Contribution of the conserved amino acids of the melanocortin-4 receptor in D-[Nle4, Phe7]-α-melanocyte-stimulating hormone binding and signaling
Contribution of the conserved amino acids of the melanocortin-4 receptor in D-[Nle4, Phe7]-α-melanocyte-stimulating hormone binding and signaling
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DOI：
10.1074/jbc.m702285200
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发表时间：
2007-07-27
影响因子：
4.8
通讯作者：
Yang, Yingkui
中科院分区：
文献类型：
--
作者：
Chen, Min;Cai, Minying;Yang, Yingkui
黑素皮质素4受体（Melanocortin 4 receptor, MC4R）在调节食物摄入和体重中起着重要作用。为了确定人类MC4R （hMC4R）负责α -促黑素皮质激素（α - msh）结合的分子基础，本研究中，我们利用受体结构域交换和定点诱变研究来研究hMC4R负责α - msh结合和信号传导的分子决定因素。α - msh对hMC4R有效，但对hMC2R无效。将hMC4R的第二、第三、第四、第五和第六跨膜区（TM）与hMC2R的同源区域进行盒体替换，并检测α - msh结合和信号传导。结果表明，每种嵌合受体均在细胞表面表达，且表达水平与野生型受体相似。hMC4R的第2、第4、第5和第6个TMs被hMC2R的同源区域取代，除了取代hMC4R的TM3外，并没有显著改变α - msh结合的亲和力和效力，这表明hMC4R TMs中的保守残基对α - msh结合和信号传导至关重要。进一步的诱变研究表明，TM2中的Glu（100）、TM3中的Asp（122）、Asp（126）和TM6中的Trp（258）、Phe（261）、His（264）等保守残基参与了α - msh的结合和信号转导。总之，我们的研究结果表明，hMC4R的TM2、TM3和TM6中的保守残基负责α - msh的结合和信号传导。
Melanocortin 4 receptor (MC4R) plays an important role in the regulation of food intake and body weight. To determine the molecular basis of human MC4R (hMC4R) responsible for alpha-melanocortin-stimulating hormone (alpha-MSH) binding, in this study, we utilized both receptor domain exchange and site-directed mutagenesis studies to investigate the molecular determinants of hMC4R responsible for alpha-MSH binding and signaling. alpha-MSH is a potent agonist at hMC4R but not at hMC2R. Cassette substitutions of the second, third, fourth, fifth, and sixth transmembrane regions (TM) of the hMC4R with the homologous regions of hMC2R were performed and alpha-MSH binding and signaling were examined. Our results indicate that each chimeric receptor was expressed at the cell surface and the expression levels remain similar to that of the wild-type receptor. The cassette substitutions of the second, fourth, fifth, and sixth TMs of the hMC4R with homologous regions of the hMC2R did not significantly alter alpha-MSH binding affinity and potency except substitution of the TM3 of the hMC4R, suggesting that the conserved residues in TMs of the hMC4R are crucial for alpha-MSH binding and signaling. Further mutagenesis studies indicate that conserved residues Glu(100) in TM2, Asp(122), Asp(126) in TM3 and Trp(258), Phe(261), His(264) in TM6 are involved in alpha-MSHbinding and signaling. In conclusion, our results suggest that the conserved residues in the TM2, TM3, and TM6 of the hMC4R are responsible for alpha-MSH binding and signaling.