Fernando Formaggio

University of Padova, Padua, Veneto, Italy

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Publications (441)1243.82 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Pulsed EPR methods, in particular PELDOR (or DEER), are very sensitive to the dipole···dipole interaction between electron spins in a pair of free radicals. Using PELDOR, the conformations of a number of double radical-containing biomolecules have been determined. In this review article we focused our attention on the application of this spectroscopy to nitroxide-labeled peptaibols. This is an emerging class of naturally-occurring, relatively short, linear, helical peptide molecules endowed with hydrophobic character, capability to interact with and to alter the structure of membranes, and antibiotic activity. We extracted detailed information on the secondary structures of specifically site-directed, double nitroxide-labeled peptaibols under a variety of experimental conditions, including biologically relevant environments. Moreover, we examined in depth peptaibol clustering, related to the marked propensity of these molecules to undergo self-association in model and whole-cell membrane systems, using mainly mono nitroxide-containing synthetic analogs. Finally, based on the PELDOR data accumulated, we proposed models of supramolecular (quaternary) structures of peptaibols and their binding modes to membranes. This article is protected by copyright. All rights reserved. © 2015 Wiley Periodicals, Inc.
    Biopolymers 08/2015; DOI:10.1002/bip.22713 · 2.39 Impact Factor
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    ABSTRACT: Among the various types of α-peptide folding motifs, δ-turn, which requires a central cis-amide disposition, has been one of the least extensively investigated. In particular, this main-chain reversal topology has been studied in-depth neither in linear/cyclic peptides nor in proteins. This Minireview article assembles and critically analyzes relevant data from a literature survey on the δ-turn conformation in those compounds. Unpublished results from recent conformational energy calculations and a preliminary solution-state analysis on a small model peptide, currently ongoing in our laboratories, are also briefly outlined. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
    Chemistry - A European Journal 08/2015; 21(40). DOI:10.1002/chem.201501467 · 5.73 Impact Factor
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    ABSTRACT: A helical hexapeptide was designed to link in a rigid parallel orientation to a gold surface. The peptide sequence of the newly synthesized compound is characterized by the presence of two 4-amino-1,2-dithiolane-4-carboxylic acid (Adt) residues (positions 1 and 4) to promote a bidentate interaction with the gold surface, two L-Ala residues (positions 2 and 5) and two-aminoisobutyric acid (Aib) residues (positions 3 and 6) to favor a high population of the 310-helix conformation. Furthermore, a ferrocenoyl (Fc) probe was inserted at the N-terminus to investigate the electronic conduction properties of the peptide. X-Ray Photoelectron Spectroscopy and Scanning Tunneling Microscopy techniques were used to characterize the binding of the peptide to the gold surface and the morphology of the peptide layer, respectively. Several electrochemical (Cyclic Voltammetry, Chronoamperometry, Square Wave Voltammetry) techniques were applied to analyze the electrochemical activity of the Fc probe, along with the influence of the peptide 3D-structure and the peptide layer morphology on electron transfer processes.
    Nanoscale 07/2015; 7(37). DOI:10.1039/C5NR03549J · 7.39 Impact Factor
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    ABSTRACT: A dual-action ligand targeting both integrin αVβ3 and vascular endothelial growth factor receptors (VEGFRs), was synthesized via conjugation of a cyclic peptidomimetic αVβ3 Arg-Gly-Asp (RGD) ligand with a decapentapeptide. The latter was obtained from a known VEGFR antagonist by acetylation at the Lys13 side chain. Functionalization of the precursor ligands was carried out in solution and in the solid phase, affording two fragments: an alkyne VEGFR ligand and the azide integrin αVβ3 ligand, which were conjugated by click chemistry. Circular dichroism studies confirmed that both the RGD and VEGFR ligand portions of the dual-action compound substantially adopt the biologically active conformation. In vitro binding assays on isolated integrin αVβ3 and VEGFR-1 showed that the dual-action conjugate retains a good level of affinity for both its target receptors, although with one order of magnitude (10/20 times) decrease in potency. The dual-action ligand strongly inhibited the VEGF-induced morphogenesis in Human Umbilical Vein Endothelial Cells (HUVECs). Remarkably, its efficiency in preventing the formation of new blood vessels was similar to that of the original individual ligands, despite the worse affinity towards integrin αVβ3 and VEGFR-1.
