C5a receptor activation - Genetic identification of critical residues in four transmembrane helices
C5a receptor activation - Genetic identification of critical residues in four transmembrane helices
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DOI：
10.1074/jbc.274.22.15757
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发表时间：
1999-05-28
影响因子：
4.8
通讯作者：
Bourne, HR
中科院分区：
文献类型：
--
作者：
Baranski, TJ;Herzmark, P;Bourne, HR
激素和感觉刺激激活蛇形受体，将信号传递给异三聚体鸟嘌呤核苷酸结合蛋白（G蛋白）的跨膜开关。为了了解开关机制，我们对人化学引诱剂C5a受体跨膜螺旋III、V、VI和VII中的93个氨基酸进行了随机饱和诱变。酵母选择鉴定出121个功能突变受体，共包含523个氨基酸取代。保守的疏水残基位于螺旋表面，在模拟的七螺旋束中面对其他螺旋（Baldwin， J. M., Schertler， G. F., and Unger， V. M. (1997) J. Mol. Biol. 272, 144-164），而预计与周围脂质接触的表面可以耐受许多取代。我们的分析确定了25个对非保守取代具有抗性的氨基酸位置。它们似乎包括受体开关的两个不同组成部分，一个位于或靠近胞外膜界面的表面和一个位于束的细胞质一半的核心簇。121个突变受体中有21个表现出组成活性，这些激活受体的氨基酸取代主要发生在III和VI螺旋上；其他激活突变截断了靠近螺旋VI的细胞外末端的受体。这些结果确定了蛇形受体开关的一般机制的关键要素。
Hormones and sensory stimuli activate serpentine receptors, transmembrane switches that relay signals to heterotrimeric guanine nucleotide-binding proteins (G proteins). To understand the switch mechanism, we subjected 93 amino acids in transmembrane helices III, V, VI, and VII of the human chemoattractant C5a receptor to random saturation mutagenesis. A yeast selection identified 121 functioning mutant receptors, containing a total of 523 amino acid substitutions. Conserved hydrophobic residues are located on helix surfaces that face other helices in a modeled seven-helix bundle (Baldwin, J. M., Schertler, G. F., and Unger, V. M. (1997) J. Mol. Biol. 272, 144-164), whereas surfaces predicted to contact the surrounding lipid tolerate many substitutions. Our analysis identified 25 amino acid positions resistant to nonconservative substitutions. These appear to comprise two distinct components of the receptor switch, a surface at or near the extracellular membrane interface and a core cluster in the cytoplasmic half of the bundle. Twenty-one of the 121 mutant receptors exhibit constitutive activity, Amino acids substitutions in these activated receptors predominate in helices III and VI; other activating mutations truncate the receptor near the extracellular end of helix VI. These results identify key elements of a general mechanism for the serpentine receptor switch.