Valentina Roviello

Hierarchical Analysis of Self-Assembled PEGylated Hexaphenylalanine Photoluminescent Nanostructures

Despite the growing literature about diphenylalanine-based peptide materials, it still remains a challenge to delineate the theoretical insight into peptide nanostructure formation and the structural features that could permit materials with enhanced properties to be engineered. Herein, we report the synthesis of a novel peptide building block composed of six phenylalanine residues and eight PEG units, PEG8 -F6. This aromatic peptide self-assembles in water in stable and well-ordered nanostructures with optoelectronic properties. A variety of techniques, such as fluorescence, FTIR, CD, DLS, SEM, SAXS, and WAXS allowed us to correlate the photoluminescence properties of the self-assembled nanostructures with the structural organization of the peptide building block at the micro- and nanoscale. Finally, a model of hexaphenylalanine in aqueous solution by molecular dynamics simulations is presented to suggest structural and energetic factors controlling the formation of nanostructures.

Photoluminescent Peptide‐Based Nanostructures as FRET Donor for Fluorophore Dye

A great interest has been recently generated by the discovery that peptide-based nanostructures (NSs) endowed with cross-β structure may show interesting photoluminescent (PL) properties. It was shown that NSs formed by PEGylated hexaphenylalanine (PEG8 -F6, PEG=polyethylene glycol) are able to emit at 460 nm when excited at 370 or 410 nm. Here, the possibility to transfer the fluorescence of these PEG8 -F6-based NSs by foster resonance electron transfer (FRET) phenomenon to a fluorescent dye was explored. To achieve this aim, the 4-chloro-7-nitrobenzofurazan (NBD) dye was encapsulated in these NSs. Structural data in solution and in solid state, obtained by a variety of techniques (circular dichroism, Fourier-transform infrared spectroscopy, wide-angle X-ray scattering, and small-angle X-ray scattering), indicated that the organization of the peptide spine of PEG8 -F6 NS, which consists of anti-parallel β-sheets separated by a dry interface made of interacting phenylalanine side chains, was maintained upon NBD encapsulation. The spectroscopic characterization of these NSs clearly showed a red-shift of the emission fluorescence peak both in solution and in solid state. This shift from 460 to 530 nm indicated that a FRET phenomenon from the peptide-based to the fluorophore-encapsulated NS occurred. FRET could also be detected in the PEG8 -F6 conjugate, in which the NBD was covalently bound to the amine of the compound. On the basis of these results, it is suggested that the red-shift of the intrinsic PL of NSs may be exploited in the bio-imaging field.

DNA- and RNA-binding ability of oligoDapT, a nucleobase-decorated peptide, for biomedical applications

Background Nucleobase-bearing peptides and their interaction with DNA and RNA are an important topic in the development of therapeutic approaches. On one hand, they are highly effective for modulating the nucleic-acid-based biological processes. On the other hand, they permit to overcome some of the main factors limiting the therapeutic efficacy of natural oligonucleotides, such as their rapid degradation by nucleases. Methods and results This article describes the synthesis and characterization of a novel thymine-bearing nucleoamino acid based on the l-diaminopropionic acid (l-Dap) and its solid phase oligomerization to α-peptides (oligoDapT), characterized using mass spectrometry, spectroscopic techniques, and scanning electron microscopy (SEM) analysis. The interaction of the obtained nucleopeptide with DNA and RNA model systems as both single strands (dA12, rA12, and poly(rA)) and duplex structures (dA12/dT12 and poly(rA)/poly(rU)) was investigated by means of circular dichroism (CD) and ultraviolet (UV) experiments. From the analysis of our data, a clear ability of the nucleopeptide to bind nucleic acids emerged, with oligoDapT being able to form stable complexes with both unpaired and double-stranded DNA and RNA. In particular, dramatic changes in the dA12/dT12 and poly(rA)/poly(rU) structures were observed as a consequence of the nucleopeptide binding. CD titrations revealed that multiple peptide units bound all the examined nucleic acid targets, with TLdap/A or TLdap/A:T(U) ratios >4 in case of oligoDapT/DNA and ~2 in oligoDapT/RNA complexes. Conclusion Our findings seem to indicate that Dap-based nucleopeptides are interesting nucleic acid binding-tools to be further explored with the aim to efficiently modulate DNA- and RNA-based biological processes.

Structural Characterization of PEGylated Hexaphenylalanine Nanostructures Exhibiting Green Photoluminescence Emission

Peptides containing aromatic residues are known to exhibit spontaneous phenomena of supramolecular organization into ordered nanostructures (NSs). In this work we studied the structural behavior and optoelectronic properties of new biocompatible materials obtained by the self-assembly of a series of hexaphenylalanines (F6) modified at the N terminus by a PEG chain of different lengths. PEG12 -F6, PEG18 -F6, and PEG24 -F6 peptides were synthesized by coupling sequentially two, three, or four units of amino-carboxy-PEG6 blocks, each one containing six oxyethylene repetitions. Changes in the length and composition of the PEG chain were found to modulate the structural organization of the phenylalanine-based nanostructures. An increase in the self-aggregation tendency was observed with longer PEG chains, whereas, independently of the PEG length, the peptide NSs display cross-β-like secondary structures with an antiparallel β-strand arrangement. WAXS/GIWAXS diffraction patterns indicate a progressive decrease in fiber order along the series. All the PEG-F6 derivatives present blue photoluminescent (PL) emission at 460 nm, with the adduct with the longest PEG chain (PEG24 -F6) showing an additional green emission at 530 nm.

