Amino Acid Side Chains That Define Muscarinic Receptor/G-protein Coupling
Amino Acid Side Chains That Define Muscarinic Receptor/G-protein Coupling
复制标题
定义毒蕈碱受体/G 蛋白偶联的氨基酸侧链
DOI：
--
复制
发表时间：
1996
影响因子：
4.8
通讯作者：
M. Brann
中科院分区：
文献类型：
--
作者：
E. Burstein;T. Spalding;M. Brann
受体第三胞内环中的氨基酸在G蛋白偶联中起关键作用。为了定义它们的结构要求，我们对m5毒蕈碱受体的该环的N-和C-末端区域（分别为Ni 3和C13）进行了随机饱和诱变。(see Burstein，E.美国，Spalding，T.一、Hill-Eubanks，D.，Brann，M. R.（1995）J.Biol.Chem.270，3141-3146和Hill-Eubanks，D.，Burstein，E.美国，Spalding，T.一、Bräuner-Osborne，H.，Brann，M. R.（1996）J.Biol.Chem.271，3058-3065）。在本研究中，我们已经扩展了我们的分析镍3通过构建受体库与所有可能的氨基酸取代的残基，我们以前确定为功能上重要的，并表征其功能表型。许多疏水性取代在Ile和Thr耐受良好，并在两种情况下引起组成型激活，建立疏水性在这些位置是结构上有利的，许多氨基酸侧链与这种结构作用相容。类似地，在Tyr处观察到疏水性和极性但不带电荷的取代，但与Thr的结果相反，Tyr处的大多数取代显著降低了最大响应并增加了卡巴胆碱的EC，表明残基217的特定侧链参与G-蛋白偶联。Arg允许最宽范围的取代的残基测试，但只有碱性残基耐受良好。所有其他取代显着增加（高达100倍）卡巴胆碱的EC，而不显着影响最大反应。这些突变受体的配体结合特性没有显著变化。我们的结论是，Ile和Thr履行的结构作用，形成的G-蛋白偶联口袋的基础，而Tyr和Arg接触G-蛋白通过特定的侧链相互作用。我们建议，G-蛋白被招募到受体的离子相互作用和疏水残基参与激活。
Amino acids in the third intracellular loops of receptors play pivotal roles in G-protein coupling. To define their structural requirements, we have subjected the N- and C-terminal regions of this loop (Ni3 and Ci3, respectively) of the m5 muscarinic receptor to random saturation mutagenesis. (see Burstein, E. S., Spalding, T. A., Hill-Eubanks, D., and Brann, M. R. (1995) J. Biol. Chem. 270, 3141-3146 and Hill-Eubanks, D., Burstein, E. S., Spalding, T. A., Bräuner-Osborne, H., and Brann, M. R.(1996) J. Biol. Chem. 271, 3058-3065). In the present study, we have extended our analysis of Ni3 by constructing libraries of receptors with all possible amino acid substitutions at the residues we previously identified as functionally important and characterizing their functional phenotypes. Numerous hydrophobic substitutions were well tolerated at Ile and Thr and caused constitutive activation in two cases, establishing that hydrophobicity is structurally favored at these positions and that many amino acid side chains are compatible with this structural role. Similarly, hydrophobic and polar, but not charged, substitutions were observed at Tyr, but in contrast to results for Thr, most substitutions at Tyr substantially decreased maximum response and increased the EC for carbachol, demonstrating that the specific side chain of residue 217 participates in G-protein coupling. Arg allowed the widest range of substitutions of the residues tested, but only basic residues were well tolerated. All other substitutions significantly increased (up to 100-fold) the EC for carbachol without significantly affecting maximal response. There were no significant changes in the ligand binding properties of these mutant receptors. We conclude that Ile and Thr fulfill a structural role, forming the foundation of the G-protein-coupling pocket, whereas Tyr and Arg contact G-proteins through specific side chain interactions. We propose that G-proteins are recruited to receptors by ionic interactions and that hydrophobic residues participate in activation.
登录
查看更多内容
DOI：
--
发表时间：
1995-12
期刊：
The Journal of pharmacology and experimental therapeutics
影响因子：
--
作者：
T. Spalding;E. Burstein;H. Bráuner-Osborne;D. Hill-Eubanks;M. Brann
通讯作者：
T. Spalding;E. Burstein;H. Bráuner-Osborne;D. Hill-Eubanks;M. Brann
嵌合毒蕈碱胆碱能受体：响应毒蕈碱激动剂而激活 Gs 的 β-肾上腺素能受体。
DOI：
--
发表时间：
1990
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Wong,SK;Parker,EM;Ross,EM
通讯作者：
Ross,EM
Hm1 毒蕈碱胆碱能受体内化需要第三个细胞质环中的结构域。
DOI：
--
发表时间：
1992
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Lameh,J;Philip,M;Sharma,YK;Moro,O;Ramachandran,J;Sadée,W
通讯作者：
Sadée,W
小鼠 m1 毒蕈碱乙酰胆碱受体的删除分析：对磷酸肌醇代谢和下调的影响。
影响因子：
2.9
作者：
Shapiro,RA;Nathanson,NM
通讯作者：
Nathanson,NM
通过在培养皿中与 β-半乳糖苷酶底物一起孵育细胞来进行 DNA 转染的简单测定。
影响因子：
2.7
作者：
Lim,K;Chae,CB
通讯作者：
Chae,CB