Real‐time Amyloid Aggregation Monitoring with a Photonic Crystal‐based Approach

We propose the application of a new label‐free optical technique based on photonic nanostructures to real‐time monitor the amyloid‐beta 1‐42 (Aβ(1‐42)) fibrillization, including the early stages of the aggregation process, which are related to the onset of the Alzheimer’s Disease (AD). The aggregation of Aβ peptides into amyloid fibrils has commonly been associated with neuronal death, which culminates in the clinical features of the incurable degenerative AD. Recent studies revealed that cell toxicity is determined by the formation of soluble oligomeric forms of Aβ peptides in the early stages of aggregation. At this phase, classical amyloid detection techniques lack in sensitivity. Upon a chemical passivation of the sensing surface by means of polyethylene glycol, the proposed approach allows an accurate, real‐time monitoring of the refractive index variation of the solution, wherein Aβ(1‐42) peptides are aggregating. This measurement is directly related to the aggregation state of the peptide throughout oligomerization and subsequent fibrillization. Our findings open new perspectives in the understanding of the dynamics of amyloid formation, and validate this approach as a new and powerful method to screen aggregation at early stages.     

Attenuation of the Aggregation and Neurotoxicity of Amyloid‐β Peptides by Catalytic Photooxygenation

Alzheimer’s disease (AD), a progressive severe neurodegenerative disorder, is currently incurable, despite intensive efforts worldwide. Herein, we demonstrate that catalytic oxygenation of amyloid‐β peptides (Aβ) might be an effective approach to treat AD. Aβ1–42 was oxygenated under physiologically‐relevant conditions (pH 7.4, 37 °C) using a riboflavin catalyst and visible light irradiation, with modifications at the Tyr¹⁰, His¹³, His¹⁴, and Met³⁵ residues. The oxygenated Aβ1–42 exhibited considerably lower aggregation potency and neurotoxicity compared with native Aβ. Photooxygenation of Aβ can be performed even in the presence of cells, by using a selective flavin catalyst attached to an Aβ‐binding peptide; the Aβ cytotoxicity was attenuated in this case as well. Furthermore, oxygenated Aβ1–42 inhibited the aggregation and cytotoxicity of native Aβ.     

Hydrophobic Effect: Solubility of Non-polar Substances in Water,Protein Denaturation and Micelle Formation

The 'hydrophobic effect' of the dissolution process of non-polar substances in water has been analysed under the light of a statistical thermodynamic molecular model. The model, based on the distinction between non-reacting and reacting systems explains the changes of the thermodynamic functions with temperature in aqueous systems. In the dissolution of non-polar substances in water, it follows from the model that the enthalpy change can be expressed as a linear function of the temperature (ΔH _app =ΔH ^ø +n _w C _p,w T ). Experimental solubility and calorimetric data of a large number of non-polar substances nicely agree with the expected function. The specific contribution of n _w solvent molecules depends on the size of solute and is related to destructuring (n _w >0) of water molecules around the solute. Then the study of 'hydrophobic effect' has been extended to the protein denaturation and micelle formation. Denaturation enthalpy either obtained by van't Hoff equation or by calorimetric determinations again depends linearly upon denaturation temperature, with denaturation enthalpy, ΔH _den , increasing with T . A portion of reaction enthalpy is absorbed by a number n _w of water molecules (n _w >0) relaxed in space around the denatured moieties. In micellization, an opposite process takes place with negative number of restructured water molecules (n _w <0) because the hydrophobic moieties of the molecules joined by hydrophobic affinity occupy a smaller cavity.     

Characterization of New Specific Copper Chelators as Potential Drugs for the Treatment of Alzheimer’s Disease

The non‐controlled redox‐active metal ions, especially copper, in the brain of patients with Alzheimer disease (AD) should be considered at the origin of the intense oxidative damage in the AD brain. Several bis(8‐aminoquinoline) ligands, such as 1 and PA1637, are able to chelate Cu²⁺ with high affinity, and are specific chelators of copper with respect to iron and zinc. They are able to efficiently extract Cu²⁺ from a metal‐loaded amyloid. In addition, these tetradentate ligands are specific for the chelation of Cu²⁺ compared with Cu⁺. Consequently, the copper ion is easily released from the bis(8‐aminoquinoline) ligand under reductive conditions, and can be trapped again by a protein having some affinity for copper such as human serum albumin (HSA) proteins. In addition, the copper is not efficiently released from [Cu(CQ)₂] in reductive conditions. The catalytic production of H₂O₂ by [Cu²⁺‐Aβ_1?28]/ascorbate is inhibited in vitro by the bis(8‐aminoquinoline) 1 , suggesting that 1 should be able to play a protective role against oxidative damages induced by copper‐loaded amyloids.     

