Functional and structural insights into interactions between β-Arrestin 1 and Gαs or Gαi1

Abstract

G proteins and arrestins are key transducers for G protein-coupled receptor (GPCR) signaling, mediating distinct downstream pathways. Recent evidence suggests that G proteins and beta-arrestins (beta arrs) can directly or functionally interact. However, the molecular details and functional consequences of G alpha-beta arr interactions remain poorly understood. Here, we quantify the binding affinities between beta arr1 and G alpha s or G alpha i1 in various activation states using microscale thermophoresis (MST). beta arr1 in the active conformational ensemble state favors binding, whereas G alpha activation status is less determinant. Hydrogen/deuterium exchange mass spectrometry reveals distinct conformational changes between G alpha s versus G alpha i1 upon beta arr1 binding, suggesting differential binding mechanism between G alpha s-beta arr1 and G alpha i1-beta arr1 complexes. Both the Ras-like domain and the alpha-helical domain of G alpha contribute to complex formation. Functionally, a BODIPY-FL-GTP gamma S assay shows that beta arr1 does not alter GDP/GTP turnover of G alpha s or G alpha i1, whereas beta-strand XX (beta XX) release assays demonstrate that G alpha s enhances beta arr1 C-tail release. Together, these results propose molecular mechanism of the interaction and asymmetric functional coupling within G alpha-beta arr complexes and uncover a previously underappreciated layer of GPCR signal transduction.