    ChemistryOpen 07/2015; 4(5). DOI:10.1002/open.201500062 · 2.11 Impact Factor
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    ABSTRACT: Total syntheses and complete characterizations of singly substituted PheCN -based analogs of alamethicin AlaP, which is active on model and natural membranes, and the TM peptide, which inserts in a trans-membrane orientation in lipid bilayers, are reported. The syntheses of the AlaP analogs were performed in solution, while those of TM and its analogs were carried out by solid phase. Using the cyanophenyl fluorescence and IR absorption probe, an in-depth investigation of the self-association, membrane-interacting, permeabilizing, and orientation properties of these peptides were conducted. The aromatic residue incorporated induces only a negligible modification to the properties of the parent peptides. The PheCN IR absorption band was located between 2228-2230 cm(-1) for all peptides, irrespective of the position of labeling. By contrast, as the width of this band varied significantly with the depth of probe insertion in the bilayer, it could represent a good marker of the PheCN position in phospholipid membranes. This article is protected by copyright. All rights reserved. © 2015 Wiley Periodicals, Inc.
    Biopolymers 05/2015; DOI:10.1002/bip.22674 · 2.39 Impact Factor
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    ABSTRACT: Two analogs of the ten-amino acid residue, membrane-active lipopeptaibiotic trichogin GA IV, mono-labeled with 4-cyano-α-methyl-L-phenylalanine, a potentially useful fluorescence and IR absorption probe of the local microenvironment, were synthesized by the solid-phase methodology and conformationally characterized. The single modification was incorporated either at the N-terminus (position 1) or near the C-terminus (position 8) of the peptide main chain. In both cases, the replaced amino acid was the equally helicogenic α-aminoisobutyric acid (Aib) residue. We performed a solution conformational analysis by use of FT-IR absorption, CD, and 2D-NMR spectroscopies. The results indicate that both labeled analogs essentially maintain the overall helical propensity of the naturally occurring lipopeptaibiotic. Peptidemembrane interactions were assessed by fluorescence and ATR-IR absorption techniques. Analogies and differences between the two peptides were highlighted. Taken together, our data confirm literature results that some of the spectroscopic parameters of the 4-cyanobenzyl chromophore are sensitive markers of the local microenvironment. Copyright © 2015 Verlag Helvetica Chimica Acta AG, Zürich.
    Chemistry & Biodiversity 04/2015; 12(4):513-27. DOI:10.1002/cbdv.201400404 · 1.52 Impact Factor

  • Physical Chemistry Chemical Physics 01/2015; DOI:10.1039/C5CP04136H · 4.49 Impact Factor
  • Hiroaki Maekawa · Gema Ballano · Fernando Formaggio · Claudio Toniolo · Nien-Hui Ge ·
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    ABSTRACT: We have used a combination of 2D IR spectroscopy with 13C═18O labeled amide-I and 15N-labeled amide-II modes to reveal how vibrational coupling between labeled peptide units depends on secondary structure. Linear and 2D IR measurements and simulations of Cα,α-diethylglycine homotetrapeptide show that this compound adopts the fully extended (2.05-helical) conformation in CDCl3, consistent with previous work on the Ac-capped peptide. The amide-I/II cross peaks of isotopomers exhibit only a marginal isotope frequency shift between labeled modes that are separated by two peptide units, indicating a very weak coupling. This result is in sharp contrast with a large cross-peak shift observed in 310-helical peptides, in which the labeled amide-I and -II modes are connected through an inter-residue C═O···H-N hydrogen bond. The discovered 3D-structural dependence indicates that the 13C═18O/15N labeled amide-I/II cross peaks can distinguish the formation of a single 310-helical turn from the fully extended polypeptide chain and increase the versatility of 2D IR spectroscopy as a conformational analysis tool of biomolecules.
    The Journal of Physical Chemistry C 12/2014; 118(50):29448-29457. DOI:10.1021/jp5091679 · 4.77 Impact Factor
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    ABSTRACT: In contrast to the extensively investigated β-turn conformation in peptides and proteins, single and multiple γ-turns have been poorly studied. Single and non-contiguous multiple γ-turns have been relatively often authenticated in small cyclic peptides, but these important peptide main-chain reversal motifs have been examined carefully neither in linear peptides nor in globular proteins. This Perspective article summarizes literature data on this aspect of peptide stereochemistry, expanding the discussion also to the rarely found, contiguous multiple γ-turns which generate incipient or fully-developed 2.27-(γ-) helices. Unpublished results of recent research activities on this topic ongoing in our laboratories are also briefly outlined.
    New Journal of Chemistry 11/2014; 39(5). DOI:10.1039/C4NJ01564A · 3.09 Impact Factor
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    ABSTRACT: Peptaibiotics, non-ribosomally synthetized peptides from various ascomycetes, are uniquely characterized by di-alkylated α-amino acids, a rigid helical conformation, and membrane permeation properties. Although generally considered antimicrobial peptides, peptaibiotics may display other toxicological properties, and their function is in many cases unknown. With the goal to define the biological activity and selectivity of the peptaibiotic trichogin GA IV from the human opportunist Trichomonas longibrachiatum we analyzed its membrane interaction, cytotoxic activity and antibacterial effect. Trichogin GA IV effectively killed several types of healthy and neoplastic human cells at doses (EC50%=4-6μM) lacking antibiotic effects on both Gram(-) and Gram(+) bacteria (MIC>64μM). The peptaibiotic distinctive C-terminal primary alcohol was found to cooperate with the N-terminal n-octanoyl group to permeate the membrane phospholipid bilayer and to mediate effective binding and active endocytosis of trichogin GA IV in eukaryotic cells, two steps essential for cell death induction. Replacement of one Gly with Lys plus the simultaneous esterification of the C-terminus, strongly increased trichogin GA IV anti-Gram(+) activity (MIC 1-4μM), but further mitigated its cytotoxicity on human cells.
    Biochimica et Biophysica Acta (BBA) - Biomembranes 10/2014; 1848(1). DOI:10.1016/j.bbamem.2014.10.005 · 3.84 Impact Factor
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    ABSTRACT: We prepared, by solution-phase methods, and fully characterized three analogs of the membrane-active peptaibiotic alamethicin F50/5, bearing a single trifluoroacetyl (Tfa) label at the N-terminus, at position 9 (central region) or at position 19 (C-terminus), and with the three Gln at positions 7, 18, and 19 replaced by Glu(OMe) residues. To add the Tfa label at position 9 or 19, a γ-trifluoroacetylated α,γ-diaminobutyric acid (Dab) residue was incorporated as a replacement for the original Val(9) or Glu(OMe)(19) amino acid. We performed a detailed conformational analysis of the three analogs (using FT-IR absorption, CD, 2D-NMR, and X-ray diffraction), which clearly showed that Tfa labeling does not introduce any dramatic backbone modification in the predominantly α-helical structure of the parent peptaibiotic. The results of an initial solid-state (19) F-NMR study on one of the analogs favor the conclusion that the Tfa group is a very promising reporter for the analysis of peptaibioticmembrane interactions. Finally, we found that the antimicrobial activities of the three newly synthesized analogs depend on the position of the Tfa label in the peptide sequence.
