A lowly populated, transient β-sheet structure in monomeric Aβ1-42 identified by multinuclear NMR of chemical denaturation

Chemical denaturation is a well-established approach for probing the equilibrium between folded and unfolded states of proteins. We demonstrate applicability of this method to the detection of a small population of a transiently folded structural element in a system that is often considered to be intrinsically fully disordered. The H-1(N), N-15, C-13(alpha), and C-13' chemical shifts of A beta(1-40) and A beta(1-42) peptides and their M35-oxidized variants were monitored as a function of urea concentration and compared to analogous urea titrations of synthetic pentapeptides of homologous sequence. Fitting of the chemical shift titrations yields a 10 +/- 1% population for a structured element at the C-terminus of A beta(1-42) that folds with a cooperativity of m = 0.06 kcal/mol.M. The fit also yields the chemical shifts of the folded state and, using a database search, for A beta(1-42) these shifts identified an antiparallel intramolecular beta-sheet for residues I32-A42, linked by a type I' beta-turn at G37 and G38. The structure is destabilized by oxidation of M35. Paramagnetic relaxation rates and two previously reported weak, mediumrange NOE interactions are consistent with this transient beta-sheet. Introduction of the requisite A42C mutation and tagging with MTSL resulted in a small stabilization of this beta-sheet. Chemical shift analysis suggests a C-terminal beta-sheet may be present in A beta(1-40) too, but the turn type at G37 is not type I'. The approach to derive Transient Structure from chemical Denaturation by NMR (TSD-NMR), demonstrated here for A beta peptides, provides a sensitive tool for identifying the presence of lowly populated, transiently ordered elements in proteins that are considered to be intrinsically disordered, and permits extraction of structural data for such elements.

Automated summary

Chemical denaturation is a useful method for detecting transiently folded structural elements in proteins that are considered to be intrinsically disordered. Analysis of chemical shift titrations can identify lowly populated structured elements in proteins.