Structure and Function of the LPLA2/LCAT Acyltransferase Family

LPLA2/LCAT 酰基转移酶家族的结构和功能

• 批准号: 9174909
• 负责人: John Tesmer
• 金额: $ 48.34万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-11-04 至 2017-08-13
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9174909
• 关键词: Active Sites; Acyltransferase; Address; Adverse effects; Affect; Alveolar Macrophages; Amiodarone; Anti-Arrhythmia Agents; Apolipoprotein A-I; Atherosclerosis; Autoimmune Diseases; Binding; Biological Assay; Biological Response Modifier Therapy; Catabolism; Catalysis; Cations; Cholesterol; Cholesterol Esters; Clinical; Complex; Cryoelectron Microscopy; Crystallization; DNA Sequence Alteration; Development; Disease; Electron Microscopy; Engineering; Enzymes; Eye diseases; Family; Fishes; Future; Glycerophospholipids; Goals; Hereditary Disease; High Density Lipoproteins; Human; Image; Infusion procedures; Kidney Failure; Knowledge; Lead; Learning; Ligands; Lipase; Lipids; Lipoprotein (a); Lipoprotein Binding; Liposomes; Liver; Maps; Membrane; Modeling; Molecular; Mutation; Negative Staining; Outcome Study; Patients; Pharmaceutical Preparations; Phosphatidylcholine-Sterol O-Acyltransferase; Phospholipase A2; Physiological; Plasma Proteins; Play; Positioning Attribute; Proteins; Pulmonary Surfactants; Reaction; Recombinants; Regulation; Reporting; Resolution; Role; Series; Serum; Site-Directed Mutagenesis; Somatic Mutation; Structure; Surface; Testing; Transacylase; Variant; acute coronary syndrome; analog; ataxia telangiectasia mutated protein; base; crystal-8; design; disease phenotype; enzyme replacement therapy; esterase; familial cholesteryl ester deficiency; flexibility; improved; insight; lecithin cholesterol acyltransferase deficiency; lipid metabolism; lipophilicity; macrophage; particle; peptidomimetics; preclinical study; public health relevance; reconstruction; reverse cholesterol transport; small molecule; theories; therapy design

DESCRIPTION (provided by applicant): Lysosomal phospholipase A2 (LPLA2) plays a major role in lipid degradation and is believed to underlie drug-induced phospholipidosis, which commonly occurs in patients taking cationic lipophilic drugs such as the antiarrhythmic amiodarone. Aberrant LPLA2 activity may also be involved in development of autoimmune disease and atherosclerosis. LPLA2 is 50% identical in sequence to lecithin-cholesterol acyltransferase (LCAT), a key enzyme in reverse cholesterol transport from arterial plaque macrophages via high density lipoproteins (HDL). Genetic mutations in LCAT are responsible for Familial LCAT Deficiency (FLD), a devastating disease characterized by low serum cholesterol ester levels and renal failure. There are no reported atomic models for either LPLA2 or LCAT, which do not have significant homology to other proteins of known structure. Thus, the molecular bases for their substrate selectivity, regulation, and disease phenotypes remain poorly understood. In this proposal, we address this critical gap in knowledge via functional analysis of our new 1.8 � crystal structure of LPLA2, determination of the atomic structure of LCAT, imaging LCAT bound to HDL particles by electron microscopy, mapping somatic mutations known to cause genetic disease, and investigating the structural basis for differences in acyl acceptor selectivity. In support of our aims, we provide multiple high resolution structure of LPLA2 in various ligand states, negative stained images of LCAT-HDL complexes, and a low resolution crystal structure of fully glycosylated LCAT. The expected outcome of these studies is a better mechanistic understanding of a structurally uncharacterized family of eukaryotic enzymes that play key roles in lipid metabolism. Our structural and functional studies will help explain the molecular basis for genetic disease and ultimately assist in the design of improved biotherapeutics and small molecule LCAT activators to treat lipid-related disorders such as atherosclerosis and LCAT deficiency.

描述（由申请人提供）：溶酶体磷脂酶 A2 (LPLA2) 在脂质降解中起主要作用，并被认为是药物诱导的磷脂沉积的基础，这种情况通常发生在服用阳离子亲脂性药物（如抗心律失常药胺碘酮）的患者中。 LPLA2 活性异常也可能与自身免疫性疾病和动脉粥样硬化的发生有关。 LPLA2 与卵磷脂胆固醇酰基转移酶 (LCAT) 的序列有 50% 相同，LCAT 是通过高密度脂蛋白 (HDL) 从动脉斑块巨噬细胞反向转运胆固醇的关键酶。 LCAT 基因突变导致家族性 LCAT 缺乏症 (FLD)，这是一种以低血清胆固醇酯水平和肾功能衰竭为特征的破坏性疾病。 LPLA2 或 LCAT 的原子模型尚未报道，它们与已知结构的其他蛋白质没有显着的同源性。因此，它们的底物选择性、调节和疾病表型的分子基础仍然知之甚少。在本提案中，我们通过对 LPLA2 的新 1.8 晶体结构进行功能分析、确定 LCAT 的原子结构、通过电子显微镜对与 HDL 颗粒结合的 LCAT 进行成像、绘制已知导致遗传疾病的体细胞突变以及研究酰基受体选择性差异的结构基础，解决了这一关键的知识空白。为了支持我们的目标，我们提供了各种配体状态下的 LPLA2 的多个高分辨率结构、LCAT-HDL 复合物的负染色图像以及完全糖基化的 LCAT 的低分辨率晶体结构。这些研究的预期结果是更好地了解在脂质代谢中发挥关键作用的结构上未表征的真核酶家族。我们的结构和功能研究将有助于解释遗传性疾病的分子基础，并最终有助于设计改进的生物治疗药物和小分子 LCAT 激活剂，以治疗脂质相关疾病，例如动脉粥样硬化和 LCAT 缺乏症。