Faculty of Life Sciences, Institute of Biochemistry, Leipzig University, Bruederstr. 34, 04103, Leipzig, Germany.
Department of Quantitative and Computational Biology, University of Southern California, Los Angeles, CA, USA.
Nat Commun. 2023 Mar 1;14(1):1151. doi: 10.1038/s41467-023-36797-2.
Understanding the molecular basis of arrestin-mediated regulation of GPCRs is critical for deciphering signaling mechanisms and designing functional selectivity. However, structural studies of GPCR-arrestin complexes are hampered by their highly dynamic nature. Here, we dissect the interaction of arrestin-2 (arr2) with the secretin-like parathyroid hormone 1 receptor PTH1R using genetically encoded crosslinking amino acids in live cells. We identify 136 intermolecular proximity points that guide the construction of energy-optimized molecular models for the PTH1R-arr2 complex. Our data reveal flexible receptor elements missing in existing structures, including intracellular loop 3 and the proximal C-tail, and suggest a functional role of a hitherto overlooked positively charged region at the arrestin N-edge. Unbiased MD simulations highlight the stability and dynamic nature of the complex. Our integrative approach yields structural insights into protein-protein complexes in a biologically relevant live-cell environment and provides information inaccessible to classical structural methods, while also revealing the dynamics of the system.
理解 arrestin 介导的 GPCR 调节的分子基础对于破译信号机制和设计功能选择性至关重要。然而，由于 GPCR-arrestin 复合物的高度动态性质，其结构研究受到阻碍。在这里，我们使用活细胞中的遗传编码交联氨基酸来剖析 arrestin-2（arr2）与类分泌素甲状旁腺激素 1 受体 PTH1R 的相互作用。我们确定了 136 个分子间接近点，这些点指导构建 PTH1R-arr2 复合物的能量优化分子模型。我们的数据揭示了现有结构中缺失的灵活受体元件，包括细胞内环 3 和近端 C 尾巴，并提出了迄今为止被忽视的 arrestin N 边缘正电荷区域的功能作用。无偏 MD 模拟强调了复合物的稳定性和动态性质。我们的综合方法提供了在生物学相关的活细胞环境中对蛋白质-蛋白质复合物的结构见解，并提供了经典结构方法无法获得的信息，同时还揭示了系统的动态。
