Molecular determinants of the granulocyte-macrophage colony-stimulating factor receptor complex assembly
Molecular determinants of the granulocyte-macrophage colony-stimulating factor receptor complex assembly
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DOI：
10.1074/jbc.274.48.34155
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发表时间：
1999-11-26
影响因子：
4.8
通讯作者：
Hoang, T
中科院分区：
文献类型：
--
作者：
Haman, A;Cadieux, C;Hoang, T
粒细胞巨噬细胞集落刺激因子（GM-CSF）受体（GMR）由属于以生长激素受体为代表的细胞因子受体超家族的两条链组成。在细胞因子受体中发现的常见结构元件是两个纤连蛋白样结构域的模块，每个结构域的特征在于分别表示为A-G和A '-G'的七条β链。alpha-链（GMR α）赋予低亲和力GM-CSF结合（Kd = 1-5 nM），而β链（β（c））本身不结合GM-CSF，但当与α（c）结合时赋予高亲和力结合。（Kd = 40-100 pM），在本研究中，我们通过几种方法的融合定义了配体识别和复合物稳定所需的分子决定簇，包括嵌合受体的构建、我们的GM.GMR复合物的三维模型的分子动力学和定点诱变。然后通过平衡时的配体结合研究和通过测定GM.GMR复合物的动力学常数来研究单个残基的功能重要性。这种三重复合物的关键是建立四个非共价键，三个决定配体识别过程的性质，涉及α链的残基Arg（280）和Tyr（226）和残基Tyr（365）由于这些残基中的任何一个的突变导致缔合速率的显著降低。最后，β（c）的残基Tyr 365起双重作用，因为它与β（c）的另一个残基Tyr（421）合作以稳定复合物，因为Tyr（365）和Tyr 421的突变导致解离速率（Koff）的急剧增加。有趣的是，这四个残基位于GMR α和β（c）的B '-C'和F '-G'环，从而在明显不对称的异二聚体结构内建立功能对称性。
The granulocyte macrophage colony-stimulating factor (GM-CSF) receptor (GMR) is composed of two chains that belong to the superfamily of cytokine receptors typified by the growth hormone receptor, A common structural element found in cytokine receptors is a module of two fibronectin-like domains, each characterized by seven beta-strands denoted A-G and A'-G', respectively. The alpha-chain (GMR alpha) confers low affinity GM-CSF binding (K-d = 1-5 nM), whereas the beta-chain (beta(c)) does not bind GM-CSF by itself but confers high affinity binding when associated with alpha (K-d = 40-100 pM), In the present study, we define the molecular determinants required for ligand recognition and for stabilization of the complex through a convergence of several approaches, including the construction of chimeric receptors, the molecular dynamics of our three-dimensional model of the GM.GMR complex, and site-directed mutagenesis. The functional importance of individual residues was then investigated through ligand binding studies at equilibrium and through determination of the kinetic constants of the GM.GMR complex, Critical to this tripartite complex is the establishment of four noncovalent bonds, three that determine the nature of the ligand recognition process involving residues Arg(280) and Tyr(226) Of the alpha-chain and residue Tyr(365) Of the beta-chain, since mutations of either one of these residues resulted in a significant decrease in the association rate. Finally, residue Tyr365 Of beta(c) serves a dual function in that it cooperates with another residue of beta(c) Tyr(421) to stabilize the complex since mutation of Tyr(365) and Tyr421 result in a drastic increase in the dissociation rate (Koff), Interestingly, these four residues are located at the B'-C' and F'-G' loops of GMR alpha and of beta(c), thus establishing a functional symmetry within an apparently asymmetrical heterodimeric structure.