PrP charge structure encodes interdomain interactions.

Javier Martínez,Adriano Aguzzi,Ilia V Baskakov,Natallia Makarava,María Gasset,Rosa Sánchez,Milagros Castellanos

doi:10.1038/srep13623

Abstract

Almost all proteins contain charged residues, and their chain distribution is tailored to fulfill essential ionic interactions for folding, binding and catalysis. Among proteins, the hinged two-domain chain of the cellular prion protein (PrPC) exhibits a peculiar charge structure with unclear consequences in its structural malleability. To decipher the charge design role, we generated charge-reverted mutants for each domain and analyzed their effect on conformational and metabolic features. We found that charges contain the information for interdomain interactions. Use of dynamic light scattering and thermal denaturation experiments delineates the compaction of the α-fold by an electrostatic compensation between the polybasic 23–30 region and the α3 electronegative surface. This interaction increases stability and disfavors fibrillation. Independently of this structural effect, the N-terminal electropositive clusters regulate the α-cleavage efficiency. In the fibrillar state, use of circular dichroism, atomic-force and fluorescence microscopies reveal that the N-terminal positive clusters and the α3 electronegative surface dictate the secondary structure, the assembly hierarchy and the growth length of the fibril state. These findings show that the PrP charge structure functions as a code set up to ensure function and reduce pathogenic routes.

Highlights

D202, E207, E221 and E228 in α 3, expose their side chains to solvent, defining electronegative surface clusters[39]
To unveil the information encoded in the complementary solvent-exposed charge structure, we constructed several mutants consisting of charge reversions and inclusions and tested their effects on the properties of both the α -folded and fibrillar states
At the globular domain (GD), the α 3 charge surface was modified by independent E200K, Q217R, Q219K, and E221K substitutions

Summary

Introduction

D202, E207, E221 and E228 in α 3, expose their side chains to solvent, defining electronegative surface clusters[39] Of these charges, the structural D144 and D147 residues and their respective salt bridges stabilize PrPC, preventing conversion to protease resistance forms, whereas D178, E196 and E211 are prone sites for pathogenic mutations upon charge alteration[38,42]. The structural D144 and D147 residues and their respective salt bridges stabilize PrPC, preventing conversion to protease resistance forms, whereas D178, E196 and E211 are prone sites for pathogenic mutations upon charge alteration[38,42] Pathogenic mutations, such as E200K and Q217R, and the dominant negative E219K polymorphism alter the α 3 surface electrostatic potential, inducing minor folding effects[39]. Effects such as the stabilization of the native α -fold, dictating the efficiency of the α -cleavage, attenuating the fibrillation propensity and yielding the most benign amyloids suggest that the charge design ensures PrPC functions

Methods

Results

Conclusion