Direct Observation of Structural Unraveling in PABPN1 Polyalanine Expansions

Abstract

Above-threshold homorepeat expansions promote alternative conformations and irreversible aggregation, driving the malfunction of numerous disease-associated proteins. A detailed understanding of the structural remodeling events in pathogenic mutants enables the identification of key elements determining aberrant structure formation. Here, we describe in high resolution the structural transitions in Poly(A) Binding Protein Nuclear 1 (PABPN1), in which polyalanine expansions beyond a +1 single additional residue cause Oculopharyngeal Muscular Dystrophy (OPMD). PABPN1’s polyalanine tract in its wild-type (containing 10 consecutive alanine residues) and +1 and +8 variants populate alternative polymorphs, with α-helical conformations being increasingly favored and rigid upon expansion. Spontaneous slow unfolding was observed across variants in real time by Nuclear Magnetic Resonance (NMR) spectroscopy, indicating that increased dynamics at the C-terminal end of the polyalanine helix emerges as an early unfolding event associated with functional impairment. Our data help to define key mechanistic features in pathogenic misfunction induced by homorepeat expansions and identify specific structural motifs as potential targets for fragment-based binders to restore protein functionality in polyalanine expansion disorders.