The interplay of receptor dynamics and ligand residence time upon ligand efficacy at adenosine receptors (C02)

导致配体停留时间延长的分子细节目前还不够清楚。这个项目中的主要问题将是利用活细胞中具有高动力学分辨率的FRET显微镜来研究与配体结合相关的受体的构象动力学。使用腺苷受体，我们将：i)研究细胞外环2在配体结合动力学中的作用；ii)比较低配体亲和力的A2B受体与高亲和力的A1和A2A受体；iii)研究动态配体结合与A1受体H251A突变的构象动力学相关的关系；iv)研究G蛋白在受体动力学和受体定位中的影响；v)以PD 81,723为原型，研究变构调节剂在停留时间和信号传递中的作用；vi)使系统适应96孔板以获得更快的吞吐能力。

The molecular details leading to increased ligand residence time are currently insufficiently understood. The major question within this project will be to address conformational dynamic of a receptor in relation to ligand binding using FRET-microscopy with high kinetic resolution in living cells. Using adenosine receptors, we will: I) Study the role of the extracellular loop 2 in ligand binding kinetics; II) Compare the low ligand affinity A2B receptor to the high affinity A1 and A2A receptors; III) Investigate dynamic ligand binding in relation to the conformational dynamics of the H251A mutation at the A1 receptor; IV) Study the influence of G-proteins in receptor dynamics and receptor localization; V) Investigate the role of allosteric modulators in residence time and signaling using PD 81,723 as prototype; VI) Adapt the system to 96-well plates for faster throughput.