Abstract
Paramagnetic relaxation enhancement is an NMR technique that has yielded important insight into the structure of folded proteins, although the perturbation introduced by the large spin probe might be thought to diminish its usefulness when applied to characterizing the structural ensembles of intrinsically disordered proteins (IDPs). We compare the computationally generated structural ensembles of the IDP amyloid-β42 (Aβ42) to an alternative sequence in which a nitroxide spin label attached to cysteine has been introduced at its N-terminus. Based on this internally consistent computational comparison, we find that the spin label does not perturb the signature population of the β-hairpin formed by residues 16-21 and 29-36 that is dominant in the Aβ42 reference ensemble. However, the presence of the tag induces a strong population shift in a subset of the original Aβ42 structural sub-populations, including a sevenfold enhancement of the β-hairpin formed by residues 27-31 and 33-38. Through back-calculation of NMR observables from the computational structural ensembles, we show that the structural differences between the labeled and unlabeled peptide would be evident in local residual dipolar couplings, and possibly differences in homonuclear (1)H-(1)H nuclear Overhauser effects (NOEs) and heteronuclear (1)H-(15)N NOEs if the paramagnetic contribution to the longitudinal relaxation does not suppress the NOE intensities in the real experiment. This work shows that molecular simulation provides a complementary approach to resolving the potential structural perturbations introduced by reporter tags that can aid in the interpretation of paramagnetic relaxation enhancement, double electron-electron resonance, and fluorescence resonance energy transfer experiments applied to IDPs.