Differences in stability among the human apolipoprotein E isoforms determined by the amino-terminal domain
Differences in stability among the human apolipoprotein E isoforms determined by the amino-terminal domain
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DOI：
10.1021/bi000099m
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发表时间：
2000-09-26
期刊：
影响因子：
2.9
通讯作者：
Weisgraber, KH
中科院分区：
文献类型：
--
作者：
Morrow, JA;Segall, ML;Weisgraber, KH
通过盐酸胍、尿素或加热对人载脂蛋白（apo）E的常见同种型（apoE 2、apoE 3和apoE 4）及其22 kDa和10 kDa片段进行变性，以研究残基112和158处半胱氨酸/精氨酸互换的影响。以前的物理特性的apoE 3建立，apoE含有两个域，10 kDa的羧基末端和22 kDa的氨基末端结构域，独立展开，并表现出很大的差异，在稳定性。然而，之前尚未比较apoE 2、apoE 3和apoE 4的物理性质。圆二色谱分析表明，不同的异构体具有相同的Ct-螺旋内容和盐酸胍变性证实，这两个域展开独立的所有三个异构体。然而，盐酸胍、尿素和热变性显示apoE亚型的22 kDa氨基末端片段之间的稳定性差异（apoE 4 <apoE 3 <apoE 2）。此外，盐酸胍变性监测圆二色性和荧光建议存在的折叠中间体中的载脂蛋白E，最显着的载脂蛋白E4。因此，这些研究表明，apoE的主要亚型，这是与不同的病理后果，表现出显着差异的稳定性。
Denaturation by guanidine-HCl, urea, or heating was performed on the common isoforms of human apolipoprotein (apo) E (apoE2, apoE3, and apoE4) and their 22-kDa and 10-kDa fragments in order to investigate the effects of the cysteine/arginine interchanges at residues 112 and 158. Previous physical characterization of apoE3 established that apoE contains two domains, the 10-kDa carboxyl-terminal and 22-kDa amino-terminal domains, which unfold independently and exhibit large differences in stability. However, the physical properties of apoE2, apoE3, and apoE4 have not been compared before. Analysis by circular dichroism showed that the different isoforms have identical ct-helical contents and guanidine-HCl denaturation confirmed that the two domains unfold independently in all three isoforms. However, guanidine-HCl, urea, and thermal denaturation showed differences in stability among the 22-kDa amino-terminal fragments of the apoE isoforms (apoE4 < apoE3 < apoE2). Furthermore, guanidine-HCl denaturation monitored by circular dichroism and fluorescence suggested the presence of a folding intermediate in apoE, most prominently in apoE4. Thus, these studies reveal that the major isoforms of apoE, which are associated with different pathological consequences, exhibit significant differences in stability.