The Role of Individual Residues in the N-Terminus of Arrestin-1 in Rhodopsin Binding

Sequences and three-dimensional structures of the four vertebrate arrestins are very similar, yet in sharp contrast to other subtypes arrestin-1 demonstrates exquisite selectivity for the active phosphorylated form of its cognate receptor, rhodopsin. The N-terminus participates in receptor binding and serves as the anchor of the C-terminus, the release of which facilitates arrestin transition into receptor-binding state. We tested the effects of substitutions of fourteen residues in the N-terminus of arrestin-1 on the binding to phosphorylated and unphosphorylated light-activated rhodopsin of wild type protein and its enhanced mutant with C-terminal deletion that demonstrates higher binding to both functional forms of rhodopsin. Profound effects of mutations identified lysine-15 as the main phosphate sensor and phenylalanine-13 as the key anchor of the C-terminus. These residues are conserved in all arrestin subtypes. Substitutions of five other residues reduced arrestin-1 selectivity, indicating that wild type residues participate in fine-tuning of arrestin- 1 binding to rhodopsin. Differential effects of numerous substitutions in wild type and an enhanced mutant arrestin-1 shows that these two proteins bind rhodopsin differently.