Because silver is toxic to microbes, but not considered toxic to humans, the metal has been used as an antimicrobial agent since ancient times. Today, silver nanoparticles and colloidal silver are used for antibacterial purposes, and silver-peptide and similar complexes are being developed as therapeutic agents. Yet, the health effects of silver exposure are not fully understood, nor are the molecular details of silver-protein interactions. In Alzheimer’s disease, the most common form of dementia worldwide, amyloid-β (Aβ) peptides aggregate to form soluble oligomers that are neurotoxic. Here, we report that monovalent silver ions (Ag⁺) bind wildtype Aβ₄₀ peptides with a binding affinity of 25 ± 12 µM in MES buffer at 20 °C. Similar binding affinities are observed for wt Aβ₄₀ peptides bound to SDS micelles, for an Aβ₄₀(H6A) mutant, and for a truncated Aβ(4–40) variant containing an ATCUN (Amino Terminal Cu and Ni) motif. Weaker Ag⁺ binding is observed for the wt Aβ₄₀ peptide at acidic pH, and for an Aβ₄₀ mutant without histidines. These results are compatible with Ag⁺ ions binding to the N-terminal segment of Aβ peptides with linear bis-his coordination. Because the Ag⁺ ions do not induce any changes in the size or structure of Aβ₄₂ oligomers, we suggest that Ag⁺ ions have a minor influence on Aβ toxicity.

Human calcitonin is a 32-residue peptide hormone that binds to the calcitonin receptor (CTR) and is involved in calcium regulation. The amino acid sequence displays a hydrophilic central segment flanked by hydrophobic C- and N-terminal regions with a net charge of zero at neutral pH. This makes the molecule amphiphilic and conformationally flexible, and different CTR variants preferentially recognize different structural conformations of calcitonin. The peptide is secreted from the thyroid gland and is overproduced in some forms of thyroid cancer and can then form cell-toxic aggregates. Characterizing the structural properties of calcitonin under different conditions is, therefore, important for understanding its receptor-binding and self-aggregation properties. Here, we used circular dichroism (CD) spectroscopy to monitor the secondary structure of human calcitonin in different environments. Calcitonin monomers were found to display a random coil structure with a significant amount of PPII-helix components in phosphate buffer, pH 7.3, at physiological temperatures. When agitated, the peptide formed soluble aggregates over time with mainly an antiparallel β-sheet secondary structure. In the presence of micelles of differently charged surfactants, monomeric calcitonin formed a pure α-helix structure with cationic CTAB, a combination of α-helix and β-sheet with anionic SDS and with zwitterionic SB3-14, and remained mainly random coil with noncharged DDM. Thus, the charge of the surfactant headgroup was found to be an important parameter for calcitonin’s interactions with membrane-mimicking micelles. Similar but not identical interactions with the surfactants were observed under the oxidizing and reducing conditions.

Interactions between molecules are fundamental in biology. They occur also between amyloidogenic peptides or proteins that are associated with different amyloid diseases, which makes it important to study the mutual influence of two polypeptides on each other's properties in mixed samples. However, addressing this research question with imaging techniques faces the challenge to distinguish different polypeptides without adding artificial probes for detection. Here, we show that nanoscale infrared spectroscopy in combination with C-13, N-15-labeling solves this problem. We studied aggregated amyloid-& beta; peptide (A & beta;) and its interaction with an inhibitory peptide (NCAM1-PrP) using scattering-type scanning near-field optical microscopy. Although having similar secondary structure, labeled and unlabeled peptides could be distinguished by comparing optical phase images taken at wavenumbers characteristic for either the labeled or the unlabeled peptide. NCAM1-PrP seems to be able to associate with or to dissolve existing A & beta; fibrils because pure A & beta; fibrils were not detected after mixing. Interactions of proteins or polypeptides with different secondary structures can be studied in a mixture by nanoscale infrared spectroscopy, however, this technique remains challenging for polypeptides with similar secondary structures. Here, the authors demonstrate clear discrimination of two polypeptides from a mixture by scattering-type scanning near-field optical microscopy when one of the components is labeled with C-13- and N-15-isotopes.

