Walsh M T, Atkinson D
Department of Biophysics, Housman Medical Research Center, Boston University School of Medicine, MA 02118-2394.
J Lipid Res. 1990 Jun;31(6):1051-62.
The unfolding of human apolipoprotein B-100 in its native lipid environment, low density lipoprotein (LDL), and in a soluble, lipid-free complex with sodium deoxycholate (NaDC) has been examined using differential scanning calorimetry (DSC) and near UV circular dichroic (CD) spectroscopy. High resolution DSC shows that LDL undergoes three thermal transitions. The first is reversible and corresponds to the order-disorder transition of the core-located cholesteryl esters (CE) (Tm = 31.1 degrees C, delta H = 0.75 cal/g CE). The second, previously unreported, is reversible with heating up to 65 degrees C (Tm = 57.1 degrees C, delta H = 0.20 cal/g apoB) and coincides with a reversible change in the tertiary structure of apoB as shown by near UV-CD. No alteration in the secondary structure of apoB is observed over this temperature range. The third transition is irreversible (Tm = 73.5 degrees C, delta H = 0.99 cal/g apoB) and coincides with disruption of the LDL particle and denaturation of apoB. The ratio of delta H/delta HvH for the reversible protein-related transition suggests that this is a two-state event that correlates with a change in the overall tertiary structure of the entire apoB molecule. The second protein-related transition is complex and coincides with irreversible denaturation. ApoB solubilized in NaDC undergoes three thermal transitions. The first two are reversible (Tm = 49.7 degrees C, delta H = 1.13 cal/g apoB; Tm = 56.4 degrees C, delta H = 2.55 cal/g apoB, respectively) and coincide with alterations in both secondary and tertiary structure of apoB. The changes in secondary structure reflect an increase in random coil conformation with a concomitant decrease in beta-structure, while the change in tertiary structure suggests that the conformation of the disulfide bonds is altered. The third transition is irreversible (Tm = 66.6 degrees C, delta H = 0.54 cal/g apoB) and coincides with complete denaturation of apoB and disruption of the NaDC micelle. The ratio of delta H/delta HvH for the two reversible transitions indicates that each of these transitions is complex which may suggest that several regions or domains of apoB are involved in each thermal event.
利用差示扫描量热法（DSC）和近紫外圆二色（CD）光谱法，研究了人载脂蛋白B-100在其天然脂质环境低密度脂蛋白（LDL）中，以及与脱氧胆酸钠（NaDC）形成的可溶性无脂复合物中的去折叠情况。高分辨率DSC显示LDL经历了三个热转变。第一个是可逆的，对应于核心位置胆固醇酯（CE）的有序-无序转变（熔点Tm = 31.1℃，焓变ΔH = 0.75 cal/g CE）。第二个热转变此前未被报道，加热至65℃时是可逆的（Tm = 57.1℃，ΔH = 0.20 cal/g载脂蛋白B），并且如近紫外CD所示，与载脂蛋白B三级结构的可逆变化一致。在此温度范围内未观察到载脂蛋白B二级结构的改变。第三个转变是不可逆的（Tm = 73.5℃，ΔH = 0.99 cal/g载脂蛋白B），与LDL颗粒的破坏和载脂蛋白B的变性一致。与可逆的蛋白质相关转变的ΔH/ΔHvH比值表明，这是一个双态事件，与整个载脂蛋白B分子整体三级结构的变化相关。第二个蛋白质相关转变较为复杂，与不可逆变性一致。溶解在NaDC中的载脂蛋白B经历了三个热转变。前两个是可逆的（分别为Tm = 49.7℃，ΔH = 1.13 cal/g载脂蛋白B；Tm = 56.4℃，ΔH = 2.55 cal/g载脂蛋白B），并且与载脂蛋白B二级和三级结构的改变一致。二级结构的变化反映出无规卷曲构象增加，同时β-结构减少，而三级结构的变化表明二硫键的构象发生了改变。第三个转变是不可逆的（Tm = 66.6℃，ΔH = 0.54 cal/g载脂蛋白B），与载脂蛋白B的完全变性和NaDC胶束的破坏一致。两个可逆转变的ΔH/ΔHvH比值表明，这些转变中的每一个都是复杂的，这可能意味着载脂蛋白B的几个区域或结构域参与了每个热事件。
