Sparks D L, Frank P G, Neville T A
Lipoproteins and Atherosclerosis Group, Ottawa Civic Hospital, University of Ottawa Heart Institute H-452, 1053 Carling Avenue, Ottawa, Ont., Canada.
Biochim Biophys Acta. 1998 Feb 16;1390(2):160-72. doi: 10.1016/s0005-2760(97)00172-0.
Characterization of the factors that regulate plasma cholesterol esterification shows that the increased activity of lecithin:cholesterol acyltransferase (LCAT) in the plasma of hyperlipidemic subjects is due to enhanced interactions with a preferred substrate. The details of how the physical properties of high density lipoproteins (HDL) may affect their ability to stimulate cholesterol esterification by LCAT have been investigated in homogeneous reconstituted HDL particles containing two molecules of apolipoprotein (apo) A-I (Lp2A-I) and palmitoyl-oleoyl phosphatidylcholine (POPC). Increasing the POPC or sphingomyelin (SPH) content in an Lp2A-I complex increases particle size and stability but decreases the negative surface charge of apoA-I. Increasing Lp2A-I POPC or SPH content also significantly inhibits cholesterol esterification by LCAT. Increase in the maximum rate of CE production (Vmax) by LCAT is directly related to an increased negative charge on the different Lp2A-I particles and to a reduced amount and stability of amphipathic alpha-helices in apoA-I. In contrast, increasing the Lp2A-I complex negative charge directly by addition of a charged lipid, phosphatidylinositol (PI), has minimal effect on apoA-I conformation and LCAT activation. While variations in Lp2A-I PI content have little effect on the interfacial binding of LCAT, increasing POPC content appears to directly increase the binding affinity of LCAT for the different Lp2A-I particles. These results show that LCAT is stimulated by an apoA-I conformation-dependent increase in negative charge but is less sensitive to electrostatic changes in the lipid interface of discoidal Lp2A-I. The activation of LCAT appears to be dependent on the exposure of both central (residues 98-132) and N-terminal (residues 2-8) domains in apoA-I. A strong relationship between the immunoreactivity of two specific mAbs, 4H1 and A11, and LCAT reactivity suggests that the N-terminus of apoA-I may interact with a central domain in a manner that may regulate the accessibility of LCAT to the edge of the disc. This indicates that the conformation and charge of apoA-I are sensitive to the surface-lipid composition of HDL particles and play a central role in regulating LCAT activation. Since alterations in the surface lipid composition of HDL particles from hyperlipidemic subjects also modify the charge and structure of these particles, this may stimulate the rates of cholesterol esterification by making these lipoproteins preferred LCAT substrates.
对调节血浆胆固醇酯化的因素进行表征表明，高脂血症患者血浆中卵磷脂胆固醇酰基转移酶（LCAT）活性增加是由于与首选底物的相互作用增强。在含有两分子载脂蛋白（apo）A-I（Lp2A-I）和棕榈酰油酰磷脂酰胆碱（POPC）的均匀重构高密度脂蛋白（HDL）颗粒中，研究了HDL的物理性质如何影响其刺激LCAT进行胆固醇酯化的能力。增加Lp2A-I复合物中的POPC或鞘磷脂（SPH）含量会增加颗粒大小和稳定性，但会降低apoA-I的负表面电荷。增加Lp2A-I的POPC或SPH含量也会显著抑制LCAT介导的胆固醇酯化。LCAT胆固醇酯生成最大速率（Vmax）的增加与不同Lp2A-I颗粒上负电荷的增加以及apoA-I中两亲性α-螺旋数量和稳定性的降低直接相关。相反，通过添加带电荷的脂质磷脂酰肌醇（PI）直接增加Lp2A-I复合物的负电荷，对apoA-I构象和LCAT激活的影响最小。虽然Lp2A-I的PI含量变化对LCAT的界面结合影响不大，但增加POPC含量似乎会直接增加LCAT对不同Lp2A-I颗粒的结合亲和力。这些结果表明，LCAT受到apoA-I构象依赖性负电荷增加的刺激，但对盘状Lp2A-I脂质界面的静电变化不太敏感。LCAT的激活似乎取决于apoA-I中中央结构域（98-132位氨基酸残基）和N端结构域（2-8位氨基酸残基）的暴露。两种特异性单克隆抗体4H1和A11的免疫反应性与LCAT反应性之间的强相关性表明，apoA-I的N端可能以某种方式与中央结构域相互作用，从而调节LCAT接近圆盘边缘的可及性。这表明apoA-I的构象和电荷对HDL颗粒的表面脂质组成敏感，并且在调节LCAT激活中起核心作用。由于高脂血症患者HDL颗粒表面脂质组成的改变也会改变这些颗粒的电荷和结构，这可能通过使这些脂蛋白成为首选的LCAT底物来刺激胆固醇酯化速率。
