IN VITRO AND IN VIVO STRUCTURE/FUNCTION ANALYSIS OF LPL AND HL

LPL和HL的体外和体内结构/功能分析

• 批准号: 5203517
• 负责人: S SANTAMARINA-FOJO
• 金额: --
• 依托单位: NATIONAL HEART, LUNG, AND BLOOD INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/5203517
• 关键词: Adenoviridae; carboxylic ester hydrolases; chimeric proteins; enzyme activity; enzyme mechanism; enzyme structure; enzyme substrate; enzyme substrate complex; laboratory mouse; lipid metabolism; lipoprotein lipase; phospholipids; protein structure function; recombinant virus; triglycerides

Hepatic lipase (HL) and lipoprotein lipase (LPL) are endothelial-bound lipolytic enzymes that play a major role in lipid metabolism by hydrolyzing triglycerides and phospholipids present in circulating plasma lipoproteins. Despite their similar organization and structure, the physiologic role that HL and LPL play in the metabolism of triglyceride-rich particles and HDL are clearly distinct, a difference that may in part be mediated by the different substrate specificities of the two enzymes. Compared to LPL, HL is the more active phospholipase and this enhanced phospholipase activity may in fact, play a major role in the ability of HL, as opposed to LPL, to modulate HDL metabolism. In order to investigate the structural basis for the different phospholipase activities between LPL and HL we have generated mutant lipases in which the HL and LPL lids, which modulates access of lipids substrates to the active sites, are exchanged. To perform these studies in vivo, we have expressed native and mutant lipases in HL-deficient mice with increased plasma phospholipids using recombinant adenoviruses. Thus, adenovirus expressing native LPL and HL as well as chimeric lipases containing either the HL backbone with the LPL lid or the LPL backbone with the HL lid were injected in a total of 16 HL-deficient mice. Animals injected with viruses expressing lipases containing HL lid had a dramatic decrease (80%) in plasma phospholipids whereas the reduction in mice injected with lipases containing the LPL lid was only 30%. Thus, regardless of the lipase backbone, the presence of the HL lid markedly enhances in vivo phospholipase activity, indicating that the lipase lid is a major determinant of the relative in vivo phospholipase activities of the two enzymes. The use of recombinant adenovirus to express mutant proteins in vivo provides a powerful new approach for performing structure-function analysis of proteins in vivo.

肝脂肪酶（HL）和脂蛋白脂肪酶（LPL）是内皮结合的