Native Glucagon Amyloids Catalyze Key Metabolic Reactions.

Glucagon is a prominent peptide hormone, playing central roles in the regulation of glucose blood-level and lipid metabolism. Formation of glucagon amyloid fibrils has been previously reported, although no biological functions of such fibrils are known. Here, we demonstrate that glucagon amyloid fibrils catalyze biologically important reactions, including esterolysis, lipid hydrolysis, and dephosphorylation. In particular, we found that glucagon fibrils catalyze dephosphorylation of adenosine triphosphate (ATP), a core metabolic reaction in cell biology. Comparative analysis of several glucagon variants allowed mapping the catalytic activity to an enzymatic pocket-like triad formed at the glucagon fibril surface, comprising the histidyl-serine domain at the N-terminus of the peptide. This study may point to previously unknown physiological roles and pathological consequences of glucagon fibrillation and supports the hypothesis that catalytic activities of native amyloid fibrils play functional roles in human physiology and disease.

Amyloid fishing: ß-Amyloid adsorption using tailor-made coated titania nanoparticles.

Amyloidoses are a family of diseases characterized by abnormal protein folding that leads to fibril aggregates, amyloids. Extensive research efforts are devoted to developing inhibitors to amyloid aggregates. Here we set to explore functionalized titania (TiO2) nanoparticles (NPs) as potential amyloid inhibiting agents. TiO2 NPs were coated by a catechol derivative, dihydroxy-phenylalanine propanoic acid (DPA), and further conjugated to the amyloids' specific dye Congo-Red (CR). TiO2-DPA-CR NPs were found to target mature fibrils of ß-amyloid (Aß). Moreover, coated NPs incubated with Aß proteins suppressed amyloid fibrillation. TiO2-DPA-CR were found to target amyloids in solution and induce their sedimentation upon centrifugation. This work demonstrates the potential utilization of TiO2-DPA NPs for labeling and facilely separating from solution mature amyloid fibrils.

Revisiting thioflavin T (ThT) fluorescence as a marker of protein fibrillation - The prominent role of electrostatic interactions.

Thioflavin T (ThT), a benzothiazole-based fluorophore, is a prominent dye widely employed for monitoring amyloid fibril assembly. Despite the near-universal presumption that ThT binds to ß-sheet domains upon fibrillar surface via hydrophobic forces, the contribution of the positive charge of ThT to fibril binding and concomitant fluorescence enhancement have not been thoroughly assessed. Here we demonstrate a considerable interdependence between ThT fluorescence and electrostatic charges of peptide fibrils. Specifically, by analyzing both fibril-forming synthetic peptides and prominent natural fibrillar peptides, we demonstrate pronounced modulations of ThT fluorescence signal that were solely dependent upon electrostatic interactions between ThT and peptide surface. The results further attest to the fact that fibril ζ-potential rather than pH-dependent assembly of the fibrils constitute the primary factor affecting ThT binding and fluorescence. This study provides the first quantitative assessment of electrostatically driven ThT fluorescence upon adsorption to amyloid fibrils.

Unravelling the role of amino acid sequence order in the assembly and function of the amyloid-ß core.

The amino acid sequence plays an essential role in amyloid formation. Here, using the central core recognition module of the Aß peptide and its reverse sequence, we show that although both peptides assemble into ß-sheets, their morphologies, kinetics and cell toxicities display marked differences. In addition, the native peptide, but not the reverse one, shows notable affinity towards bilayer lipid model membranes that modulates the aggregation pathways to stabilize the oligomeric intermediate states and function as the toxic agent responsible for neuronal dysfunction.

Deciphering the Rules for Amino Acid Co-Assembly Based on Interlayer Distances.

Metabolite materials are extremely useful to obtain functional bioinspired assemblies with unique physical properties for various applications in the fields of material science, engineering, and medicine by self-assembly of the simplest biological building blocks. Supramolecular co-assembly has recently emerged as a promising extended approach to further expand the conformational space of metabolite assemblies in terms of structural and functional complexity. Yet, the design of synergistically co-assembled amino acids to produce tailor-made functional architectures is still challenging. Herein, we propose a design rule to predict the supramolecular co-assembly of naturally occurring amino acids based on their interlayer separation distances observed in single crystals. Using diverse experimental techniques, we demonstrate that amino acids with comparable interlayer separation strongly interact and co-assemble to produce structural composites distinctly different from their individual properties. However, such an interaction is hampered in a mixture of differentially layer-separated amino acids, which self-sort to generate individual characteristic structures. This study provides a different paradigm for the modular design of supramolecular assemblies based on amino acids with predictable properties.

Inhibitory Effect of Naphthoquinone-Tryptophan Hybrid towards Aggregation of PAP f39 Semen Amyloid.

PAP248⁻286, a 39 amino acid peptide fragment, derived from the prostatic acid phosphatase secreted in human semen, forms amyloid fibrils and facilitates the attachment of retroviruses to host cells that results in the enhancement of viral infection. Therefore, the inhibition of amyloid formation by PAP248⁻286 (termed PAP f39) may likely reduce HIV transmission in AIDS. In this study, we show that the naphthoquinone tryptophan (NQTrp) hybrid molecule significantly inhibited PAP f39 aggregation in vitro in a dose-dependent manner as observed from the ThT assay, ANS assay, and transmission electron microscopy imaging. We found that even at a sub-molar concentration of 20:1 [PAP f39:NQTrp], NQTrp could reduce >50% amyloid formation. NQTrp inhibition of PAP f39 aggregation resulted in non-toxic intermediate species as determined by the vesicle leakage assay. Isothermal titration calorimetry and molecular docking revealed that the binding of NQTrp and PAP f39 is spontaneous, and NQTrp predominantly interacts with the polar and charged residues of the peptide by forming hydrogen bonds and hydrophobic contacts with a strong binding energy. Collectively, these findings indicate that NQTrp holds significant potential as a small molecule inhibitor of semen amyloids.

Chiral modulation of amyloid beta fibrillation and cytotoxicity by enantiomeric carbon dots.

Enantiomeric carbon dots (C-dots) synthesized from l-lysine or d-lysine, modulate aggregation and cytotoxicity of amyloid beta-42 (Aß42), the primary constituent of the amyloid plaques associated with Alzheimer's disease. In particular, l-Lys-C-dots dramatically remodeled Aß42 secondary structure and fibril morphologies, as well as inhibited Aß42 cytotoxicity and membrane interactions.