A Proposed Mechanism for the Promotion of Prion Conversion Involving a Strictly Conserved Tyrosine Residue in the β2-α2 Loop of PrPC

Timothy D Kurt,Lin Jiang,Cyrus Bett,David Eisenberg,Christina J Sigurdson

doi:10.1074/jbc.m114.549030

Abstract

The transmission of infectious prions into different host species requires compatible prion protein (PrP) primary structures, and even one heterologous residue at a pivotal position can block prion infection. Mapping the key amino acid positions that govern cross-species prion conversion has not yet been possible, although certain residue positions have been identified as restrictive, including residues in the β2-α2 loop region of PrP. To further define how β2-α2 residues impact conversion, we investigated residue substitutions in PrP(C) using an in vitro prion conversion assay. Within the β2-α2 loop, a tyrosine residue at position 169 is strictly conserved among mammals, and transgenic mice expressing mouse PrP having the Y169G, S170N, and N174T substitutions resist prion infection. To better understand the structural requirements of specific residues for conversion initiated by mouse prions, we substituted a diverse array of amino acids at position 169 of PrP. We found that the substitution of glycine, leucine, or glutamine at position 169 reduced conversion by ∼ 75%. In contrast, replacing tyrosine 169 with either of the bulky, aromatic residues, phenylalanine or tryptophan, supported efficient prion conversion. We propose a model based on a requirement for tightly interdigitating complementary amino acid side chains within specific domains of adjacent PrP molecules, known as "steric zippers," to explain these results. Collectively, these studies suggest that an aromatic residue at position 169 supports efficient prion conversion.

Highlights

Single residue differences can block conversion of the cellular prion protein (PrP) to the pathogenic conformation
We recently found that transgenic mice expressing mouse PrP (MoPrP) having the Y169G substitution together with the S170N and N174T substitutions completely resist infection with either mouse or deer prions, implicating tyrosine 169 as critical for prion conversion [36]
Similar to protein-misfolding cyclic amplification [45], the mixtures of PrPC and PrPSc were subjected to 24 h of intermittent bursts of sonication followed by incubation at 37 °C

Summary

Background

Single residue differences can block conversion of the cellular prion protein (PrP) to the pathogenic conformation. We propose a model based on a requirement for tightly interdigitating complementary amino acid side chains within specific domains of adjacent PrP molecules, known as “steric zippers,” to explain these results These studies suggest that an aromatic residue at position 169 supports efficient prion conversion. Transgenic mice expressing mouse PrP with the D167S substitution (MoPrP167), which results in a well defined ␤2-␣2 loop by NMR [31], show no detectable change in species barriers [32], suggesting that the PrP primary sequence may override the secondary structure in promoting prion conversion. We found robust differences in prion conversion among the PrPC variants, and we propose a structural model based on amino acid side chain interactions within a steric zipper comprising PrP residues 167–176 to explain these results

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION