Structure-based analysis of GPCR function:: Conformational adaptation of both agonist and receptor upon leukotriene B4 binding to recombinant BLT1
Structure-based analysis of GPCR function:: Conformational adaptation of both agonist and receptor upon leukotriene B4 binding to recombinant BLT1
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DOI：
10.1016/s0022-2836(03)00438-8
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发表时间：
2003-06-13
影响因子：
5.6
通讯作者：
Parello, J
中科院分区：
文献类型：
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作者：
Baneres, JL;Martin, A;Parello, J
我们在大肠杆菌中产生了人白三烯B-4（LTB 4）受体BLT 1，一种G蛋白偶联受体，其产量足以用于该受体在溶液中的第一次结构表征。通过密码子优化和寻找从包涵体中回收的BLT 1的最佳复性条件实现了BLT 1的过表达。洗涤剂增溶的受体显示出与七个跨膜（TM）结构域相容的3D折叠，其中约50% α-螺旋和一个必需的二硫键（圆二色性证据）;与LTB 4结合，K-a = 7.8（+/-0.2）× 10（8）M-1，化学计量比为0.98（+/-0.02）。使用与LTB 4具有共同结构特征的合成分子研究拮抗作用。我们报告的证据表明，这两个合作伙伴，LTB 4和BLT 1，经历了一个重排后，其各自的构象复杂的形成：（i）从平面性的LTB，共轭三烯部分的偏离;（ii）在Trp 234（TM-VI螺旋）的环境中的变化，并在该跨膜螺旋的细胞质区域的暴露。(C)2003爱思唯尔科技有限公司版权所有。
We produced the human leukotriene B-4 (LTB4) receptor BLT1, a G-protein-coupled receptor, in Escherichia coli with yields that are sufficient for the first structural characterization of this receptor in solution. Overexpression was achieved through codon optimization and the search for optimal refolding conditions of BLT1 recovered from inclusion bodies. The detergent-solubilized receptor displays a 3D-fold compatible with a seven transmembrane (TM) domain with ca. 50% alpha-helix and an essential disulfide bridge (circular dichroism evidence); it binds LTB4 With K-a = 7.8(+/-0.2) X 10(8) M-1 and a stoichiometric ratio of 0.98(+/-0.02). Antagonistic effects were investigated using a synthetic molecule that shares common structural features with LTB4. We report evidence that both partners, LTB4 and BLT1, undergo a rearrangement of their respective conformations upon complex formation: (i) a departure from planarity of the LTB, conjugated triene moiety; (ii) a change in the environment of Trp234 (TM-VI helix) and in the exposure of the cytoplasmic region of this transmembrane helix. (C) 2003 Elsevier Science Ltd. All rights reserved.