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Introduction: It is recently becoming clear that there is a significant and underestimated effect of metal coordination on the antimicrobial activity of antimicrobial peptides (AMPs) - potential treasure troves for the design of novel antibiotics. Beyond any doubt, understanding the molecular mechanisms underlying these disorders is a serious challenge for modern science. Aim: The way in which metal ions affect the mode of action of AMPs is far from being understood. They may alter the charge/structure/morphology of a given AMP and therefore enhance the AMP antimicrobial potency. The aim of the project is to understand the relationship between metal coordination, thermodynamic stability, structure, morphology and antimicrobial activity of AMPs. Material and methods: Achieving the overall aim requires a complex experimental approach. Mass spectrometry, potentiometry, ITC and a fruitful combination of nMR, UV-Vis, CD and EPR spectroscopic techniques are used to describe the beautiful bioinorganic chemistry of the studied systems. Antimicrobial assays and studies on regular cell lines establish the biological potency of the novel systems. Results: We have recently elucidated the relationship between Zn(II) coordination, structural, morphological change and antifungal activity of human amylin analogues (pramlintide) and for peptides from the shepherin group (from the plant Capsella bursa pastorsis). We observe a logical sequence of phenomena, each of which is the result of the previous one: (i) Zn(II) coordinates to histidyl imidazoles, (ii) causes a structural change of the peptide, which in turn (iii) results in the formation of fibrils. Conclusions: Ultimately, this chain of structural changes has (iv) biological consequences - the Zn(II)-pramlintide and Zn(II)-shepherin complex fibrils have a strong antifungal effect. A similar effect is not observed neither in the case of free ligands, nor in the case of their complexes with Cu(II). A working hypothesis on the mode of action of such Zn(II) complexes assumes that the formed fibrils can act as needles, physically damaging the fungal cell wall or membrane. [ABSTRACT FROM AUTHOR]

Introduction: Undoubtedly, neurodegenerative Diseases (nD) are one of the most serious diseases in modern society. Understanding the molecular mechanisms underlying these disorders is a challenge for global science. nD are still based on huge number of hypothesis, therefore it is important to find out the mechanism which leads to the pathogenic process. Amyloidogenic proteins are associated with severe neurodegenerative disorders afflicting millions of people worldwide. Aim: The misfolding process leading to protein aggregates is far to be completely understood. However, it is well accepted that many factors are able to influence the morphology and kinetics of amyloids formation. The aim of the project is the understanding of the bioinorganic chemistry of biologically significant copper complexes of the amyloidogenic proteins, in the presence of micelles, which mimic the lipid bilayer, to which the proteins are anchored in vivo. Material and methods: Understanding the coordination properties of the metalpeptide complexes will require a complex experimental approach. Mass spectrometry will be used to study the stoichiometry of metal-complex formation. Potentiometry was allow us to determine the thermodynamic parameters. In order to precisely identify the binding sites, the donor atoms and the coordination geometry of complex species formed in solution at given pH, a fruitful combination of nMR, UV-Vis, CD and EPR spectroscopic techniques have been used. Results: The coordination ability of the amyloidogenic fragment of human Prion Protein (hPrP) is significantly different in the aqueous solution when compared to that containing micelles. Our results prove that the metal binding mode strongly depends on the protein backbone structure. The α-helix structuring of the amyloid hPrP domain influences both the metal coordination sphere and the binding affinity. Conclusions: Essential metal ions are actively involved in several biological events associated with neurodegenerative disorders. The molecular understanding basis of the metal homeostasis and regulations in the cells are critical in identifying the underlying causes for diseases pathophysiology, providing proper diagnosis and treatments. It is also necessary for the development of new therapeutic agents able to treat and prevent their occurrence. [ABSTRACT FROM AUTHOR]

Consecutive histidine repeats are chosen both by nature and by molecular biologists due to their high affinity towards metal ions. Screening of the human genome showed that transcription factors are extremely rich in His tracts. In this work, we examine two of such His-rich regions from forkhead box and MAFA proteins-MB3 (contains 18 His) and MB6 (with 21 His residues), focusing on the affinity and binding modes of Cu and Zn towards the two His-rich regions. In the case of Zn species, the availability of imidazole nitrogen donors enhances metal complex stability. Interestingly, an opposite tendency is observed for Cu complexes at above physiological pH, in which amide nitrogens participate in binding. [ABSTRACT FROM AUTHOR]

