Electrostatic Collapse of Intrinsically Disordered Acid-Rich Protein Is Sensitive to Counterion Valency

Abstract

Intrinsically disordered proteins (IDPs) respond sensitively to their ionic environment, yet the mechanisms driving ion-induced conformational changes remain incompletely understood. Here, we investigate how counterion valency modulates the dimensions of an extremely charged model IDP, the aspartic and glutamic acid-rich protein AGARP. Fluorescence correlation spectroscopy and size exclusion chromatography reveal a pronounced, valency-dependent reduction in its hydrodynamic radius, with divalent cations (Ca2+, Mg2+) inducing collapse at much lower activities than monovalent cations (Na+, K+). Molecular dynamics simulations, direct sampling, and polyampholyte theory quantitatively capture the Debye–Hückel screening by monovalent ions but not the enhanced compaction driven by divalent ion binding thereby suggesting that, beyond differences in screening strength, a valency-sensitive mode of interaction is at play. Circular dichroism spectroscopy shows that compaction occurs without secondary structure formation. Our results demonstrate a structure-free electrostatic collapse and suggest that specific chelation of divalent ions by disordered polyanionic protein chains is a key mechanism regulating IDP compaction, with implications for understanding their behavior in biologically relevant ionic environments.