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The role of proline residues in the structure and function of human MT2 melatonin receptor.
Journal of Pineal Research ( IF 10.3 ) Pub Date : 2008-06-12 , DOI: 10.1111/j.1600-079x.2008.00598.x
Petr Mazna 1 , Lenka Grycova , Ales Balik , Hana Zemkova , Eliska Friedlova , Veronika Obsilova , Tomas Obsil , Jan Teisinger
Affiliation  

Melatonin functions as an essential regulator of various physiological processes in all vertebrate species. In mammals, two G protein-coupled melatonin receptors (GPCR) mediate some melatonin's actions: MT1 and MT2. Transmembrane domains (TM) of most GPCRs contain a set of highly conserved proline residues that presumably play important structural and functional roles. As TM segments of MT2 receptor display several interesting differences in expression of specific proline residues compared to other rhodopsin-like receptors (rGPCRs), we investigated the role of proline residues in the structure and function of this receptor. All prolines in TM segments of MT2 receptor were individually replaced with alanine and/or glycine. In addition, the unusual NAxxY motif located in TM7 was mutated to generate highly conserved NPxxY motif found in the majority of rGPCR proteins. Following transient expression in CHO-K1 cells, binding properties of the mutant receptors and their ability to transduce signals were analyzed using (125)I-mel- and [(35)S]GTPgammaS-binding assays, respectively. The impact of the performed mutations on the receptor structure was assessed by molecular dynamic simulations of MT2 receptors embedded in the fully hydrated phospholipid bilayer. Our results indicate that residues P174, P212 and P266 are important for the ligand binding and/or signaling of the human MT2 receptor. We also show that changes within the unusual NAxxY sequence in the TM7 (mutations A305P and A305V) produce defective MT2 receptors indicating an important role of this motif in the function of melatonin receptors.

中文翻译:

脯氨酸残基在人MT2褪黑激素受体的结构和功能中的作用。

褪黑激素是所有脊椎动物物种中各种生理过程的重要调节剂。在哺乳动物中,两个G蛋白偶联的褪黑激素受体(GPCR)介导了一些褪黑激素的作用:MT1和MT2。大多数GPCR的跨膜结构域(TM)包含一组高度保守的脯氨酸残基,这些残基可能起重要的结构和功能作用。与其他视紫红质样受体(rGPCR)相比,由于MT2受体的TM片段在特定脯氨酸残基的表达中表现出几个有趣的差异,因此我们研究了脯氨酸残基在该受体的结构和功能中的作用。MT2受体的TM区段中的所有脯氨酸分别被丙氨酸和/或甘氨酸替代。此外,将位于TM7中的异常NAxxY基序突变,以生成在大多数rGPCR蛋白中发现的高度保守的NPxxY基序。在CHO-K1细胞中瞬时表达后,分别使用(125)I-mel-和[(35)S] GTPgammaS结合分析法分析了突变受体的结合特性及其转导信号的能力。通过嵌入完全水合磷脂双层中的MT2受体的分子动力学模拟,评估了所执行的突变对受体结构的影响。我们的结果表明,残基P174,P212和P266对于配体结合和/或人MT2受体的信号传导很重要。
更新日期:2019-11-01
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