Membrane signaling protein modulation by lipids and drugs

脂质和药物对膜信号蛋白的调节

• 批准号: 10592319
• 负责人: Daniel Rosenbaum
• 金额: $ 40.68万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-01 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10592319
• 关键词: Address; Architecture; Asthma; Binding; Binding Sites; Biological; Biophysics; CNR1 gene; Cardiovascular Diseases; Cholesterol; Cholesterol Homeostasis; Complex; Cryoelectron Microscopy; Data; Disease; Drug Modulation; Endoplasmic Reticulum; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; GTP-Binding Proteins; Goals; Hormones; Integral Membrane Protein; Length; Ligand Binding; Lipids; Membrane; Membrane Proteins; Molecular Conformation; NMR Spectroscopy; Neurotransmitters; Parkinson Disease; Pathway interactions; Pharmaceutical Preparations; Phospholipids; Physiological; Protein Conformation; Public Health; Regulation; Roentgen Rays; Signal Transduction; Signaling Protein; Structure; Surface; System; Transcriptional Regulation; atomic data; biophysical techniques; fatty liver disease; hormonal signals; nanodisk; therapeutic target; transcription factor

Project Summary/Abstract The goal of this project is to understand at the atomic level how cholesterol modulates integral membrane protein signaling. We seek to address this question in two physiologically important biological pathways, cholesterol homeostasis by the SREBP regulatory machinery and hormone signaling by G protein-coupled receptors (GPCRs). The SREBP machinery centers on Scap, a membrane protein that resides in the endoplasmic reticulum. Scap binds to the SREBP transcription factors and controls their maturation in a cholesterol-dependent manner. There are currently no atomic structures of any vertebrate Scap, and little is known at a detailed biophysical level about Scap's cholesterol binding or conformational changes. Like Scap, GPCRs bind ligands (e.g. hormones, neurotransmitters, and lipids) and undergo conformational changes that trigger intracellular signaling. Multiple GPCR X-ray structures have revealed cholesterol binding interactions, however the functional significance of these interactions is largely unknown. We found that the CB1 cannabinoid receptor can bind allosteric modulators at a membrane-embedded surface that overlaps with its cholesterol binding site. This finding suggests that cholesterol may be an allosteric modulator of CB1 as well as other GPCRs. We are studying the interactions of cholesterol with Scap and GPCRs to better understand how this essential lipid can control diverse functions through these different membrane proteins. We will use cryo-EM to determine the atomic structure of a full-length vertebrate Scap, either bound to its co-regulatory membrane protein Insig or a soluble fragment of SREBP2. These structures will elucidate the architecture of this lipid-sensing machine and provide an atomic basis for cholesterol regulation. In parallel, we will obtain the first NMR spectroscopy data and cryo- EM structural data for GPCRs embedded in native-like phospholipid nanodiscs, with and without cholesterol. These data will reveal whether embedded cholesterol can stabilize particular conformations of a GPCR, and whether cholesterol interactions are maintained in G protein signaling complexes. We propose to use this diverse array of biophysical techniques to obtain unprecedented atomic data on cholesterol regulation of these systems, and our findings may be used to help develop drugs that modulate Scap or different GPCRs through the membrane.

项目总结/摘要 这个项目的目标是在原子水平上了解胆固醇如何调节膜蛋白 发信号。我们试图解决这个问题在两个生理上重要的生物途径，胆固醇 通过SREBP调节机制和通过G蛋白偶联受体的激素信号传导的体内平衡 （GPCR）。SREBP机制以Scap为中心，Scap是一种存在于内质网中的膜蛋白。 网状细胞。Scap与SREBP转录因子结合，并以胆固醇依赖性的方式控制它们的成熟。 方式目前还没有任何脊椎动物Scap的原子结构，并且在详细的结构方面知之甚少。 生物物理水平上的Scap的胆固醇结合或构象变化。像Scap一样，GPCR结合配体 (e.g.激素、神经递质和脂质）并经历构象变化， 发信号。多个GPCR X射线结构揭示了胆固醇结合相互作用，然而，功能性 这些相互作用的重要性在很大程度上是未知的。我们发现CB 1大麻素受体可以结合 在与其胆固醇结合位点重叠的膜包埋表面处的变构调节剂。这 这一发现表明胆固醇可能是CB 1以及其他GPCR的变构调节剂。我们正在研究 胆固醇与Scap和GPCR的相互作用，以更好地了解这种必需的脂质如何控制 通过这些不同的膜蛋白发挥不同的功能。我们会用低温电镜来确定 全长脊椎动物Scap的结构，其结合到其共调节膜蛋白Insig或可溶性 SREBP 2片段。这些结构将阐明这种脂质传感机器的架构，并提供 胆固醇调节的原子基础。同时，我们将获得第一个核磁共振光谱数据和低温- 包埋在天然样磷脂纳米盘中的GPCR的EM结构数据，有和没有胆固醇。 这些数据将揭示嵌入的胆固醇是否可以稳定GPCR的特定构象， 胆固醇相互作用是否维持在G蛋白信号复合物中。我们建议利用这一多样化的 一系列生物物理技术，以获得前所未有的原子数据对胆固醇调节这些系统， 我们的研究结果可用于帮助开发药物，通过调节Scap或不同的GPCR， 膜的