Molecular characteristics of a protease-resistant, amyloidogenic and neurotoxic peptide homologous to residues 106-126 of the prion protein.
In the prion-related encephalopathies the prion protein is converted to an altered form, known as PrPSc, that is partially resistant to protease digestion. This abnormal isoform accumulates in the brain and its protease-resistant core aggregates extracellularly into amyloid fibrils. We have investigated the conformational properties, aggregation behaviour and sensitivity to protease digestion of a synthetic peptide homologous to residues 106-126 of human PrP, which was previously found to form amyloid-like fibrils in vitro and displayed neurotoxic activity toward primary cultures of rat hippocampal neurons. A scrambled sequence of peptide PrP 106-126 was used as a control. By circular dichroism, PrP 106-126 exhibited a secondary structure composed largely of beta-sheet, whereas the scrambled sequence of PrP 106-126 showed a random coil structure. The beta-sheet content of PrP 106-126 was much higher in 200 mM phosphate buffer at pH 5.0 than in the same buffer at pH 7.0. Laser light scattering analysis showed that PrP 106-126 aggregated immediately after dissolution in 20 mM or 200 mM phosphate buffer, pH 5.0 and 7.0, whereas scrambled PrP 106-126 did not. PrP 106-126 aggregates had an average hydrodinamic diameter of 100 nm and an average molecular weight of 12 x 10(6) +/- 30% Daltons, corresponding to the aggregation of 6000 +/- 30% molecules. Peptide PrP 106-126 showed partial resistance to digestion with Proteinase K and Pronase, whereas scrambled PrP 106-126 was completely degraded by incubation with the enzymes at 37 degrees C for 30 minutes.
Selvaggini
C
C
Istituto di Ricerche Farmacologiche Mario Negri, Milano, Italy.
De Gioia
L
L
Cantù
L
L
Ghibaudi
E
E
Diomede
L
L
Passerini
F
F
Forloni
G
G
Bugiani
O
O
Tagliavini
F
F
Salmona
M
M
D000595:Amino Acid Sequence; D002942:Circular Dichroism; D019286:Endopeptidase K; D006801:Humans; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry*; D010446:Peptide Fragments / Q000737:chemistry* / Q000378:metabolism*; D018620:PrPSc Proteins; D011328:Prions / Q000737:chemistry* / Q000378:metabolism*; D011402:Pronase / Q000378:metabolism; D017433:Protein Structure, Secondary; D017386:Sequence Homology, Amino Acid; D012697:Serine Endopeptidases / Q000378:metabolism; D012995:Solubility
D009419:Nerve Tissue Proteins; D010446:Peptide Fragments; D018620:PrPSc Proteins; D011328:Prions; C082632:prion protein (106-126); D012697:Serine Endopeptidases; D019286:Endopeptidase K; D011402:Pronase