Meyer D F, Nealis A S, Macphee C H, Groot P H, Suckling K E, Bruckdorfer K R, Perkins S J
Department of Biochemistry and Molecular Biology, Royal Free Hospital School of Medicine, London, U.K.
Biochem J. 1996 Oct 1;319 ( Pt 1)(Pt 1):217-27. doi: 10.1042/bj3190217.
Low-density lipoproteins (LDLs) in plasma are constructed from a single molecule of apolipoprotein B-100 (apoB) (M(r) 512,000) in association with lipid [approximate M(r) (2-3) x 10(6)]. LDL oxidation is an important process in the development of atherosclerosis, and can be imitated by the addition of Cu2+ ions. Synchrotron X-ray scattering of LDL yields curves without radiation damage effects at concentrations close to physiological. The radius of gyration RG for preparations of LDL from different donors ranged between 12.1 and 16.0 nm, with a mean of 13.9 nm. At 4 degrees C, the distance distribution curve P(r) indicated a maximum dimension of 25-27 nm for LDL, a peak at 19.5 nm which corresponds to a surface shell of protein and phospholipid head groups in LDL, and submaxima between 1.7 and 13.5 nm, which correspond to an ordered lipid core in LDL. LDL from different donors exhibited distinct P(r) curves. For oxidation studies of LDL by X-rays, data are best obtained at 4 degrees C at a concentration of > or = 2 mg of LDL protein/ml together with controls based on non-oxidized LDL. LDL oxidation (2 mg of apoB/ml) was studied at 37 degrees C in the presence of 6.4, 25.6 and 51.2 mu of Cu2+/g of apoB. Large changes in P(r) were reproducibly observed in the inter-particle distance range between 13 and 16 nm shortly after initiation of oxidation. This corresponds to the phospholipid hydrocarbon in LDL, which has either increased in electron density during oxidation or become increasingly disordered. After 25 h, the structural changes subsequently spread to regions of the P(r) curves assigned to surface apoB and the central core of cholesteryl esters and triacyl-glycerols. Lipid analyses were carried out under the same solution conditions. The alpha-tocopherol and beta-carotene antioxidant contents of LDL were consumed within 1-2 h. Analyses of the formation of thiobarbituric acid-reactive substances and lipid hydroperoxides indicated that arachidonic acid was preferentially oxidized before the maximal formation of lipid hydroperoxides at 8-12 h after initiation of oxidation. High-performance TLC showed that phosphatidylcholine was continuously converted into lysophosphatidylcholine during oxidation, which is consistent with the early changes in the X-ray P(r) curves. The neutral core lipids became modified only after 12-15 h of oxidation. The combination of X-ray scattering structural analyses with biochemical analyses shows that the oxidation of LDL first affects the outer shell of surface phospholipid, then it spreads towards damage of apoB and the internal neutral lipid core of LDL.
血浆中的低密度脂蛋白（LDL）由单个载脂蛋白B - 100（apoB）分子（相对分子质量512,000）与脂质结合构成[相对分子质量约为(2 - 3)×10⁶]。LDL氧化是动脉粥样硬化发展过程中的一个重要过程，可通过添加Cu²⁺离子来模拟。LDL的同步加速器X射线散射在接近生理浓度时产生无辐射损伤效应的曲线。来自不同供体的LDL制剂的回转半径RG在12.1至16.0 nm之间，平均值为13.9 nm。在4℃时，距离分布曲线P(r)表明LDL的最大尺寸为25 - 27 nm，在19.5 nm处有一个峰值，对应于LDL中蛋白质和磷脂头部基团的表面壳层，在1.7至13.5 nm之间有次最大值，对应于LDL中有序的脂质核心。来自不同供体的LDL表现出不同的P(r)曲线。对于通过X射线对LDL进行氧化研究，最好在4℃、LDL蛋白浓度≥2 mg/ml的条件下获取数据，并设置基于未氧化LDL的对照。在37℃、存在6.4、25.6和51.2 μmol Cu²⁺/g apoB的条件下研究了LDL氧化（2 mg apoB/ml）。氧化开始后不久，在13至16 nm的颗粒间距离范围内可重复观察到P(r)的巨大变化。这对应于LDL中的磷脂烃，其在氧化过程中电子密度要么增加，要么变得越来越无序。25小时后，结构变化随后扩展到P(r)曲线中分配给表面apoB以及胆固醇酯和三酰甘油中心核心的区域。在相同溶液条件下进行脂质分析。LDL中的α - 生育酚和β - 胡萝卜素抗氧化剂含量在1 - 2小时内被消耗。硫代巴比妥酸反应性物质和脂质氢过氧化物形成的分析表明，在氧化开始后8 - 12小时脂质氢过氧化物最大形成之前，花生四烯酸优先被氧化。高效薄层层析显示，氧化过程中磷脂酰胆碱持续转化为溶血磷脂酰胆碱，这与X射线P(r)曲线的早期变化一致。中性核心脂质仅在氧化12 - 15小时后才发生改变。X射线散射结构分析与生化分析相结合表明，LDL的氧化首先影响表面磷脂的外壳，然后扩展到对apoB和LDL内部中性脂质核心的损伤。
