Insights into the Activation and Self-Association\nof Arrestin‑1

Arrestins\nhalt signal transduction by binding to the phosphorylated\nC-termini of activated G protein-coupled receptors. Arrestin-1, the\nfirst subtype discovered, binds to rhodopsin in rod cells. Mutations\nin <i>SAG</i>, the gene encoding Arrestin-1, are linked\nto Oguchi disease, characterized by delayed dark adaptation. Since\nthe discovery of Arrestin-1, substantial progress has been made in\nunderstanding the role of these regulatory proteins in phototransduction,\nincluding the characterization of visual phenotypes of animals and\nhumans lacking this protein, discovery of splice variants, and documentation\nof its binding to inositol-polyphosphates. Arrestin-1 was one of the\nfirst structurally characterized proteins in the phototransduction\ncascade. However, there are knowledge gaps regarding the conformational\nintermediates leading to its binding to phosphorylated rhodopsin.\nAmong various mammalian Arrestin-1 conformations captured via crystallography,\nthe preactivated state is represented by the mutant R175E-Arrestin-1\nand by a C-terminally truncated splice variant (p44). This report\ndescribes a novel purification method of Arrestin-1 from bovine retinas\nfollowed by limited proteolysis to obtain a protein resembling p44.\nWe solved the crystal structure of this preactivated, shortened <sup>3–367</sup>Arrestin-1 at a resolution of 1.40 Å. The\nstructure reveals a more complete picture of the finger loop structure\nand of the role of the polar core in the activation of Arrestin-1.\nThe structure of <sup>3–367</sup>Arrestin-1 captures an intermediate\nform halfway between the inactive and fully activated conformations\nof Arrestin-1. Finally, we addressed the question of Arrestin-1 oligomerization\nby comparing the packing interfaces in different Arrestin-1 crystals\nand dimer models predicted by AlphaFold 3.