用前沿色谱法测定蛋白质吸附等温线时增浓方式的研究

用连续前沿色谱法和断续前沿色谱法测定了七种标准蛋白在疏水色谱填料上的吸附等温线,两种方法之间存在着一定的差别。从流出曲线突跃处斜率测定的不确定性、色谱过程中动力学因素和实验方法本身存在的问题等几个方面探讨了误差的来源,指出了可能解决这一问题的途径。     

Investigation of Intrinsically Disordered Proteins through Exchange with Hyperpolarized Water

Hyperpolarized water can selectively enhance NMR signals of rapidly exchanging protons in osteopontin (OPN), a metastasis‐associated intrinsically disordered protein (IDP), at near‐physiological pH and temperature. The transfer of magnetization from hyperpolarized water is limited to solvent‐exposed residues and therefore selectively enhances signals in ¹H‐¹⁵N correlation spectra. Binding to the polysaccharide heparin was found to induce the unfolding of preformed structural elements in OPN.     

Influence of Aromatic Residues on the Material Characteristics of Aβ Amyloid Protofibrils at the Atomic Scale

Amyloid fibrils, which cause a number of degenerative diseases, are insoluble under physiological conditions and are supported by native contacts. Recently, the effects of the aromatic residues on the Aβ amyloid protofibril were investigated in a ThT fluorescence study. However, the relationship between the material characteristics of the Aβ protofibril and its aromatic residues has not yet been investigated on the atomic scale. Here, we successfully constructed wild‐type (WT) and mutated types of Aβ protofibrils by using molecular dynamics simulations. Through principle component analysis, we established the structural stability and vibrational characteristics of F20L Aβ protofibrils and compared them with WT and other mutated models such as F19L and F19LF20L. In addition, structural stability was assessed by calculating the elastic modulus, which showed that the F20L model has higher values than the other models studied. From our results, it is shown that aromatic residues influence the structural and material characteristics of Aβ protofibrils.     

Constrained Peptides with Target‐Adapted Cross‐Links as Inhibitors of a Pathogenic Protein–Protein Interaction

Bioactive conformations of peptides can be stabilized by macrocyclization, resulting in increased target affinity and activity. Such macrocyclic peptides proved useful as modulators of biological functions, in particular as inhibitors of protein–protein interactions (PPI). However, most peptide‐derived PPI inhibitors involve stabilized α‐helices, leaving a large number of secondary structures unaddressed. Herein, we present a rational approach towards stabilization of an irregular peptide structure, using hydrophobic cross‐links that replace residues crucially involved in target binding. The molecular basis of this interaction was elucidated by X‐ray crystallography and isothermal titration calorimetry. The resulting cross‐linked peptides inhibit the interaction between human adaptor protein 14‐3‐3 and virulence factor exoenzyme S. Taking into consideration that irregular peptide structures participate widely in PPIs, this approach provides access to novel peptide‐derived inhibitors.     

Molecular Structure of Amyloid Fibrils Formed by Residues 127 to 147 of the Human Prion Protein

Amyloid fibrils are filamentous and insoluble forms of peptides or proteins. Proline has long been considered to be incompatible with the cross‐β structural motif of amyloid fibrils. On the basis of solid‐state NMR spectroscopy data, we present a structural model of an in‐register parallel β sheet for the amyloid fibrils formed from a human prion protein fragment, huPrP_127–47. We have developed a simple solid‐state NMR spectroscopy technique to identify solvent‐protected backbone amide protons in a H/D exchange experiment without disaggregating the amyloid fibrils, from which we find that proline residue P₁₃₇ does not disrupt the β‐sheet structure from G₁₂₇ to G₁₄₂. We suggest that the resultant kink at P₁₃₇ generates a twist between adjacent peptide strands to maintain hydrogen bonding in the β‐sheet regions flanking the P₁₃₇ residue. Although proline can be well integrated into the cross‐β structure of amyloid fibrils, the kink formed at the position of the proline residue will considerably weaken the hydrogen bonding between the neighboring strands, especially when the mutation site is near the central region of a β sheet.