Cross-linking and lipid efflux properties of ApoA-I mutants suggest direct association between ApoA-I helices and ABCA1
Cross-linking and lipid efflux properties of ApoA-I mutants suggest direct association between ApoA-I helices and ABCA1
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DOI：
10.1021/bi035813p
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发表时间：
2004-02-24
期刊：
影响因子：
2.9
通讯作者：
Zannis, VI
中科院分区：
文献类型：
--
作者：
Chroni, A;Liu, T;Zannis, VI
为了探索apoA-I和ABCA1之间的功能相互作用，我们将几个apoA-I突变体的交联性与它们促进胆固醇外流的能力联系起来。在竞争性交联实验中，apoA-I的氨基末端缺失突变体和双氨基和双端缺失突变体有效地竞争了WT I-125-apoA-I与ABCA1的交联性，而羧基末端缺失突变体apoA-I[Delta(220-243)]竞争较弱。WT apoA-I、apoA-I的氨基末端和双缺失突变体与ABCA1的直接交联组的表观K-d值相似(49-74 nM)，而羧基末端缺失突变体apoA-I[Delta(185-243)]和apoA-I[Delta(220-243)]的表观K-d值增加了3倍。对apoA-I的几个内部缺失和点突变的分析表明，apoA-I[Delta(61-78)]、apoA-I[Delta(89-99)]、apoA-I[Delta(136-143)]、apoA-I[Delta(144-165)]、apoA-I[D102A/D103A]、apoA-I[E125K/E128K/K133E/E139K]、apoA-I[L141R]、apoA-I[R160V/H162A]和WT apoA-I具有相似的AB1介导的脂溢流并且Wt I-125-apoA-I与ABCA1的交联物均能有效竞争。WT apoA-I和ABCA1可以用3A交联剂进行交联。ApoA-I的WT apoA-I、氨基酸、羧基和双缺失突变体与WT ABCA1和突变体ABCA1[W590S]的交联性不同。这一发现与apoA-I螺旋的不同组合与互补的ABCA1结构域直接相关。改变ABCA1/apoA-I关联的突变会影响胆固醇外流并抑制高密度脂蛋白的生物生成。
To explore the functional interactions between apoA-I and ABCA1, we correlated the cross-linking properties of several apoA-I mutants with their ability to promote cholesterol efflux. In a competitive cross-linking assay, amino-terminal deletion and double amino- and carboxy-terminal deletion mutants of apoA-I competed effectively the cross-linking of WT I-125-apoA-I to ABCA1, while the carboxy-terminal deletion mutant apoA-I[Delta(220-243)] competed poorly. Direct cross-linking of WT apoA-I, amino-terminal, and double deletion mutants of apoA-I to ABCA1 showed similar apparent K-d values (49-74 nM), whereas the apparent K-d values of the carboxy-terminal deletion mutants apoA-I[Delta(185-243)] and apoA-I[Delta(220-243)] were increased 3-fold. Analysis of several internal deletions and point mutants of apoA-I showed that apoA-I[Delta(61-78)], apoA-I[Delta(89-99)], apoA-I[Delta(136-143)], apoA-I[Delta(144-165)], apoA-I[D102A/D103A], apoA-I[E125K/E128K/K133E/E139K], apoA-I[L141R], apoA-I[R160V/H162A], and WT apoA-I had similar ABCA1-mediated lipid efflux, and all competed efficiently the cross-linking of WT I-125-apoA-I to ABCA1. WT apoA-I and ABCA1 could be cross-linked with a 3 A cross-linker. The WT apoA-I, amino, carboxy and double deletion mutants of apoA-I showed differences in the cross-linking to WT ABCA1 and the mutant ABCA1[W590S]. The findings are consistent with a direct association of different combinations of apoA-I helices with a complementary ABCA1 domain. Mutations that alter ABCA1/apoA-I association affect cholesterol efflux and inhibit biogenesis of HDL.