Rescuing Functional Protein from Amyloid-Like Structure

Abstract

Formation of amyloid deposits is the molecular background of several diseases. Protein oligomers and aggregates formed in the process are connected to the observed pathogenesis. Amyloids represent the end stage of a multi-step aggregation cascade. Although some recovery of the enzyme activity from amyloid deposits has been reported in the case of the lysozyme (Booth II et al., Nature 385 (1997) 787-93), the recovery of the active structure from amyloid or amyloid-like deposits has not been studied. Here we show that phosphoglycerate kinase can be refolded into the biologically active structure from amyloid-like fibrils. First, amyloid-like fibrils were grown from phosphoglycerate kinase. The conversion of the protein structure was confirmed by electron microscopy, enzyme activity assays, as well as by Congo red and Thioflavin T binding measurements. Next, the protein was refolded into its native structure. Biological equivalence of the reference and recovered enzyme was confirmed by enzyme activity and differential scanning calorimetry measurements. We found that stabilizing the native fold is not enough for the efficient recovery of the native enzyme. The aggregates have to be destabilized before the formation of the native structure is initiated.