Céline Chapuis

The oligomerization properties of prion protein are restricted to the H2H3 domain

The propensity of the prion protein (PrP) to adopt different structures is a clue to its pathological behavior. The determination of the region involved in the PrPC to PrPSc conversion is fundamental for the understanding of the mechanisms underlying this process at the molecular level. In this paper, the polymerization of the helical H2H3 domain of ovine PrP (OvPrP) was compared to the full‐length construct (using chromatography and light scattering). We show that the oligomerization patterns are identical, although the H2H3 domain has a higher polymerization rate. Furthermore, the depolymerization kinetics of purified H2H3 oligomers compared to those purified from the full‐length PrP reveal that regions outside H2H3 do not significantly contribute to the oligomerization process. By combining rational mutagenesis and molecular dynamics to investigate the early stages of H2H3 oligomerization, we observe a conformationally stable β‐sheet structure that we propose as a possible nucleus for oligomerization; we also show that single point mutations in H2 and H3 present structural polymorphisms and oligomerization properties that could constitute the basis of species or strain variability.—Chakroun, N., Prigent, S., Dreiss, C. A., Noinville, S., Chapuis, C., Fraternali, F., Rezaei, H. The oligomerization properties of prion protein are restricted to the H2H3 domain. FASEB J. 24, 3222–3231 (2010). www.fasebj.org

Prion fibrillization is mediated by a native structural element that comprises helices H2 and H3.

Aggregation and misfolding of the prion protein (PrP) are thought to be the cause of a family of lethal neurodegenerative diseases affecting humans and other animals. Although the structures of PrP from several species have been solved, still little is known about the mechanisms that lead to the misfolded species. Here, we show that the region of PrP comprising the hairpin formed by the helices H2 and H3 is a stable independently folded unit able to retain its secondary and tertiary structure also in the absence of the rest of the sequence. We also prove that the isolated H2H3 is highly fibrillogenic and forms amyloid fibers morphologically similar to those obtained for the full-length protein. Fibrillization of H2H3 but not of full-length PrP is concomitant with formation of aggregates. These observations suggest a “banana-peeling” mechanism for misfolding of PrP in which H2H3 is the aggregation seed that needs to be first exposed to promote conversion from a helical to a β-rich structure.

Substitutions at residue 211 in the prion protein drive a switch between CJD and GSS syndrome, a new mechanism governing inherited neurodegenerative disorders

Human prion diseases are a heterogeneous group of fatal neurodegenerative disorders, characterized by the deposition of the partially protease-resistant prion protein (PrP(res)), astrocytosis, neuronal loss and spongiform change in the brain. Among inherited forms that represent 15% of patients, different phenotypes have been described depending on the variations detected at different positions within the prion protein gene. Here, we report a new mechanism governing the phenotypic variability of inherited prion diseases. First, we observed that the substitution at residue 211 with either Gln or Asp leads to distinct disorders at the clinical, neuropathological and biochemical levels (Creutzfeldt-Jakob disease or Gerstmann-Sträussler-Scheinker syndrome with abundant amyloid plaques and tau neurofibrillar pathology). Then, using molecular dynamics simulations and biophysical characterization of mutant proteins and an in vitro model of PrP conversion, we found evidence that each substitution impacts differently the stability of PrP and its propensity to produce different protease resistant fragments that may contribute to the phenotypical switch. Thus, subtle differences in the PrP primary structure and stability are sufficient to control amyloid plaques formation and tau abnormal phosphorylation and fibrillation. This mechanism is unique among neurodegenerative disorders and is consistent with the prion hypothesis that proposes a conformational change as the key pathological event in prion disorders.

Vesicle Permeabilization by Purified Soluble Oligomers of Prion Protein: A Comparative Study of the Interaction of Oligomers and Monomers with Lipid Membranes

The conversion of normal cellular prion protein (PrP) into its pathological isoform, scrapie PrP, may occur at the cell surface or, more probably, in late endosomes. The early events leading to the structural conversion of PrP appear to be related to the presence of more or less stable soluble oligomers, which might mediate neurotoxicity. In the current study, we investigate the interaction of alpha-rich PrP monomers and beta-rich size-exclusion-chromatography-purified PrP oligomers with lipid membranes. We compare their structural properties when associated with lipid bilayers and study their propensities to permeabilize the membrane at physiological pH. We also study the influence of the N-terminal flexible region (residues 24-103) by comparing full-length PrP(24-234) and N-terminally truncated PrP(104-234) oligomers. We showed that both 12-subunit oligomers cause an immediate and large increase in the permeability of the membrane, whereas equivalent amounts of monomeric forms cause no detectable leakage. Although the two monomeric PrP constructs undergo an alpha-to-beta conformational change when bound to the negatively charged membrane, only the full-length form of monomeric PrP has a weak fusogenic effect. Finally, the oligomers affect the integrity of the membrane differently from the monomers, independently of the presence of the N-terminal flexible domain. As for other forms of amyloidogenesis, a reasonable mechanism for the toxicity arising from PrP fibrillization must be associated with low-molecular-weight oligomeric intermediates, rather than with mature fibrils. Knowledge of the mechanism of action of these soluble oligomers would have a high impact on the development of novel therapeutic targets.