    Chemistry & Biodiversity 08/2014; 11(8):1163-91. DOI:10.1002/cbdv.201300394 · 1.52 Impact Factor
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    ABSTRACT: Oxo-dipeptides and thio-dipeptides are built via condensation between couples of amino acids and amino thioacids, the latter with the carbonyl oxygen replaced by an sp(2) sulfur. We explored via in silico methods (PBE0/6-31G(d,p) and PBE0/6-311G(d,p)) all the possible combinations and built 800 dipeptides, whose structures were fully optimized. Maps of condensation energies are presented to highlight optimal partners leading to stable dipeptides and critical situations for which lower stability or instability is predicted in terms of Gibbs reaction free energies. To validate the feasibility of our computational investigation, we synthesized and compared the stabilities of two thionated dimers, namely -Gly[Ψ(CSNH)]Gly- and -Phe[Ψ(CSNH)]Phe-, characterized by diverging physico-chemical properties. To the best of our knowledge, this is the first systematic analysis reported for dipeptides built from natural amino acids as well as for their corresponding thio-analogs.
    Physical Chemistry Chemical Physics 07/2014; DOI:10.1039/c4cp02680b · 4.49 Impact Factor
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    ABSTRACT: Interactions between peptides are relevant from a biomedical point of view, in particular for the role played by their aggregates in different important pathologies, and also because peptide aggregates represent promising scaffolds for innovative materials.In the present article, the aggregation properties of the homo-peptides formed by α-aminoisobutyric acid (U) residues are discussed. The peptides investigated have chain lengths between six and 15 residues and comprise benzyl and naphthyl groups at the N- and C-termini, respectively. Spectroscopic experiments and molecular dynamics simulations show that the shortest homo-peptide, constituted by six U, does not exhibit any tendency to aggregate under the conditions examined. On the other hand, the homologous peptide with 15 U forms very stable and compact aggregates in 70/30(v/v) methanol/water solution. Atomic force microscopy images indicate that these aggregates promote formation of long fibrils once they are deposited on a mica surface. The aggregation phenomenon is mainly due to hydrophobic interactions occurring between very stable helical structures, and the aromatic groups in the peptides seem to play a minor role. Copyright © 2014 European Peptide Society and John Wiley & Sons, Ltd.
    Journal of Peptide Science 07/2014; 20(7). DOI:10.1002/psc.2648 · 1.55 Impact Factor
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    ABSTRACT: Trichogin GA IV, an antimicrobial peptaibol, exerts its function by augmenting membrane permeability, but the molecular aspects of its pore-forming mechanism are still debated. Several lines of evidence indicate a ‘barrel-stave’ channel structure, similar to that of alamethicin, but the length of a trichogin helix is too short to span a normal bilayer. Herein, we present electrophysiology measurements in planar bilayers, showing that trichogin does form channels of a well-defined size (R=4.2⋅109 Ω; corresponding at least to a trimeric aggregate) that span the membrane and allow ion diffusion, but do not exhibit voltage-dependent rectification, unlike those of alamethicin.
    Chemistry & Biodiversity 07/2014; 11(7). DOI:10.1002/cbdv.201300334 · 1.52 Impact Factor
  • Andrea Orlandin · Fernando Formaggio · Antonio Toffoletti · Cristina Peggion ·
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    ABSTRACT: Three approaches for the chemical ligation of peptides to cotton fibers are described and compared. This investigation was encouraged by the need to create peptide-decorated natural textiles, furnished with useful properties (e.g. antimicrobial activity). IR absorption spectroscopy is proved to be an easy and fast method to check the covalent anchorage of a peptide to cotton, whereas for a quantitative determination, a UV absorption method was employed. We also analyzed the usefulness of electron paramagnetic resonance spectroscopy to characterize our peptide-cotton conjugates. Copyright © 2014 European Peptide Society and John Wiley & Sons, Ltd.