Prion proteins (PrP) from different species have the ability to tightly bind Cu2+ ions. Copper coordination sites are located in the disordered and flexible N-terminal region which contains several His anchoring sites. Among them, two His residues are found in the so called amyloidogenic PrP region which is believed to play a key role in the process leading to oligomer and fibril formation. Both chicken and human amyloidogenic regions have a hydrophobic C-terminal region rich in Ala and Val amino acids. Recent findings revealed that this domain undergoes random coil to α-helix structuring upon interaction with membrane models. This interaction might strongly impact metal binding abilities either in terms of donor sets or affinity. In this study we investigated Cu2+ interaction with an amyloidogenic fragment, chPrP105–140, derived from chicken prion protein (chPrP), in different solution environments. The behavior of the peptide and its metal complexes was analyzed in water and in the presence of negative and positive charged membrane mimicking environments formed by sodium dodecyl sulfate (SDS) and dodecyl trimethyl ammonium chloride (DTAC) micelles. The metal coordination sphere, the metal binding affinity and stoichiometry were evaluated by combining spectroscopic and potentiometric methods. Finally we compare copper(ii) interactions with human and chicken amyloidogenic fragments. Our results indicate that the chicken amyloidogenic fragment is a stronger copper ligand than the human amyloidogenic fragment. [ABSTRACT FROM AUTHOR]

Prion diseases, known as Transmissible Spongiform Encephalopathies (TSEs), are a group of fatal neuronal, and to some extent infectious disorders, associated with a pathogenic protein agent called prion protein (PrP). The human prion protein (hPrP) fragment encompassing the 91–127 region, also known as the amyloidogenic domain, comprises two copper-binding sites corresponding to His-96 and His-111 residues that act as anchors for Cu2+ binding. In this work, we investigated Cu2+ interaction with hPrP91–127 in the presence of the anionic surfactant sodium dodecyl sulfate (SDS), which induces a partial α-helix folding of the peptide. Our data indicate that the Cu2+ coordination ability of the amyloidogenic fragment in the presence of SDS micelles is significantly different to that observed in aqueous solution. This is mainly due to the fact that SDS micelles strongly stabilize the formation of the α-helical structure of the peptide backbone, which is well conserved also upon Cu2+ binding, contrary to the random coil conformation mainly assumed by hPrP91–127 in aqueous solutions. Potentiometric and spectroscopic studies clearly indicate that in the case of SDS containing solutions, Cu2+ ions coordinate simultaneously to both imidazoles, while in the case of water solutions, metal ion coordination involves only a single His side chain, which individually acts as an independent Cu2+ anchoring site. [ABSTRACT FROM AUTHOR]

It is supposed that the presence of poly-His regions in close proximity to poly-Gly domains in snake venoms is related to their biological activity; poly-His/poly-Gly (pHpG) peptides inhibit the activity of metalloproteinases during venom storage via the chelation metal ions, necessary for their proper functioning. This work shows that only the histidyl residues from the N-terminal VDHDHDH motif (but not from the poly-His tag) were the primary Zn(II) binding sites and that the poly-Gly domain situated in the proximity of a central proline residue may play a regulatory role in venom gland protection. The proline induces a kink of the peptide, resulting in steric hindrance, which may modulate the accessibility of potential metal binding sites in the poly-His domain and may, in turn, be one of the regulators of Zn(II) accessibility in the venom gland and therefore a modulator of metalloproteinase activity during venom storage.

Prion proteins (PrP) from different species have the ability to tightly bind Cu 2+ ions. Copper coordination sites are located in the disordered and flexible N-terminal region which contains several His anchoring sites. Among them, two His residues are found in the so called amyloidogenic PrP region which is believed to play a key role in the process leading to oligomer and fibril formation. Both chicken and human amyloidogenic regions have a hydrophobic C-terminal region rich in Ala and Val amino acids. Recent findings revealed that this domain undergoes random coil to α-helix structuring upon interaction with membrane models. This interaction might strongly impact metal binding abilities either in terms of donor sets or affinity. In this study we investigated Cu 2+ interaction with an amyloidogenic fragment, chPrP105-140, derived from chicken prion protein (chPrP), in different solution environments. The behavior of the peptide and its metal complexes was analyzed in water and in the presence of negative and positive charged membrane mimicking environments formed by sodium dodecyl sulfate (SDS) and dodecyl trimethyl ammonium chloride (DTAC) micelles. The metal coordination sphere, the metal binding affinity and stoichiometry were evaluated by combining spectroscopic and potentiometric methods. Finally we compare copper(ii) interactions with human and chicken amyloidogenic fragments. Our results indicate that the chicken amyloidogenic fragment is a stronger copper ligand than the human amyloidogenic fragment.