A Chemical Mutagenesis Approach to Insert Post-translational Modifications in Aggregation-Prone Proteins

[Image: see text] Neurodegenerative diseases are a class of disorders linked to the formation in the nervous system of fibrillar protein aggregates called amyloids. This aggregation process is affected by a variety of post-translational modifications, whose specific mechanisms are not fully understood yet. Emerging chemical mutagenesis technology is currently striving to address the challenge of introducing protein post-translational modifications, while maintaining the stability and solubility of the proteins during the modification reaction. Several amyloidogenic proteins are highly aggregation-prone, and current modification procedures can lead to unexpected precipitation of these proteins, affecting their yield and downstream characterization. Here, we present a method for maintaining amyloidogenic protein solubility during chemical mutagenesis. As proof-of-principle, we applied our method to mimic the phosphorylation of serine-26 and the acetylation of lysine-28 of the 40-residue long variant of amyloid-β peptide, whose aggregation is linked to Alzheimer’s disease.