Probing conformational changes of human serum albumin due to unsaturated fatty acid binding by chemical cross-linking and mass spectrometry
Probing conformational changes of human serum albumin due to unsaturated fatty acid binding by chemical cross-linking and mass spectrometry
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DOI：
10.1042/bj20041624
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发表时间：
2005-05-01
影响因子：
4.1
通讯作者：
Kim, HY
中科院分区：
文献类型：
--
作者：
Huang, BX;Dass, C;Kim, HY
采用化学交联质谱技术研究了人血清白蛋白(HSA)与单不饱和油酸(OA)、多不饱和AA(花生四烯酸)或二十二碳六烯酸(DHA)相互作用时的构象变化。用赖氨酸特异的交联剂修饰无脂肪酸或结合的人血清白蛋白，并用胰酶消化。用纳米电喷雾电离质谱仪对交联肽进行了分析，确定了交联肽的位置。我们的数据表明，当这三种脂肪酸结合时，涉及IIIA亚域的Lys-402或IIIB亚域的Lys-541侧链移动的局部构象发生了变化。我们的数据还表明，当与AA或DHA结合时，Lys-205(IIA)和Lys-466(IIIA)的侧链更靠近彼此，而不是OA，这表明HSA与单不饱和脂肪酸和多不饱和脂肪酸结合时的构象明显不同。虽然这些观察结果与以前的X射线结晶学研究一致，但大多数交联型赖氨酸对的epsilon-氨基之间的距离比晶体结构预测的要短，这可能反映了溶液和晶体结构之间的差异。这种方法可以作为X射线结晶学的有益补充，特别是在探测蛋白质在溶液中的结构方面。
Mass spectrometry with chemical cross-linking was used to probe the conformational changes of HSA (human serum albumin) in solution on interaction with monounsaturated OA (oleic acid) or polyunsaturated AA (arachidonic acid) or DHA (docosahexaenoic acid). Fatty acid-free or -bound HSA was modified with lysine-specific cross-linkers and digested with trypsin. Cross-linked peptides were analysed by nano-electrospray ionization MS to localize the sites of cross-linking. Our data indicated that a local conformational change involving movement of the side chains of Lys-402 of subdomain IIIA or Lys-541 of subdomain IIIB occurred upon binding of all three fatty acids. Our data also indicated that the side chains of Lys-205 (IIA) and Lys-466 (IIIA) moved closer towards each other upon binding AA or DHA, but not OA, suggesting that the conformations of HSA when bound to mono- and poly-unsaturated fatty acids are distinctively different. While these observations agreed with previous X-ray crystallographic studies, the distances between epsilon-amino groups of most cross-linked lysine pairs were shorter than-the crystal structure predicted, possibly reflecting a discrepancy between the solution and crystal structures. This method can serve as a useful complement to X-ray crystallography, particularly in probing the structure of a protein in solution.