IDENTIFICATION OF PROTEINS ASSOCIATED WITH APOLIPOPROTEIN A-I-CONTAINING LIPOPROTEINS PURIFIED BY SELECTED-AFFINITY IMMUNOSORPTION
IDENTIFICATION OF PROTEINS ASSOCIATED WITH APOLIPOPROTEIN A-I-CONTAINING LIPOPROTEINS PURIFIED BY SELECTED-AFFINITY IMMUNOSORPTION
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DOI：
10.1021/bi00174a003
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发表时间：
1994-03-01
期刊：
影响因子：
2.9
通讯作者：
KANE, JP
中科院分区：
文献类型：
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作者：
KUNITAKE, ST;CARILLI, CT;KANE, JP
通过选择性亲和免疫吸附分离含有载脂蛋白 A-I 的脂蛋白 [Lp(A-I)]，可最大程度地减少通过连续超速离心分离高密度脂蛋白 (HDL) 过程中发生的相关蛋白的损失。我们使用二维凝胶电泳分析来分离与 Lp(A-I) 相关的蛋白质。通过结合转印迹蛋白的氨基酸测序和带有特定抗血清的蛋白质印迹，我们鉴定了许多相关蛋白。载脂蛋白 (ape) A-I、A-II、A-IV、C-III、D 和 E 的位置位于凝胶上。离心后，卵磷脂胆固醇酰基转移酶和胆固醇酯转移蛋白与 Lp(A-I) 的关联程度高于与 HDL 的关联程度。除了之前鉴定出的与 HDL 相关的蛋白质外，我们还检测到了许多与 Lp(A-I) 相关的血浆蛋白，即纤维蛋白原、触珠蛋白、富含脯氨酸的蛋白（C4b 结合蛋白）和载脂蛋白 J（SP40,40 硫酸化糖蛋白）。这些蛋白质与 Lp(A-I) 的共分离似乎不是人为因素，因为它们对含有共价结合的预免疫山羊 IgG 代替抗 apoA-I IgG 的假柱具有非常低的亲和力。这些发现表明，除了主要与脂蛋白相关地存在的载脂蛋白之外，还有另一类蛋白质以脂蛋白相关形式和分散状态存在。这些脂蛋白相关蛋白的检测和鉴定可能有助于机械确定大量由 HDL 引起的观察到的功能。
The isolation of apolipoprotein A-I-containing lipoproteins [Lp(A-I)] by selected-affinity immunosorption minimizes the loss of associated proteins that occurs during the isolation of high-density lipoproteins (HDL) by sequential ultracentrifugation. We have used two-dimensional gel electrophoretic analysis to separate the proteins associated with Lp(A-I). Using a combination of amino acid sequencing of transblotted proteins and Western blotting with specific antisera, we have identified a number of associated proteins. The positions of the apolipoproteins (ape) A-I, A-II, A-IV, C-III, D, and E were located on the gels. Lecithin-cholesterol acyltransferase and cholesteryl ester transfer protein were identified in association with Lp(A-I) to a greater extent than found associated with HDL after centrifugation. In addition to those proteins previously identified in association with HDL, we detected a number of plasma proteins associated with Lp(A-I), namely, fibrinogen, haptoglobin, proline-rich protein (C4b-binding protein), and apolipoprotein J (SP40,40 sulfated grycoprotein). The coisolation of these proteins with Lp(A-I) does not appear to be an artifact in that they have very low affinity for a sham column containing covalently bound preimmune goat IgG in place of the anti-apoA-I IgG. These findings suggest that in addition to apolipoproteins that exist largely in association with lipoproteins there is another class of proteins which exist both in lipoprotein- associated form and in the dispersed state. Detection and identification of these lipoprotein-associated proteins may aid in the mechanistic determination of a humber of observed functions attributed to HDL.