    Journal of Peptide Science 07/2014; 20(7). DOI:10.1002/psc.2659 · 1.55 Impact Factor
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    ABSTRACT: The PELDOR technique was used to obtain the spectra of distances between spin labels for mono and double TOAC substituted analogues of [Glu(OMe)(7,18,19)] alamethicin F50/5 (Alm') peptaibiotic on the surface of the organic sorbent Oasis HLB and in ethanol solution at 77 K. For the double-labeled Alm', the free radical probes are at positions 1 and 16 (Alm'1,16). The intra- and intermolecular contributions to the PELDOR time traces were separated, with regard to the fractality of the system studied. We established that on HLB the labeled Alm' molecules are prone to aggregation. The distance spectra for Alm'1,16 show that, in both adsorbed state and in ethanol solution, the peptaibiotic is predominantly folded in the α-helix conformation. We assign the asymmetry of the distance spectrum in both cases to the occurrence of an admixture of more elongated α/310-helical conformers. The portion of these conformers is higher for the peptide adsorbed on HLB. We speculate that both the broadening of the basic spectrum line at rmax = 2.0 nm and the increase in the contribution of elongated conformers might be associated with the spread of the peptaibiotic adsorption sites on HLB as compared with the more uniform Alm'1,16 trap structure in frozen ethanol solution. The aggregates of mono-labeled Alm'1 and Alm'16 also studied. The intermolecular distance spectrum for Alm'1 on HLB is shifted toward longer distances as compared with those of Alm'16. This result suggests that in the aggregates Alm' molecules are preferentially oriented with their C-terminal regions in the vicinity.
    The Journal of Physical Chemistry B 06/2014; 118(25). DOI:10.1021/jp503691n · 3.30 Impact Factor
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    ABSTRACT: Photoinduced electron transfer (PET) experiments have been carried out on peptide self-assembled monolayers (SAM) chemisorbed on a gold substrate. The oligopeptide building block was exclusively formed by Cα-tetrasubstituted α-aminoisobutyric residues to attain a helical conformation despite the shortness of the peptide chain. Furthermore, it was functionalized at the C-terminus by a pyrene chromophore to enhance the UV photon capture cross-section of the compound and by a lipoic group at the N-terminus for linking to gold substrates. Electron transfer across the peptide SAM has been studied by photocurrent generation experiments in an electrochemical cell employing a gold substrate modified by chemisorption of a peptide SAM as a working electrode and by steady state and time-resolved fluorescence experiments in solution and on a gold-coated glass. The results show that the electronic flow through the peptide bridge is strongly asymmetric, i.e. PET from the C-terminus to gold is highly favored with respect to PET in the opposite direction. This effect arises from the polarity of the Au-S linkage (Auδ+-Sδ-, junction effect) and from the electrostatic field generated by the peptide helix.
    The Journal of Physical Chemistry A 06/2014; 118(33). DOI:10.1021/jp503791w · 2.69 Impact Factor
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    ABSTRACT: In this article, we review the relevant results obtained during almost 60 years of research on a specific aspect of stereochemistry, namely handedness preference and switches between right-handed and left-handed helical peptide structures generated by protein amino acids or appropriately designed, side-chain modified analogs. In particular, we present and discuss here experimental and theoretical data on three categories of those screw-sense issues: (i) right-handed/left-handed α-helix transitions underwent by peptides rich in Asp, specific Asp β-esters, and Asn; (ii) comparison of the preferred conformations adopted by helical host–guest peptide series, each characterized by an amino acid residue (e.g. Ile or its diastereomer aIle) endowed with two chiral centers in its chemical structure; and (iii) right-handed (type I)/left-handed (type II) poly-(Pro)n helix transitions monitored for peptides rich in Pro itself or its analogs with a pyrrolidine ring substitution, particularly at the biologically important position 4. The unique modular and chiral properties of peptides, combined with their relatively easy synthesis, the chance to shape them into the desired conformation, and the enormous chemical diversity of their coded and non-coded α-amino acid building blocks, offer a huge opportunity to structural chemists for applications to bioscience and nanoscience problems. Copyright © 2014 European Peptide Society and John Wiley & Sons, Ltd.