Synthesis and biological evaluation of a novel Amadori compound.

Here, we report the synthesis, purification, ESI MS and NMR characterization, as well as the SEM analysis of a fructosyl thiophenyl-substituted triazolyl-thione l-alanine (denominated Fru-l-TTA). This novel fructosyl derivative was obtained by solution synthesis using the Amadori reaction, in analogy to other natural fructosyl-amino acids, and fully characterized. In particular, we report an accurate NMR/MS/SEM characterization of Fru-l-TTA alongside some biological properties, and investigated to compare the properties of the artificial derivative of this work with the natural counterparts. In particular, Fru-l-TTA shares with natural fructosyl-amino acids the possibility to inhibit the colony formation of prostate cancer cells and additionally decreases their migration.

Nucleic acid binding and other biomedical properties of artificial oligolysines

In the present study, we report the interaction of an artificial oligolysine (referred to as AOL) realized in our laboratory with targets of biomedical importance. These included polyinosinic acid (poly rI) and its complex with polycytidylic acid (poly I:C), RNAs with well-known interferon-inducing ability, and double-stranded (ds) DNA. The ability of the peptide to bind both single-stranded poly rI and ds poly I:C RNAs emerged from our circular dichroism (CD) and ultraviolet (UV) studies. In addition, we found that AOL forms complexes with dsDNA, as shown by spectroscopic binding assays and UV thermal denaturation experiments. These findings are encouraging for the possible use of AOL in biomedicine for nucleic acid targeting and oligonucleotide condensation, with the latter being a key step preceding their clinical application. Moreover, we tested the ability of AOL to bind to proteins, using serum albumin as a model protein. We demonstrated the oligolysine–protein binding by CD experiments which suggested that AOL, positively charged under physiological conditions, binds to the protein regions rich in anionic residues. Finally, the morphology characterization of the solid oligolysine, performed by scanning electron microscopy, showed different crystal forms including cubic-shaped crystals confirming the high purity of AOL.

Evaluating the biological properties of synthetic 4-nitrophenyl functionalized benzofuran derivatives with telomeric DNA binding and antiproliferative activities

Four 4-nitrophenyl-functionalized benzofuran (BF1, BF2) and benzodifuran (BDF1, BDF2) compounds were synthesized by a convenient route based on the Craven reaction. All the compounds underwent a detailed chemical-physical characterization to evaluate their structural, thermal and optical properties. An investigation on the therapeutic potential of the reported compounds was performed by analyzing their antiproliferative activity on prostatic tumour cells (PC-3). In both classes of compounds, anticancer potential is in direct correlation with the lipophilicity. From our study it emerged that antiproliferative activity was higher for benzofuran derivatives as compared to benzodifuran systems. Moreover, we report a mechanistic study relative to the most promising molecule, i.e. the apolar benzofuran BF1, that relates the antiproliferative properties found in our investigation to its ability to bind telomeric DNA (proven by CD and fluorescence techniques on tel22 G4 DNA), and highlights its unexpected impact on cell cycle progression.

Self-Assembling of Fmoc-GC Peptide Nucleic Acid Dimers into Highly Fluorescent Aggregates

The study of molecules that self-assemble through noncovalent interactions is one of the most attractive topics in supramolecular chemistry. The use of short peptides or modified nucleotides as building blocks for the aggregates is particularly intriguing because these are very easy to synthesize; moreover, subtle changes in the chemical structure of such building blocks may drastically affect the properties of the aggregates. The ability of peptide nucleic acids (PNA) to aggregate has been very little explored, despite its practical applications. In this work we investigated the self-assembling properties of a PNA dimer, conjugated at the N-terminus to a fluorenylmethoxycarbonyl group. This PNA dimer forms nano-aggregates at low concentration in CHCl3 /CH3 OH mixtures. The aggregates retain very interesting fluorescent properties (high quantum yield in the visible region with lifetimes on the nanosecond scale), which make them promising materials for applications in optoelectronics.

Lac-l-TTA, a novel lactose-based amino acid–sugar conjugate for anti-metastatic applications

Here we describe the synthesis, chromatographic purification, MS and NMR characterization of a new lactosyl-derivative, i.e. a lactosyl thiophenyl-substituted triazolyl-thione l-alanine (Lac-L-TTA). This amino acid–sugar conjugate was prepared by solution synthesis in analogy to the natural fructosyl-amino acids. Furthermore, we investigated the inhibition of PC-3 prostate cancer cell colony formation by this lactose derivative in comparison with the less polar fructose-based derivative, Fru-L-TTA. This let us to compare the properties of the artificial derivative, object of the present work, with the monosaccharide-based counterpart and to obtain a preliminary information on the influence of polarity on such biological activity. A significantly higher anticancer effect of Lac-L-TTA with respect to the fructose analogue emerged from our study suggesting that the anti-metastatic potential of fructosyl-amino acids can be enhanced by increasing the polarity of the compounds, for example by introducing disaccharide moieties in place of fructose.

Correction: Solid phase synthesis of TyrT, a thymine–tyrosine conjugate with poly(A) RNA-binding ability

Correction for ‘Solid phase synthesis of TyrT, a thymine–tyrosine conjugate with poly(A) RNA-binding ability’ by Giovanni N. Roviello et al., RSC Adv., 2016, 6, 27607–27613.