Tuning the conformational properties of the prion peptide.
Previously, we disclosed that O-linked glycosylation of Ser-132 or Ser-135 could dramatically change the amyloidogenic property of the hamster prion peptide (sequence 108-144). This peptide, which corresponds to the flexible loop and the first beta-strand in the structure of the prion protein, is a random coil when it is initially dissolved in buffer, but amyloid fibrils are formed with time. Thus, it offers a convenient model system to observe and compare how different chemical modifications and sequence mutations alter the amyloidogenic property of the peptide within a reasonable experimental time frame. In our earlier study, aside from uncovering a site-specificity of the glycosylation on the fibrillogenesis, different effects of alpha-GalNAc and beta-GlcNAc were observed. In this work, we explore further how different sugar configurations affect the conformational property of the polypeptide chain. We compare the effects of O-linked glycosylation by the common sugars alpha-GalNAc, beta-GlcNAc with their non-native analogs beta-GalNAc, alpha-GlcNAc in an effort to uncover the origin of the sugar-specificity on the fibril formation. We find that the anomeric configuration of the sugar is the most important factor affecting the fibrillogenesis. Sugars with the glycosidic bond in the alpha-configuration at Ser-135 have a dramatic inhibitory effect on the structural conversion of the glycosylated peptide. Because O-glycosylation of Ser-135 with alpha-linked sugars also promote the formation of three slowly converting conformations at the site of glycosylation, we surmise that the amyloidogenic property of the peptide is related to its conformational flexibility, and the proclivity of this region of the peptide to undergo the structural conversion from the random coil to form the beta-structure. Upon O-glycosylation with an alpha-linked sugar, this conversion is inhibited and the nucleation of fibril formation is largely retarded. Consistent with this scenario, Arg-136 is the residue most affected in the TOCSY NMR spectra of the glycosylated peptides, other than the serine site modified. In addition, when Arg-136 is substituted by Gly, a mutation that should provide higher structural flexibility in this part of the peptide, the amyloidogenic property of the peptide is greatly enhanced, and the inhibition effect of glycosylation is largely diminished. These results are consistent with Ser-135 and Arg-136 being part of the kink region involved in the structural conversion.
Ho
Chai-Chi
CC
Institute of Biological Chemistry, Academia Sinica, Taipei 115, Taiwan, Republic of China.
Lee
Lily Y-L
LY
Huang
Kuo-Ting
KT
Lin
Chun-Cheng
CC
Ku
Mei-Yun
MY
Yang
Chien-Chih
CC
Chan
Sunney I
SI
Hsu
Ruei-Lin
RL
Chen
Rita P-Y
RP
D000595:Amino Acid Sequence; D000682:Amyloid / Q000737:chemistry / Q000378:metabolism; D005944:Glucosamine / Q000737:chemistry* / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism; D011328:Prions / Q000737:chemistry* / Q000378:metabolism*; D011487:Protein Conformation; D017510:Protein Folding
D000682:Amyloid; D010455:Peptides; D011328:Prions; D005944:Glucosamine