FUNCTION OF LIPOPROTEIN LIPASE PROTEOGLYCAN INTERACTION

LPL是负责富含甘油三酯的水解的主要酶 脂蛋白 这种酶在决定 血浆脂蛋白的浓度和组成，包括LDL和 HDL组分。 因此，理解LPL的机制方面 调节将有助于对动脉粥样硬化的理解。 在 细胞产生LPL，大部分合成的酶被降解 降解速率决定了酶的净流出量。 当前 该提案侧重于乙酰肝素功能的分子方面 硫酸蛋白聚糖（HSPG）和低密度脂蛋白受体 相关蛋白（LRP）在LPL降解中的作用>据推测， 细胞表面上的LPL结合HSPG和HSPG并被其内化。 LRP。 在脂肪细胞中，HSPG介导的降解占主导地位。 我们将 测试HSPG核心蛋白发挥信号功能的概念 在靶向与LPL结合的HS链中， 输出到细胞外液。 也有人假设，LRP 介导的途径依赖于HSPG结合LPL和LPL的最大通量 还有LRP。 将纯化脂肪细胞HSPG和紧密结合的蛋白质， 克隆和制备抗体。 LPL结合的鉴定 结构域将通过定点诱变、化学诱变和化学诱变来完成。 用单克隆抗体修饰HSPG保护的碱性残基 probes. 通过LRP降解的LPL的分数，和特定的HSPG将是 在脉冲追踪研究中定量测定。 RAP（受体相关 蛋白），X-ch-LPL-3-6a，一种抑制LPL结合的单克隆抗体 HSPG、磷脂酰肌醇特异性磷脂酶C和缺乏HSPG的细胞 HS链将用于选择性抑制LPL途径 降解 野生型或突变型LPL的速率常数（Kon和Koff） 在细胞表面存在或不存在HS链的情况下与LRP结合 将确定评估HSPG在LPL/LRP中的功能 互动

LPL is the major enzyme responsible for the hydrolysis of triglyceride rich lipoproteins. The enzyme plays a major role in determining the concentration and composition of plasma lipoproteins including the LDL and HDL fractions. Therefore, understanding mechanistic aspects of LPL regulation will contribute to an understanding of atherosclerosis. In cells producing LPL a large fraction of the synthesized enzyme is degraded and degradation rate determines the net efflux of enzyme. The current proposal focuses on the molecular aspects of the function of heparan sulfate proteoglycans(HSPGs) and the low density lipoprotein receptor related protein (LRP) in the degradation of LPL> It is hypothesized that LPL on the surface of cells binds and is internalized by both HSPGs and LRP. In adipocytes, the HSPG mediated degradation predominates. We will test the notion that the core proteins of HSPGs plays a signaling function in targeting HS chains to which LPL is bound to endocytic pathways or to export to the extracellular fluid. It is also hypothesized that the LRP mediated pathway is dependent for maximal flux on HSPG binding to both LPL and LRP. Adipocyte HSPGs and tightly associated proteins will be purified, cloned, and antibodies prepared. The identification of the LPL binding domains will be completed by site directed mutagenesis, chemical modification of HSPG protected basic residues and with monoclonal antibody probes. The fraction of LPL degraded via LRP, and specific HSPGs will be quantitatively determined in pulse chase studies. RAP (Receptor-Associated Protein), X-ch-LPL-3-6a, a monoclonal antibody which inhibits LPL binding to HSPGs, phosphatidylinositol specific phospholipase C and cells devoid of HS chains will be utilized to inhibit selectively pathways of LPL degradation. Rate constants (Kon and Koff) of wild type or mutant LPL binding to LRP in the presence or absence of HS chains on cell surfaces will be determined to evaluate the function of HSPGs in the LPL/LR P interaction.