Mazna Petr, Obsilova Veronika, Jelinkova Irena, Balik Ales, Berka Karel, Sovova Zofie, Ettrich Rüdiger, Svoboda Petr, Obsil Tomas, Teisinger Jan
Institute of Physiology, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
J Neurochem. 2004 Nov;91(4):836-42. doi: 10.1111/j.1471-4159.2004.02758.x.
A model of the helical part of the human MT2 melatonin (hMT2) receptor, a member of the G protein-coupled receptors superfamily has been generated, based on the structure of bovine rhodopsin. Modeling has been combined with site-directed mutagenesis to investigate the role of the specific amino acid residues within the transmembrane domains (TM) numbers V, VI and VII of hMT2 receptor in the interaction with 2-iodomelatonin. Saturation binding assays with 2-iodomelatonin demonstrated that the substitution V204A (TMV) resulted in total loss of binding while the mutation V205A had no effect. The replacement of F209 with alanine led to a significant decrease in the Bmax value of receptor binding while mutations V205A and F209A also within TM V did not significantly change binding properties of the hMT2 receptor. In the case of TM VI, the substitution G271T caused substantial decrease in 2-iodomelatonin binding to the hMT2 receptor. The change L272A (TM VI) as well as mutation Y298A within TM VII completely abolished ligand binding to the receptor. These data suggest that several new amino acid residues within TM V, VI and VII are involved in ligand-MT2 receptor interaction.
基于牛视紫红质的结构，构建了人MT2褪黑素（hMT2）受体螺旋部分的模型，hMT2受体是G蛋白偶联受体超家族的成员。建模与定点诱变相结合，以研究hMT2受体跨膜结构域（TM）V、VI和VII内特定氨基酸残基在与2-碘褪黑素相互作用中的作用。用2-碘褪黑素进行的饱和结合试验表明，V204A（TMV）取代导致结合完全丧失，而V205A突变没有影响。用丙氨酸取代F209导致受体结合的Bmax值显著降低，而同样在TM V内的V205A和F209A突变并没有显著改变hMT2受体的结合特性。在TM VI的情况下，G271T取代导致2-碘褪黑素与hMT2受体的结合大幅减少。TM VI中的L272A变化以及TM VII中的Y298A突变完全消除了配体与受体的结合。这些数据表明，TM V、VI和VII内的几个新氨基酸残基参与了配体与MT2受体的相互作用。
