Complex folding and misfolding effects of deer-specific amino acid substitutions in the β2-α2 loop of murine prion protein.
The β2-α2 loop of PrP(C) is a key modulator of disease-associated prion protein misfolding. Amino acids that differentiate mouse (Ser169, Asn173) and deer (Asn169, Thr173) PrP(C) appear to confer dramatically different structural properties in this region and it has been suggested that amino acid sequences associated with structural rigidity of the loop also confer susceptibility to prion disease. Using mouse recombinant PrP, we show that mutating residue 173 from Asn to Thr alters protein stability and misfolding only subtly, whilst changing Ser to Asn at codon 169 causes instability in the protein, promotes oligomer formation and dramatically potentiates fibril formation. The doubly mutated protein exhibits more complex folding and misfolding behaviour than either single mutant, suggestive of differential effects of the β2-α2 loop sequence on both protein stability and on specific misfolding pathways. Molecular dynamics simulation of protein structure suggests a key role for the solvent accessibility of Tyr168 in promoting molecular interactions that may lead to prion protein misfolding. Thus, we conclude that 'rigidity' in the β2-α2 loop region of the normal conformer of PrP has less effect on misfolding than other sequence-related effects in this region.
The Roslin Institute and R(D)SVS, University of Edinburgh, Easter Bush Veterinary Centre, Roslin, Edinburgh EH25 9RG, UK.
Sonya
S
Agarwal
The Roslin Institute and R(D)SVS, University of Edinburgh, Easter Bush Veterinary Centre, Roslin, Edinburgh EH25 9RG, UK.
Kristina
K
Döring
School of Life Sciences, University of Warwick, Gibbet Hill Road, Coventry, CV4 7AL, UK.
Leszek A
LA
Gierusz
The Roslin Institute and R(D)SVS, University of Edinburgh, Easter Bush Veterinary Centre, Roslin, Edinburgh EH25 9RG, UK.
Pooja
P
Iyer
The Roslin Institute and R(D)SVS, University of Edinburgh, Easter Bush Veterinary Centre, Roslin, Edinburgh EH25 9RG, UK.
Fiona M
FM
Lane
The Roslin Institute and R(D)SVS, University of Edinburgh, Easter Bush Veterinary Centre, Roslin, Edinburgh EH25 9RG, UK.
James F
JF
Graham
The Roslin Institute and R(D)SVS, University of Edinburgh, Easter Bush Veterinary Centre, Roslin, Edinburgh EH25 9RG, UK.
Wilfred
W
Goldmann
School of Life Sciences, University of Warwick, Gibbet Hill Road, Coventry, CV4 7AL, UK.
Teresa J T
TJ
Pinheiro
The Roslin Institute and R(D)SVS, University of Edinburgh, Easter Bush Veterinary Centre, Roslin, Edinburgh EH25 9RG, UK.
Andrew C
AC
Gill
D000595:Amino Acid Sequence / Q000235:genetics; D019943:Amino Acid Substitution / Q000235:genetics*; D000818:Animals; D003670:Deer / Q000235:genetics; D006801:Humans; D051379:Mice; D017096:Prion Diseases / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D011328:Prions / Q000737:chemistry / Q000235:genetics*; D017510:Protein Folding; D055550:Protein Stability; D017433:Protein Structure, Secondary / Q000235:genetics; D057165:Proteostasis Deficiencies / Q000235:genetics* / Q000378:metabolism / Q000473:pathology
D011328:Prions