MOLECULAR BASIS FOR APOLIPOPROTEIN AI FUNCTION

载脂蛋白 AI 功能的分子基础

• 批准号: 6184875
• 负责人: Christie G. Brouillette
• 金额: $ 32.01万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-07-01 至 2003-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6184875
• 关键词: analytical ultracentrifugation; apolipoproteins; chemical binding; conformation; crosslink; fluorescence resonance energy transfer; high density lipoproteins; infrared spectrometry; interferometry; mutant; phosphatidylcholine sterol acyltransferase; physical model; protein structure function; structural biology; thermodynamics

DESCRIPTION (Adapted from abstract): Apolipoprotein A-I (apo A-I) is the major protein found in high density lipoprotein (HDL) an has been implicated in all of the known physiological functions of HDL, including the protective effect of HDL against coronary artery disease and atherosclerosis. One important question in this field is the role that apo A-I structure plays in the conversion of the protein from its lipid-free to lipid-bound conformation(s). This question has been the subject of substantial testing by a variety of biochemical, biophysical, and genetic techniques. The diverse approaches used to address this question have led to the development of several competing models for the structure of apo A-I in its lipid-free and lipid-bound conformations. Rigorous testing of the predictions of these competing models is the focus of this revised application. The research team determined the first atomic resolution for an apo A-I mutant [Borhani, et al 1997]. The information contained in this structure (currently being refined at 3 angstroms) provides a framework for testing the predictions of the various competing models for apo A-I structure. The two specific aims are: 1. What is the physiologically relevant structure of lipid free apo A-I? (a) Is it a globule, a rod or both? (b) Does it have a discrete tertiary structure or is it a "molten globule" with no distinct tertiary structure? 2. What is the conformation of lipid-bound apo A-I? (a) Is it a discrete structure when bound to discoidal lipid complexes (described as either a "picket fence" or a "belt" with defined interhelical pairing), or are there less defined conformations that have no distinct tertiary structure or interhelical pairing? A variety of methods will be employed to test predictions of the various models, such as cysteine crosslinking, pyrene excimer formation, fluorescence resonance energy transfer (FRET), analytical ultracentrifugation, thermodynamic stability, polarized attenuated internal reflection Fourier transform infrared spectroscopy (PATIR-FITR), and lipid binding kinetics. In the long term, these studies will help determine which of the competing models may be physiologically relevant and help us understand lipid and cholesterol metabolism.

描述（改编自摘要）：载脂蛋白 A-I (apo A-I) 是高密度脂蛋白 (HDL) 中发现的主要蛋白质，与 HDL 的所有已知生理功能有关，包括 HDL 对冠状动脉疾病和心血管疾病的保护作用。动脉粥样硬化。这一领域的一个重要问题是apo A-I 结构在蛋白质从无脂构象向脂结合构象转变中所起的作用。这个问题一直是通过各种生物化学、生物物理和遗传技术进行大量测试的主题。用于解决这个问题的多种方法导致了针对 apo A-I 的无脂和脂结合构象结构的几种竞争模型的开发。对这些竞争模型的预测进行严格测试是本次修订后的应用程序的重点。研究小组首次确定了 apo A-I 突变体的原子分辨率 [Borhani, et al 1997]。该结构中包含的信息（目前正在精炼至 3 埃）提供了一个框架，用于测试 apo A-I 结构的各种竞争模型的预测。两个具体目标是： 1. 无脂apo A-I 的生理相关结构是什么？ (a) 它是球状、棒状还是两者兼而有之？ (b) 它是否具有离散的三级结构，还是没有明显三级结构的“熔球”？ 2. 脂质结合的apo A-I的构象是什么？ (a) 当与盘状脂质复合物结合时，它是离散结构（被描述为具有明确的螺旋间配对的“栅栏”或“带”），还是存在没有明显的三级结构或螺旋间配对的不太明确的构象？将采用多种方法来测试各种模型的预测，例如半胱氨酸交联、芘准分子形成、荧光共振能量转移（FRET）、分析超速离心、热力学稳定性、偏振衰减内反射傅立叶变换红外光谱（PATIR-FITR） ）和脂质结合动力学。从长远来看，这些研究将有助于确定哪些竞争模型可能具有生理相关性，并帮助我们了解脂质和胆固醇代谢。