Metal ion mediated transition from random coil to β-sheet and aggregation of Bri2-23, a natural inhibitor of Aβ aggregation

Abstract

Furin-dependent maturation of the BRI2 protein generates the Bri2-23 fragment that is able to arrest the aggregation of amyloidβ, the peptide implicated in Alzheimer's disease (AD). Bri2-23 contains cysteines at positions 5 and 22, which are likely to bind to metal ions such as Cu(i). Metal ions may play a role in the etiology of neurodegenerative disorders such as AD, and in this work we explore the metal ion induced folding and aggregation of Bri2-23 using Hg(ii) and Ag(i) as spectroscopic probes with structural and ligand preferences similar to those of Cu(i), while not displaying redox activity under the experimental conditions. In general, interaction of Bri2-23 with soft metal ions changes the structural properties and solution behavior of the peptide that tune to increasing metal to peptide stoichiometry. Potentiometric, (199m)Hg PAC and ESI-MS data indicate that addition of up to 0.5 equivalents of Hg(ii) to Bri2-23 yields a two-coordinated HgS2 structure at the metal site. While the free peptide is inherently unstructured, the presence of Ag(i) and Hg(ii) gives rise to β-sheet formation. NMR spectroscopy supports the formation of β-sheet structure in the presence of 0.5 equivalents of Hg(ii), and displays an interesting and marked change in the TOCSY spectra when increasing the Hg(ii) to peptide stoichiometry from 0.5 to 0.7 equivalents, indicating the equilibrium between two structural analogues of the complex. Addition of more than 0.7 equivalents of Hg(ii) gives rise to line broadening, presumably reflecting aggregation. This is further supported by ThT fluorescence studies showing that the Bri2-23 peptide does not aggregate over 24 hours, while addition of over 0.7 equivalents of Ag(i) or Hg(ii) leads to increase of fluorescence, indicating that these metal ions induce aggregation. Thus, a model integrating all data into a coherent picture is that the metal ion binding to the two thiolates gives rise to folding of the peptide into a structure that is prone to aggregation, forming aggregates with a considerable amount of β-sheets. Molecular dynamics simulations initiated with structures that agree with NMR data additionally support this model.

Influence of Ion Specificity and Concentration on the Conformational Transition of Intrinsically Disordered Sheep Prion Peptide

“…Much of the early works support that multivalent cations such as Cu 2+ , Zn 2+ , Ca 2+ , Mn 2+ , Fe 3+ promote the aggregation‐prone molten globule state of IDPs by binding with the charged amino acids at the protein surface [32–36] or increasing the mean residence time of hydration‐shell waters [37] . Additionally, the transitional metal ions such as Hg 2+ are reported to favor the transition of the unstructured coil to β‐sheet conformation for IDPs like tau protein [38] . Moreover, Cu 2+ and Zn 2+ are also reported to exhibit a significant effect on prion misfolding and the formation of toxic oligomeric species [39,40] .…”

“…[37] Additionally, the transitional metal ions such as Hg 2 + are reported to favor the transition of the unstructured coil to βsheet conformation for IDPs like tau protein. [38] Moreover, Cu 2 + and Zn 2 + are also reported to exhibit a significant effect on prion misfolding and the formation of toxic oligomeric species. [39,40] Thus, the patients with neurodegenerative disorders are found to have elevated concentrations of divalent metal ions (metal dyshomeostasis) in brain and body fluids.…”

TDPAC andβ-NMR applications in chemistry and biochemistry

“…TDPAC spectroscopy was applied to study the metal-binding features of various metalloproteins using the 199m Hg isotope as a spectroscopic probe substituting the native metal ions at the probed metal sites. Studies have been performed with the Hg II -substituted rubredoxin [49] and in the systems of Hg II /Cd II and various zinc-finger model oligopeptides [50], the Hg II -binding of the human metallochaperone HAH1 [51] as well as the 23-mer oligopeptide fragment cleaved off near the C-terminus of the transmembrane protein BRI2 (BRI2-23) [52]. The first two of these examples are discussed here and the latter two in the following sections.…”

“…Cu, Zn, Fe) are believed to play various roles in neurodegeneration, however, the metal binding ability of BRI2-23, possessing several potential metal coordinating amino acid residues, including two cysteines at position 5 and 22, is largely unexplored. In this multi-method study, Hg II and Ag I ions were used to probe the Cu I -binding characteristics of BRI2-23 and the effect of the metal ion on the folding and aggregation of the oligopeptide molecules [52]. BRI2-23 inherently possesses a disordered, random coil structure and despite the relatively large distance between the two Cys residues, Hg II binding was observed in a broad pH-range, when the metal ion-peptide ratio was not more than 0.5: 1.…”

An Extremely Stable Interprotein Tetrahedral Hg(Cys)4 Core Forms in the Zinc Hook Domain of Rad50 Protein at Physiological pH

“…[21] Intermolecular Hg II binding to macromolecules is usually related to the toxic activity of the metal and mechanistically relies on the formation of aggregates. Also here, in most cases the metal is majorly bound to thiolate groups, [22][23][24] though the metal promoted aggregation may as well be fulfilled through binding to other medium/soft donors such as imidazole nitrogen. [25] Formation of digonal Hg II species is observed regardless of the accessibility of potential binding sites.…”