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Molecular basis of activation of the prototypic class B G protein-coupled secretin receptor

原型 B 类 G 蛋白偶联促胰液素受体激活的分子基础

基本信息

批准号：
10468293
负责人：
LAURENCE J MILLER
金额：
$ 37.23万
依托单位：
MAYO CLINIC ARIZONA
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-09-01 至 2024-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10468293
关键词：
Address Affect Affinity Agonist Antibodies Anxiety Binding Biochemical Biological Biological Assay Cardiovascular Diseases Cardiovascular Physiology Cholestasis Complement Complex Coupled Cryoelectron Microscopy Cyclic AMP Cysteine Data Dependence Development Diabetes Mellitus Dimerization Distal Docking Epitopes Event Exhibits Extracellular Domain Family member Fluorescence Resonance Energy Transfer Funding Mechanisms G-Protein-Coupled Receptors GTP-Binding Proteins Global Change Goals Grant Heterotrimeric GTP-Binding Proteins Immunologics Investigation Knowledge Length Ligand Binding Ligands Maps Mental Depression Metabolic Metabolic syndrome Methodology Migraine Modeling Molecular Molecular Conformation Mutagenesis N-terminal Nature Obesity Oral Osteoporosis Pathway interactions Peptides Pharmaceutical Preparations Photoaffinity Labels Play Positioning Attribute Production Proteins Pruritus Receptor Activation Receptor Signaling Refractory Resolution Role Secretin Series Signal Pathway Signal Transduction Site Specificity Structure Surface Techniques Testing Therapeutic Variant Work analog antagonist design disulfide bond drug development experience gastrointestinal insight novel particle pharmacophore receptor receptor function receptor structure function secretin receptor small molecule three dimensional structure tool

项目摘要

PROJECT SUMMARY/ABSTRACT Our OVERALL OBJECTIVE is to elucidate the molecular basis of activation of class B GPCRs, using the prototypic secretin receptor (SecR) as a model. Insights will fill key gaps in knowledge and facilitate ultimate development of drugs exhibiting various activity profiles. Class B GPCRs include established targets for treatment of diabetes, obesity, osteoporosis, migraine, anxiety, and depression. However, therapeutics remain suboptimal, and the receptor class has been refractory to development of small molecule, orally active drugs, at least in part, due to lack of understanding of the structure and functional dynamics required for receptor activation. We now have novel, unique insights into both of these, including high-resolution structures of related, full-length, G protein-coupled holoreceptors, that highlight the importance of the interface between the receptor N-terminal extracellular domain (ECD) and the transmembane domain core in ligand binding and receptor activation. Component aims are directed toward understanding the conformational dynamics of this interface for agonist binding and receptor activation, and using our breakthroughs in use of single particle cryo- EM to provide a structural framework for this work. Aim 1, elucidates molecular events involved in secretin peptide engagement with its receptor core, and key determinants for its activity, testing the hypothesis that orientation (and interaction) of ECD and core domains plays a critical role in directing and positioning the orthosteric agonist pharmacophore near its site of action. We will explore this locus using cysteine trapping to compare spatial approximations for analogous inactive and active probes, applied to wild type receptor, as well as dimerization-deficient receptor constructs. Rational structure-activity analysis for binding and a broad range of biological activities of the agonist pharmacophore will also be performed, with results used to provide insights into determinants of activation and effector specificity. Aim 2, investigates the relative orientations and interactions between SecR ECD and core, exploring functional implications of this interrelationship, testing the hypothesis that these domains can interact in various ways that affect states of quiescence and activation. This will be approached by receptor mutagenesis to modify domain interactions, establishment of domain-domain disulfide bonds by incorporating cysteines at the top of the receptor core and bottom of the ECD, as well as using immunologic probes of predicted surfaces of the receptor amino terminus to determine access and to be used in resonance transfer techniques. Aim 3, elucidates SecR inactive and active/holostructure using single particle cryo-EM, testing the hypothesis that mapping of biochemical and functional data onto high resolution 3D structures, including agonist-receptor-heterotrimeric G protein and inactive antagonist-occupied receptor, will help to elucidate the molecular basis for receptor signaling. Together, this work will provide fundamental advances in understanding of class B GPCR structure and function, with insights highly useful in drug development targeting these receptors.

项目总结/文摘

项目成果

期刊论文数量（6）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Secretin Amino-Terminal Structure-Activity Relationships and Complementary Mutagenesis at the Site of Docking to the Secretin Receptor.

促胰液素氨基末端结构-活性关系和与促胰液素受体对接位点的互补诱变。

DOI：
10.1124/molpharm.122.000502
发表时间：
2022
期刊：
Molecular pharmacology
影响因子：
3.6
作者：
Milburn,JulianaE;Harikumar,KaleeckalG;Piper,SarahJ;Raval,Sweta;Christopoulos,Arthur;Wootten,Denise;Sexton,PatrickM;Miller,LaurenceJ
通讯作者：
Miller,LaurenceJ

Targeting VIP and PACAP Receptor Signaling: New Insights into Designing Drugs for the PACAP Subfamily of Receptors.