Helix 6 of subdomain III A of human serum albumin is the region primarily photolabeled by ketoprofen, an arylpropionic acid NSAID containing a benzophenone moiety
Helix 6 of subdomain III A of human serum albumin is the region primarily photolabeled by ketoprofen, an arylpropionic acid NSAID containing a benzophenone moiety
复制标题
DOI：
10.1016/s0167-4838(99)00174-0
复制
发表时间：
1999-09-14
期刊：
影响因子：
--
通讯作者：
Otagiri, M
中科院分区：
文献类型：
--
作者：
Chuang, VTG;Kuniyasu, A;Otagiri, M
众所周知，人血清白蛋白(HSA)的亚区III A(Site II)与多种内源性和外源性物质结合。然而，结合部位的微环境的性质仍不清楚。酮洛芬(KP)是一种含有二苯甲酮部分的芳基丙酸类非甾体抗炎药，作为光亲和标记剂标记结合区域。随后对光标记人血清白蛋白的CNBR裂解表明，11.6 kDa和9.4 kDa片段包含了大部分被掺入的放射性。竞争实验表明，该11.6 kDa片段含有与II位点配体的共同结合区。该片段经Achromobacter lyticus Protease I(AP-I)双酶切后，确定其氨基酸序列为XCTESLVNRR，与HSA的476C-485K序列相对应。相应的AP-I酶切人血清白蛋白多肽的氨基酸序列由476到499个残基组成，形成了IIIA亚区的第5和第6个螺旋。HSA-MyR X射线结晶学数据表明，第5螺旋与配体结合的程度最小。对接模型提供了进一步的支持，即螺旋6代表Kp的光标记区域。(C)1999 Elsevier Science B.V.保留所有权利。
It is well known that the subdomain III A (site II) of human serum albumin (HSA) binds a variety of endogenous and exogenous substances. However, the nature of the microenvironment of the binding site remains unclear. Ketoprofen (KP), an arylpropionic acid NSAID which contains a benzophenone moiety, was used as a photoaffinity labeling agent to label the binding region. Subsequent CNBr cleavage of the photolabeled HSA revealed that the 11.6 kDa and 9.4 kDa fragments contained most of the incorporated radioactivity. Competition experiments showed that the 11.6 kDa fragment contains the common binding region for site II ligands. This fragment was redigested with Achromobacter lyticus protease I (AP-I) and the amino acid sequence of the photolabeled peptide was determined to be XCTESLVNRR, which corresponds to the sequence 476C-485K of HSA. The complete amino acid sequence of the corresponding AP-I digested HSA peptide encompasses residues 476 to 499, which form helices 5 and 6 of subdomain III A. The HSA-Myr X-ray crystallography data showed that helix 5 is involved to the least extent in ligand binding. A docking model provided further support that helix 6 represents the photolabeled region of KP. (C) 1999 Elsevier Science B.V. All rights reserved.