Pre-Homonuclear Decoupling Enables High-Resolution NMR Analysis of Intrinsically Disordered Proteins in Solution

Probing the atomic details of intrinsically disordered proteins is crucial to understanding their biological function and relation to pathogenesis. Although amide detected NMR experiments are widely employed in protein studies, (3)J(HNH alpha) couplings between amide (H-1(N)) and alpha (H-1(alpha)) protons impose an intrinsic limit on the achievable H-1(N) linewidth. Here, we present a homonuclear decoupling method that narrows the alpha-synuclein H-1(N) linewidths to 3-5 Hz. Tightly distributed (1)J(C alpha H alpha) coupling values were employed to generate homogeneous antiphase coherences of 2H(z)(a)H(y)(N) and 4H(alpha)(2)(z)H-alpha(3)(z)H-y(N) for nonglycine and glycine residues, respectively, which were combined with their in-phase H-y(N) counterparts to achieve homonuclear decoupling. By reducing the multiplet structure to a singlet, the width of the H-1(N) cross-peak was reduced by similar to 3-fold in the 2D HSQC and 3D intra-HNCA spectra, and good spectral quality was achieved without the need for postprocessing.