    Journal of Peptide Science 05/2014; 20(5). DOI:10.1002/psc.2638 · 1.55 Impact Factor
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    ABSTRACT: Model peptides containing N-substituted 3-aminoazetidine-3-carboxylic acids have been synthesized to investigate the conformational features of these Cα-tetrasubstituted amino acids. The target peptides were designed and prepared by classical synthetic methods in solution, exploiting the convenient N-substituted 3-azidoazetidine-3-carboxylic acids as precursors. The conformational preferences of the newly synthesized peptides were investigated in solution by IR and NMR spectroscopy. It was observed that the 3-aminoazetidine-3-carboxylic acid moiety is likely a β-turn inducer. In addition, an interesting main-chain-to-side-chain hydrogen bond that forms a six-membered pseudo-cycle was detected. It connects the nitrogen (acceptor) of the azetidine ring to the amide NH (donor) of the immediately following residue. This unexpected hydrogen bond increases the number of conformational options offered by N-substituted 3-aminoazetidine-3-carboxylic acids when designing foldamers with new and predictable 3D structures.
    European Journal of Organic Chemistry 04/2014; 2014(11). DOI:10.1002/ejoc.201301741 · 3.07 Impact Factor
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    ABSTRACT: The aggregation properties of two Ala-based pentapeptides were investigated by spectroscopic techniques and molecular dynamics (MD) simulations. The two peptides, both functionalized at the N-terminus with a pyrenyl group, differ in the insertion of an α-aminoisobutyric acid residue at position 4. We showed that this single modification of the homo-peptide sequence inhibits the aggregation of the pentapeptide in aqueous solutions. Atomic force microscopy imaging revealed that the two peptides form mesoscopic aggregates of very different morphologies when deposited on mica. MD experiments showed that the two peptides have a very different propensity to form β-pleated sheet structures, as confirmed by our spectroscopic measurements. The implications of these findings for our understanding of the mechanism leading to the formation of amyloid structures, primary responsible for numerous neurodegenerative diseases, are also discussed.
    Soft Matter 03/2014; 10(15):2508-19. DOI:10.1039/c3sm52831f · 4.03 Impact Factor

Publication Stats

7k Citations
1,243.82 Total Impact Points


  • 1986-2015
    • University of Padova
      • Department of Chemical Sciences
      Padua, Veneto, Italy
    • Second University of Naples
      Caserta, Campania, Italy
  • 2002-2009
    • University of Rome Tor Vergata
      • Dipartimento di Scienze e Tecnologie Chimiche
      Roma, Latium, Italy
  • 2000-2009
    • National Research Council
      • • Institute of Biomolecular Chemistry ICB
      • • Laboratory of Organic Chemistry
      Roma, Latium, Italy
    • Russian Academy of Sciences
      • Institute of Chemical Kinetics and Combustion
      Moscow, Moscow, Russia
    • McMaster University
      • Department of Chemistry and Chemical Biology
      Hamilton, Ontario, Canada
  • 2007
    • Leiden University
      Leyden, South Holland, Netherlands
  • 2006-2007
    • It-Robotics
      Vicenza, Veneto, Italy
  • 1998
    • Lodz University of Technology
      • Institute of Organic Chemistry
      Łódź, Łódź Voivodeship, Poland
  • 1995
    • Osaka University of Pharmaceutical Sciences
      Ōsaka, Ōsaka, Japan
  • 1986-1995
    • University of Illinois at Chicago
      • Department of Chemistry
      Chicago, Illinois, United States
  • 1994
    • Université Bordeaux 1
      Talence, Aquitaine, France
  • 1992
    • Naples Eastern University
      Napoli, Campania, Italy