Influence of the Dynamically Disordered N-Terminal Tail Domain on the Amyloid Core Structure of Human Y145Stop Prion Protein Fibrils.

Zhe Qi,Krystyna Surewicz,Christopher P Jaroniec,Witold K Surewicz

doi:10.3389/fmolb.2022.841790

Abstract

The Y145Stop mutant of human prion protein (huPrP23-144) is associated with a familial prionopathy and provides a convenient in vitro model for investigating amyloid strains and cross-seeding barriers. huPrP23-144 fibrils feature a compact and relatively rigid parallel in-register β-sheet amyloid core spanning ∼30 C-terminal amino acid residues (∼112–141) and a large ∼90-residue dynamically disordered N-terminal tail domain. Here, we systematically evaluate the influence of this dynamic domain on the structure adopted by the huPrP23-144 amyloid core region, by investigating using magic-angle spinning solid-state nuclear magnetic resonance (NMR) spectroscopy a series of fibril samples formed by huPrP23-144 variants corresponding to deletions of large segments of the N-terminal tail. We find that deletion of the bulk of the N-terminal tail, up to residue 98, yields amyloid fibrils with native-like huPrP23-144 core structure. Interestingly, deletion of additional flexible residues in the stretch 99–106 located outside of the amyloid core yields shorter heterogenous fibrils with fingerprint NMR spectra that are clearly distinct from those for full-length huPrP23-144, suggestive of the onset of perturbations to the native structure and degree of molecular ordering for the core residues. For the deletion variant missing residues 99–106 we show that native huPrP23-144 core structure can be “restored” by seeding the fibril growth with preformed full-length huPrP23-144 fibrils.

Highlights

Most peptide and protein molecules are capable of undergoing conformational conversion from their native state into highly ordered, β-sheet rich amyloid fibrils (Dobson, 1999), and for ~50 human proteins such misfolding and amyloid formation can occur under physiological conditions in vivo leading to development of disease (Chiti and Dobson, 2006)
To investigate this further we prepared the Δ92-98 variant, which was found to form WT-like fibrils suggesting that deletion of huPrP23-144 N-terminal residues up to aa 98 does not have a significant impact on formation of the native huPrP23-144 amyloid core structure
On the basis of effectively identical fibril assembly kinetics, morphologies and fingerprint solid-state nuclear magnetic resonance (NMR) spectra for all huPrP23-144 variants studied containing large deletions up to residue 98, our results indicate that the bulk of the dynamically disordered N-terminal tail domain of huPrP23-144 is not essential for amyloid formation under autocatalytic conditions and ability of the resulting β-core region to adopt a WT-like structure

Summary

Introduction

Most peptide and protein molecules are capable of undergoing conformational conversion from their native state into highly ordered, β-sheet rich amyloid fibrils (Dobson, 1999), and for ~50 human proteins such misfolding and amyloid formation can occur under physiological conditions in vivo leading to development of disease (Chiti and Dobson, 2006). The highly homologous human (hu), mouse (mo) and Syrian hamster (Sha) PrP23-144 proteins (pairwise amino acid, aa, sequence identities of ~90–95%) have been shown to provide a valuable in vitro model for detailed investigation of the structural basis of amyloid strains and transmissibility barriers (Kundu et al, 2003; Vanik et al, 2004; Jones and Surewicz, 2005; Surewicz et al, 2006). Our previous structural and dynamic solidstate NMR studies of huPrP23-144 fibrils revealed the presence of a structured ~30-residue parallel in-register β-amyloid core (aa ~112–141) exhibiting limited protein backbone motions on the ~0.1–1 ms time scale located near the C-terminus and a large dynamically disordered ~90-residue N-terminal tail domain (aa ~23–110) (Helmus et al, 2008; Helmus et al, 2010; Helmus et al, 2011; Theint et al, 2018; Aucoin et al, 2019; Shannon et al, 2019). Additional studies of PrP23-144 amyloids containing mutations and deletions corresponding to different huPrP23144 core residues enabled these sequence modifications to be correlated with structural and dynamic changes in the PrP23-144 amyloid core and provided initial insights into mammalian PrP23-144 cross-seeding specificities (Jones et al, 2011; Theint et al, 2017a; Theint et al, 2017b; Dao et al, 2021)

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Results

Conclusion