Structural analysis of apolipoprotein A-I: limited proteolysis of methionine-reduced and -oxidized lipid-free and lipid-bound human apo A-I.
Structural analysis of apolipoprotein A-I: limited proteolysis of methionine-reduced and -oxidized lipid-free and lipid-bound human apo A-I.
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载脂蛋白 A-I 的结构分析：甲硫氨酸还原和氧化的无脂和脂结合人载脂蛋白 A-I 的有限蛋白水解。
DOI：
10.1021/bi962952g
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发表时间：
1997
期刊：
影响因子：
--
通讯作者：
Brouillette,CG
中科院分区：
文献类型：
--
作者：
Roberts,LM;Ray,MJ;Shih,TW;Hayden,E;Reader,MM;Brouillette,CG
用限制性蛋白水解法分析了含有还原和氧化蛋氨酸的脱脂和脂结合载脂蛋白A-I(apo A-I)的结构。无脂的载脂蛋白A-I主要在极端的羧基末端被切割，在蛋白质的中心区域115和136之间的程度要小得多。甲硫氨酸112和148分别氧化为可能的两亲性螺旋4(P99-E120)和6(P143-A164)中的相应亚硫醚，导致螺旋1(L44-G65)、2(P66-S87)和7(P165-G186)变得容易被蛋白酶消化。这些结果与由两亲螺旋1、2、4、6和7最低限度形成的无脂蛋白的离散球状三级结构是一致的。与无脂的apo A-I形成鲜明对比的是，脂结合的apo A-I最容易在极端的氨基末端断裂，其次是在中央和羧基末端。观察到的还原的脂结合蛋白的切割模式支持通过脂结合蛋白的序列分析预测的螺旋之间存在许多转折。蛋氨酸氧化脂质结合蛋白导致极端氨基末端的蛋白酶敏感性降低，而伴随而来的是中央和羧基末端的蛋白酶敏感性增加。蛋氨酸氧化的结果表明，蛋白质的氧化状态是决定脱脂和结合载脂蛋白A-I构象的重要决定因素。
The domain structures of lipid-free and lipid-bound apolipoprotein A-I (apo A-I) containing reduced and oxidized methionines were analyzed by limited proteolysis. Lipid-free apo A-I is cleaved primarily in the extreme carboxy-terminus and, to a much lesser extent, in the central region of the protein between residues 115 and 136. Oxidation of methionines 112 and 148 to the corresponding sulfoxides in putative amphipathic helices 4 (P99-E120) and 6 (P143-A164), respectively, causes helices 1 (L44-G65), 2 (P66-S87), and 7 (P165-G186) to become susceptible to protease digestion. These results are consistent with a discrete, globular tertiary structure for the lipid-free protein minimally formed from amphipathic helices 1, 2, 4, 6, and 7. In distinct contrast to lipid-free apo A-I, lipid-bound apo A-I is most susceptible to cleavage in the extreme amino-terminus and, to a lesser extent, in both the central and carboxy-terminal regions. The observed cleavage pattern for the reduced lipid-bound protein supports the existence of many of the turns between helices predicted by sequence analysis of the lipid-bound protein. Methionine oxidation of lipid-bound protein results in a decreased protease susceptibility in the extreme amino-terminus and a concomitant increase in protease susceptibility in the central and carboxy-terminal regions. The results from methionine oxidation indicate the oxidation state of the protein is an important determinant in defining the conformation of both lipid-free and lipid-bound apo A-I.