Uranium (U) is naturally present in ambient air, water, and soil, and depleted uranium (DU) is released into the environment via industrial and military activities. While the radiological damage from U is rather well understood, less is known about the chemical damage mechanisms, which dominate in DU. Heavy metal exposure is associated with numerous health conditions, including Alzheimer’s disease (AD), the most prevalent age-related cause of dementia. The pathological hallmark of AD is the deposition of amyloid plaques, consisting mainly of amyloid-β (Aβ) peptides aggregated into amyloid fibrils in the brain. However, the toxic species in AD are likely oligomeric Aβ aggregates. Exposure to heavy metals such as Cd, Hg, Mn, and Pb is known to increase Aβ production, and these metals bind to Aβ peptides and modulate their aggregation. The possible effects of U in AD pathology have been sparsely studied. Here, we use biophysical techniques to study in vitro interactions between Aβ peptides and uranyl ions, UO₂²⁺, of DU. We show for the first time that uranyl ions bind to Aβ peptides with affinities in the micromolar range, induce structural changes in Aβ monomers and oligomers, and inhibit Aβ fibrillization. This suggests a possible link between AD and U exposure, which could be further explored by cell, animal, and epidemiological studies. General toxic mechanisms of uranyl ions could be modulation of protein folding, misfolding, and aggregation. 

Cell-penetrating peptides (CPPs) are highly promising transfection agents that can deliver various compounds into living cells, including nucleic acids (NAs). Positively charged CPPs can form non-covalent complexes with negatively charged NAs, enabling simple and time-efficient nanoparticle preparation. However, as CPPs have substantially different chemical and physical properties, their complexation with the cargo and characteristics of the resulting nanoparticles largely depends on the properties of the surrounding environment, i.e., solution. Here, we show that the solvent used for the initial dissolving of a CPP determines the properties of the resulting CPP particles formed in an aqueous solution, including the activity and toxicity of the CPP–NA complexes. Using different biophysical methods such as dynamic light scattering (DLS), atomic force microscopy (AFM), transmission and scanning electron microscopy (TEM and SEM), we show that PepFect14 (PF14), a cationic amphipathic CPP, forms spherical particles of uniform size when dissolved in organic solvents, such as ethanol and DMSO. Water-dissolved PF14, however, tends to form micelles and non-uniform aggregates. When dissolved in organic solvents, PF14 retains its α-helical conformation and biological activity in cell culture conditions without any increase in cytotoxicity. Altogether, our results indicate that by using a solvent that matches the chemical nature of the CPP, the properties of the peptide–cargo particles can be tuned in the desired way. This can be of critical importance for in vivo applications, where CPP particles that are too large, non-uniform, or prone to aggregation may induce severe consequences.

Misfolding of the cellular prion protein (PrP^C) is associated with the development of fatal neurodegenerative diseases called transmissible spongiform encephalopathies (TSEs). Metal ions appear to play a crucial role in PrP^C misfolding. PrP^C is a combined Cu(II) and Zn(II) metal-binding protein, where the main metal-binding site is located in the octarepeat (OR) region. Thus, the biological function of PrP^C may involve the transport of divalent metal ions across membranes or buffering concentrations of divalent metal ions in the synaptic cleft. Recent studies have shown that an excess of Cu(II) ions can result in PrP^C instability, oligomerization, and/or neuroinflammation. Here, we have used biophysical methods to characterize Cu(II) and Zn(II) binding to the isolated OR region of PrP^C. Circular dichroism (CD) spectroscopy data suggest that the OR domain binds up to four Cu(II) ions or two Zn(II) ions. Binding of the first metal ion results in a structural transition from the polyproline II helix to the β-turn structure, while the binding of additional metal ions induces the formation of β-sheet structures. Fluorescence spectroscopy data indicate that the OR region can bind both Cu(II) and Zn(II) ions at neutral pH, but under acidic conditions, it binds only Cu(II) ions. Molecular dynamics simulations suggest that binding of either metal ion to the OR region results in the formation of β-hairpin structures. As the formation of β-sheet structures can be a first step toward amyloid formation, we propose that high concentrations of either Cu(II) or Zn(II) ions may have a pro-amyloid effect in TSE diseases.

