Structural transitions in full-length human prion protein detected by xenon as probe and spin labeling of the N-terminal domain.

Sunilkumar Puthenpurackal Narayanan,Divya Gopalakrishnan Nair,Daniel Schaal,Marisa Barbosa De Aguiar,Werner Kremer,Stephan Schwarzinger,Hans Robert Kalbitzer,Sabine Wenzel

doi:10.1038/srep28419

Sunilkumar Puthenpurackal Narayanan, Divya Gopalakrishnan Nair + Show 6 more

Open Access

https://doi.org/10.1038/srep28419

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Abstract

Fatal neurodegenerative disorders termed transmissible spongiform encephalopathies (TSEs) are associated with the accumulation of fibrils of misfolded prion protein PrP. The noble gas xenon accommodates into four transiently enlarged hydrophobic cavities located in the well-folded core of human PrP(23–230) as detected by [1H, 15N]-HSQC spectroscopy. In thermal equilibrium a fifth xenon binding site is formed transiently by amino acids A120 to L125 of the presumably disordered N-terminal domain and by amino acids K185 to T193 of the well-folded domain. Xenon bound PrP was modelled by restraint molecular dynamics. The individual microscopic and macroscopic dissociation constants could be derived by fitting the data to a model including a dynamic opening and closing of the cavities. As observed earlier by high pressure NMR spectroscopy xenon binding influences also other amino acids all over the N-terminal domain including residues of the AGAAAAGA motif indicating a structural coupling between the N-terminal domain and the core domain. This is in agreement with spin labelling experiments at positions 93 or 107 that show a transient interaction between the N-terminus and the start of helix 2 and the end of helix 3 of the core domain similar to that observed earlier by Zn2+-binding to the octarepeat motif.

Highlights

Residue 166 to 172 that occurs in two conformational states, in one state it contains a 310-helix, in the other state a type-I β-turn
In the present study xenon is used to probe structural transitions coupled to fluctuations of hydrophobic cavities in huPrP(23–230) and correlate them with the transitions detected by high pressure NMR-spectroscopy
Zn2+-binding to the octarepeat induces an almost complete shift of conformational equilibrium to a state where the octarepeat region is close to T200 at the start of helix 3 suggesting that this closed conformation protects the PrP from fibril formation[41]

Summary

Introduction

Residue 166 to 172 that occurs in two conformational states, in one state it contains a 310-helix, in the other state a type-I β-turn. These regions often can be identified by their typical strong, non-linear pressure response in high-pressure NMR spectroscopy[24]; the corresponding structural transition can be characterized by thermodynamic analysis of the data Such a study has been reported for the human and the Syrian hamster prion protein[15,25,26] where four different states of the folded core with differences of Gibbs free energies ΔG 0 of approximately 3, 11, and 19 kJ/mol could be defined. Already in the early paper by Zahn et al.[8] the authors realized from an analysis of their NMR data that between the unfolded part of the protein and the well-folded part at least transient interactions have to exist since in the folded part chemical shift changes were observed when the N-terminal residues were removed. For getting more experimental information about such an conformational equilibrium, we introduced spin labels just after the octarepeat in position 93 and in front of the AGAAAAGA motif in position 107 and observed their relaxation enhancement effects on the [1H, 15N]-HSQC spectra of human prion protein as presented in the following

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Conclusion