Functional Interplay of Lipid Membrane Components: Activation, Inhibition, and Raft Formation

脂膜成分的功能相互作用：激活、抑制和筏形成

• 批准号: 9382509
• 负责人: Benjamin James Wylie
• 金额: $ 34.6万
• 依托单位: TEXAS TECH UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-10 至 2022-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9382509
• 关键词: Affinity; Alzheimer' s Disease; Antibiotic Resistance; Behavior; Binding Sites; Biological; Biological Assay; Biological Process; Burkholderia pseudomallei; Cardiolipins; Cellular Membrane; Cholesterol; Communication; Disease; Environment; Etiology; GEM gene; HIV; Health; Heart Diseases; Human; Lipid Bilayers; Lipid Binding; Lipids; Membrane; Membrane Lipids; Membrane Microdomains; Membrane Proteins; Nature; Organism; Parkinson Disease; Pathogenesis; Pharmacologic Substance; Phase; Process; Proteins; Research; Resolution; Signal Transduction; Site; Sterols; Structure; Therapeutic; Transport Process; Virulence; improved; interest; membrane assembly; novel; pathogenic bacteria; programs; proteoliposomes; saturated fat; solid state nuclear magnetic resonance

We will determine how the lipid bilayer organizes around membrane proteins to regulate vital biological functions, including signal transduction and molecular transport. Many lipids and membrane proteins associate to form platforms called lipid rafts, which are phase-separated from the surrounding membrane. The dynamic structure and functional importance of these intermediate-sized (5-200 nm), non-crystalline assemblies are difficult to characterize. Many pathogenic bacteria organize lipid rafts which can increase virulence and antibiotic resistance. In humans, rafts form to facilitate multiple signaling processes. These processes are, in turn, involved in the pathogenesis of diseases, including Alzheimer’s, Parkinson’s, and heart disease. Atomic- resolution dynamic structural details of these assemblies will broaden our understanding of signaling processes and inform disease etiology. We will confront this problem using solid-state NMR (SSNMR) and functional assays in proteoliposomes and biological membranes. Our research program is built around three thematic thrusts: (1) To understand how the lipid environment regulates membrane proteins site-specifically. (2) To determine how membrane proteins, in turn, order their environment. (3) To determine the degree of long-range order and dynamic timescales of these membrane assemblies. Our first target is the KirBac1.1 prokaryotic inward-rectifier K+ (Kir) channel and an array of functional lipids, including synthetic lipids and biological lipid extracts, known to associate with rafts. KirBac1.1 shares many behaviors with eukaryotic Kir channels. It is activated by anionic lipids (especially cardiolipin) and has a high affinity for saturated lipids, cholesterol, and other lipid microdomain-forming components (including hopanoids from the native organism Burkholderia Pseudomallei). The shared regulatory and structural features between KirBac1.1 and eukaryotic Kir channels have inspired several topics of interest: (a) How does the lipid cardiolipin maximally activate KirBac1.1 and trigger transmembrane allostery? Cardiolipin is an essential functional lipid throughout nature, and understanding membrane allostery will inform not only the mechanism of K+ conductance, but the means of transmembrane communications. (b) What is the locus and mechanism of cholesterol/hopanoid induced channel activation? Understanding this is key to determining both how sterols regulate proteins and how they contribute to bilayer organization. (c) How do functional lipid binding sites nucleate rafts? Cardiolipin, cholesterol, and hopanoids are all associated with modulating protein activity and membrane organization; our aim is to understand how they create protein-lipid and lipid-lipid interactions in this process. (d) How does the organization of the annular/nonannular lipid shell act as a secondary regulator of membrane proteins? Kir channels are inactivated by cholesterol, but have a high affinity for rafts. How do cellular membranes organize such that Kir channels can be in rafts, yet retain activity? (e) What is the long-range order and lifetime of these assemblies? It is still unknown if these assemblies persist on the timescale of signaling processes.

我们将确定脂质双层如何围绕膜蛋白组织以调节重要的生物学 功能，包括信号转导和分子运输许多脂质和膜蛋白相关。 形成称为脂筏的平台，该平台与周围的膜相分离。 这些中等尺寸（5-20​​0 nm）非晶体组件的结构和功能重要性 许多病原菌组织脂筏，可以增加毒力并降低其特征。 在人类中，筏的形成是为了促进多种信号传导过程。 反过来，参与疾病的发病机制，包括阿尔茨海默病、帕金森病和心脏病。 解析这些组件的动态结构细节将拓宽我们对信号传导的理解 我们将使用固态核磁共振 (SSNMR) 来应对这个问题。 我们的研究计划围绕三个方面展开。 主题要点：（1）了解脂质环境如何对膜蛋白进行位点特异性调节。 (2) 确定膜蛋白如何依次排列其环境。 (3) 确定其环境的程度。 这些膜组件的长程有序和动态时间尺度是 KirBac1.1。 原核内向整流 K+ (Kir) 通道和一系列功能性脂质，包括合成脂质和 已知与筏相关的生物脂质提取物与真核 Kir 具有许多相同的行为。 它被阴离子脂质（尤其是心磷脂）激活，并且对饱和脂质具有高亲和力， 胆固醇和其他脂质微结构域形成成分（包括来自天然生物体的藿香类化合物） KirBac1.1 和真核生物之间共有的调控和结构特征。 Kir 通道激发了几个有趣的话题：（a）脂质心磷脂如何最大程度地激活 KirBac1.1 和触发跨膜变构？心磷脂是自然界中必需的功能性脂质， 了解膜变构不仅可以了解 K+ 电导的机制，还可以了解 K+ 电导的方法 (b) 胆固醇/藿香类物质诱导的位点和机制是什么。 了解这一点是确定甾醇如何调节蛋白质以及它们如何调节的关键 (c) 功能性脂质结合位点如何使心磷脂成核？ 胆固醇和藿香类化合物都与调节蛋白质活性和膜组织有关； 目的是了解它们如何在此过程中产生蛋白质-脂质和脂质-脂质相互作用。 环状/非环状脂质壳的组织充当膜蛋白的二级调节器？ 通道被胆固醇灭活，但对筏具有高亲和力。细胞膜如何组织。 这样 Kir 通道可以位于筏中，但仍保持活动性？ (e) 这些通道的长期顺序和寿命是多少？ 集会吗？这些集会是否持续存在于信号传递过程的时间尺度上仍然未知。