Alzheimer's disease (AD) is the most common cause of dementia worldwide. AD brains display deposits of insoluble amyloid plaques consisting mainly of aggregated amyloid-beta (A beta) peptides, and A beta oligomers are likely a toxic species in AD pathology. AD patients display altered metal homeostasis, and AD plaques show elevated concentrations of metals such as Cu, Fe, and Zn. Yet, the metal chemistry in AD pathology remains unclear. Ni(II) ions are known to interact with A beta peptides, but the nature and effects of such interactions are unknown. Here, we use numerous biophysical methods-mainly spectroscopy and imaging techniques-to characterize A beta/Ni(II) interactions in vitro, for different A beta variants: A beta(1-40), A beta(1-40)(H6A, H13A, H14A), A beta(4-40), and A beta(1-42). We show for the first time that Ni(II) ions display specific binding to the N-terminal segment of full-length A beta monomers. Equimolar amounts of Ni(II) ions retard A beta aggregation and direct it towards non-structured aggregates. The His6, His13, and His14 residues are implicated as binding ligands, and the Ni(II)center dot A beta binding affinity is in the low mu M range. The redox-active Ni(II) ions induce formation of dityrosine cross-links via redox chemistry, thereby creating covalent A beta dimers. In aqueous buffer Ni(II) ions promote formation of beta sheet structure in A beta monomers, while in a membrane-mimicking environment (SDS micelles) coil-coil helix interactions appear to be induced. For SDS-stabilized A beta oligomers, Ni(II) ions direct the oligomers towards larger sizes and more diverse (heterogeneous) populations. All of these structural rearrangements may be relevant for the A beta aggregation processes that are involved in AD brain pathology.

Mercury intoxication typically produces more severe outcomes in people with the APOE-ε4 gene, which codes for the ApoE4 variant of apolipoprotein E, compared to individuals with the APOE-ε2 and APOE-ε3 genes. Why the APOE-ε4 allele is a risk factor in mercury exposure remains unknown. One proposed possibility is that the ApoE protein could be involved in clearing of heavy metals, where the ApoE4 protein might perform this task worse than the ApoE2 and ApoE3 variants. Here, we used fluorescence and circular dichroism spectroscopies to characterize the in vitro interactions of the three different ApoE variants with Hg(I) and Hg(II) ions. Hg(I) ions displayed weak binding to all ApoE variants and induced virtually no structural changes. Thus, Hg(I) ions appear to have no biologically relevant interactions with the ApoE protein. Hg(II) ions displayed stronger and very similar binding affinities for all three ApoE isoforms, with K_D values of 4.6 μM for ApoE2, 4.9 μM for ApoE3, and 4.3 μM for ApoE4. Binding of Hg(II) ions also induced changes in ApoE superhelicity, that is, altered coil–coil interactions, which might modify the protein function. As these structural changes were most pronounced in the ApoE4 protein, they could be related to the APOE-ε4 gene being a risk factor in mercury toxicity.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by motor neuron loss and widespread muscular atrophy. Despite intensive investigations on genetic and environmental factors, the cause of ALS remains unknown. Recent data suggest a role for metal exposures in ALS causation. In this study we present a patient who developed ALS after a traditional medical procedure in Kenya. The procedure involved insertion of a black metal powder into several subcutaneous cuts in the lower back. Four months later, general muscle weakness developed. Clinical and electrophysiological examinations detected widespread denervation consistent with ALS. The patient died from respiratory failure less than a year after the procedure. Scanning electron microscopy and X-ray diffraction analyses identified the black powder as potassium permanganate (KMnO4). A causative relationship between the systemic exposure to KMnO4 and ALS development can be suspected, especially as manganese is a well-known neurotoxicant previously found to be elevated in cerebrospinal fluid from ALS patients. Manganese neurotoxicity and exposure routes conveying this toxicity deserve further attention.

Alzheimer’s disease (AD) is an incurable disease and the main cause of age-related dementia worldwide, despite decades of research. Treatment of AD with lithium (Li) has shown promising results, but the underlying mechanism is unclear. The pathological hallmark of AD brains is deposition of amyloid plaques, consisting mainly of amyloid-β (Aβ) peptides aggregated into amyloid fibrils. The plaques contain also metal ions of e.g. Cu, Fe, and Zn, and such ions are known to interact with Aβ peptides and modulate their aggregation and toxicity. The interactions between Aβ peptides and Li+ions have however not been well investigated. Here, we use a range of biophysical techniques to characterize in vitro interactions between Aβ peptides and Li+ions. We show that Li+ions display weak and non-specific interactions with Aβ peptides, and have minor effects on Aβ aggregation. These results indicate that possible beneficial effects of Li on AD pathology are not likely caused by direct interactions between Aβ peptides and Li+ions.