Toxin Variant Yeasts airway inflammation analog base high throughput screening in vitro testing in vivo inhibitor/antagonist interest member mutant novel prevent protein expression protein purification public health relevance pyroglyphid receptor response success toll-like receptor 4

项目摘要

DESCRIPTION (provided by applicant): Allergens are known to have a direct affect on the induction of airway inflammation and asthma. The immunological mechanisms involved are now beginning to be understood. Recent evidence suggests that some allergens may be particularly potent at eliciting an immune response because the same molecule (allergen) has the ability to bind to innate receptors and serve as a conventional B and T cell antigen. Thus, Derp2 from the dust mite has been shown to behave like the mammalian protein MD-2 in recruiting bacterial lipopolysaccharide (LPS) to stimulate toll-like receptor 4 (TLR4). According to this hypothesis, the presence of Derp2 and a source of LPS (or similar lipid-based compounds), could initiate activation of the innate system, ultimately leading to stimulation of TH cells and IgE allergic reactions and associated pulmonary disease such as asthma. We propose to study the molecular details of MD-2, Derp2, and other related allergens in their binding to LPS by using a high-throughput protein expression and analysis system called yeast display. This technology will also be used for protein engineering in an effort to develop dominant negative inhibitors of MD- 2 that might serve as modulators of such adverse reactions. The specific aims are to: 1) characterize the MD-2 and Derp2 residues important in LPS and TLR4 binding, 2) analyze LPS and TLR4 binding by a panel of allergen homologs of MD-2, and 3) engineer soluble MD-2 analogs that act as dominant negative inhibitors of Derp2 in LPS recruitment. PUBLIC HEALTH RELEVANCE: For most patients with asthma, it is clear that allergies can induce the inflammatory cascade. Recent evidence suggests that some allergens may be particularly potent at eliciting an immune response because the same molecule (allergen) has the ability to bind to innate receptors and serve as a conventional B and T cell antigen. We propose to use high-throughput protein engineering and analysis to understand the molecular basis of the innate interactions, and to develop proteins that could act as specific anti-inflammatory drugs.

描述（由申请人提供）：已知过敏原对诱导气道炎症和哮喘有直接影响。所涉及的免疫学机制现在开始被理解。最近的证据表明，某些过敏原在引发免疫反应方面可能特别有效，因为相同的分子（过敏原）具有与先天受体结合并作为常规B细胞和T细胞抗原的能力。因此，尘螨的Derp2表现出与哺乳动物蛋白MD-2相似的行为，可以募集细菌脂多糖（LPS）来刺激toll样受体4 （TLR4）。根据这一假设，Derp2和LPS（或类似的脂质化合物）的存在可能启动先天系统的激活，最终导致TH细胞和IgE过敏反应的刺激，以及相关的肺部疾病，如哮喘。我们建议使用一种称为酵母显示的高通量蛋白表达和分析系统来研究MD-2， Derp2和其他相关过敏原与LPS结合的分子细节。这项技术也将用于蛋白质工程，努力开发MD- 2的显性负抑制剂，可能作为这些不良反应的调节剂。具体目标是：1)表征在LPS和TLR4结合中重要的MD-2和Derp2残基，2)通过一组MD-2的过敏原同源物分析LPS和TLR4的结合，以及3)设计可溶性MD-2类似物，在LPS募集中作为Derp2的主要负抑制剂。