Conformational Differences Between Arrestin2 and Pre-activated Mutants as Revealed by Hydrogen Exchange Mass Spectrometry

摘要: Arrestins are regulatory proteins that bind specifically to ligand-activated phosphorylated G protein-coupled receptors terminate protein-mediated signaling, cause the internalization of receptor-arrestin complex, and initiate additional intracellular signaling cascades. Multiple lines evidence suggest arrestin normally exists in an inactive basal state undergoes conformational activation process receptor binding. "Pre-activated" phosphorylation-independent mutants display increased binding but unphosphorylated receptors. The mutations believed expose key receptor-binding regions, allowing mimic, some extent, transition its active state. In present study, amide hydrogen exchange (HX) mass spectrometry (MS) were used examine conformation wild-type arrestin2 compare solution with two pre-activated (R169E 3A (I385A, V386A, F387A)). results unexpected level structural organization within elements containing clathrin adaptin2-binding sites previously be completely disordered. Increased deuterium incorporation was observed both mutant forms compared wild-type, indicating a change mutants. Three regions demonstrated significant differences incorporation: first 33 residues N terminus 243-255 (both implicated interaction), 271-299. subtle responsible for difference biological activity displayed by similar changes occur receptor.