One O-linked sugar can affect the coil-to-β structural transition of the prion peptide

Abstract

It has been known that the structural transition from PrP C to PrP Sc leads to the prion formation. This putative conformational change challenges the central dogma of the protein folding theory-''one sequence, one structure.'' Generally, scientists believe that there must be either a posttranslational modification or environmental factors involved in this event. However, all of the efforts to solve the mystery of the PrP C to PrP Sc transition have ended in vain so far. Here we provide evidence linking O-linked glycosylation to the structural transition based on prion peptide studies. We find that the O-linked ␣-GalNAc at Ser-135 suppresses the formation of amyloid fibril formation of the prion peptide at physiological salt concentrations, whereas the peptide with the same sugar at Ser-132 shows the opposite effect. Moreover, this effect is sugar specific. Replacing ␣-GalNAc with ␤-GlcNAc does not yield the same effect.T he prion protein (PrP, 254 aa for hamster PrP) has been found to be associated with the prion infectivity. The normal product of the prion gene is expressed as a glycophosphatidyl inositol-anchored glycoprotein on the outer cell membrane (1-3). The solution NMR structure of recombinant syrian hamster prion protein rPrP(90-231) was determined in 1997 (4). The secondary structure consists of three ␣-helices (144-153, 172-194, 200-227) interdispersed between two short ␤-strands (129)(130)(131)(161)(162)(163); the N terminus (90-111) is largely unstructured ( Fig. 1). A posttranslational process that converts the cellular prion protein (PrP C ) to the abnormal, insoluble, pathogenic isoform (PrP Sc ) has been implicated in the prion formation during the development of prion diseases.CD spectroscopy and Fourier transform infrared spectroscopy studies have indicated that approximately half of the ␣-helical and coiled structure in PrP C is transformed into ␤-sheet in PrP Sc (5). However, the thermodynamics of the ␣-to-␤ transition, including the factors that influence it, remain unknown. PrP C has a high turnover rate in vivo. This high turnover makes the purification of PrP C extremely difficult (6). The low yield of purified PrP C from animal brains or low level of expression in mammalian cells as well as the insolubility of PrP Sc have remained a bottleneck in prion research. It has been difficult to discern whether there are chemical modifications in PrP C that are absent in PrP Sc or vice versa. Although Edman sequencing data suggest that the amino acid sequence of PrP Sc is identical with the translational product of the prion gene (7), we cannot rule out the possibility of low levels of unidentified chemical modifications in PrP C or PrP Sc as the culprit of the medical malady. To date, the search for differential modifications between PrP C and PrP Sc has focused on the two N-linked glycosylation sites at N181 and N197. However, based on the results of tunicamycin treatment, an N-linked glycosylation inhibitor (8), as well as experiments on the expression of unglycosylated mutated prion prot...

How Post‐Translational Modifications Influence Amyloid Formation: A Systematic Study of Phosphorylation and Glycosylation in Model Peptides

“…Previous studies on O-glycosylated prion protein (PrP) fragments revealed that even the orientation of a single hydroxyl group at the C4 of the sugar can dictate the process of fibril formation. [44] Additionally, it was shown that a N-linked glycan can also indirectly reduce the rate of fibril formation of a PrP-fragment by promoting intermolecular disulfide formation. [45] Conclusion Post-translational modifications of proteins play a key role in their structure, function, and stability.…”

Glycosylation and Specific Deamidation of Ribonuclease B Affect the Formation of Three-dimensional Domain-swapped Oligomers

“…The results presented here suggest that the formation of threedimensional domain-swapped oligomers is less sensitive to glycosylation or deamidation than later stages in amyloid formation. Recently, the addition of a single sugar residue has been found to greatly accelerate or inhibit amyloid formation by a highly amyloidogenic human prion domain, depending on its position (27). The deamidation of the Asn residue in the amyloidogenic peptide amylin has been shown to decisively accelerate amyloid fibril formation (63), whereas the oxidation of Met residues inhibits fibril formation in the Alzheimer's disease A␤ peptide (64) and in ␣-synuclein (65).…”

“…However, a single sugar residue in a human prion protein domain has been found not only to inhibit amyloidogenesis, but also to accelerate it, depending on its position (27). One key objective of this work is to test the ability of RNase B to form domain-swapped oligomers.…”

Dissociation of recombinant prion autocatalysis from infectivity

“…Indeed, N-linked glycans are known to have a chaperone-like activity during protein folding and to enhance overall protein conformational stability. 19 In addition, covalently linked PrP glycans have been shown to alter the rate of amyloid fibril formation, 20 limit the prion strain susceptibility of host animals, 21 and even influence interspecies transmission barriers. 22 The role of the GPI anchor modification in protein folding is less well studied.…”