Linking Mechanochemistry to Protein Folding at the Single Bond Level

Abstract

The impact of post translational modifications (PTM) on protein folding remains elusive. S-sulfenylation is the first oxidation step of cysteinyl thiols to create sulfenic acid. This PTM has emerged as a key sensor that modulates protein function and redox signalling. Here we use single molecule force clamp spectroscopy to create a stable sulfenic acid conformation by promoting an OH- nucleophilic attack on a single disulfide bond embedded within the core of the 27th immunoglobulin domain of titin (I27). By applying a tailor-designed force protocol, we set out to study the effect of S-sulfenylation on the oxidative folding of the I27 protein. We observe that sulfenic acid is a necessary precursor that facilitates enzymatic-free disulfide bond reformation. Furthermore, this disulfide reformation mechanism competes with the evolution of sulfenic acid to higher oxidation state species, which ultimately triggers protein misfolding. These results demonstrate that the oxidation state of a single amino acid within a protein has the power to completely alter the outcome of protein folding.