129 results on '"Notomista, E."'
Search Results
2. WS11.3 GVF27, HVA36 and IMY47: human-derived host defence peptides for alternative cystic fibrosis therapeutic intervention strategies
- Author
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Bosso, A., primary, Gaglione, R., additional, Pane, K., additional, Cafaro, V., additional, Arciello, A., additional, Notomista, E., additional, and Pizzo, E., additional
- Published
- 2018
- Full Text
- View/download PDF
3. A new cryptic host defense peptide identified in human 11-hydroxysteroid dehydrogenase-1 β-like: from in silico identification to experimental evidence
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Bosso, A., primary, Pirone, L., additional, Gaglione, R., additional, Pane, K., additional, Del Gatto, A., additional, Zaccaro, L., additional, Di Gaetano, S., additional, Diana, D., additional, Fattorusso, R., additional, Pedone, E., additional, Cafaro, V., additional, Haagsman, H.P., additional, van Dijk, A., additional, Scheenstra, M.R., additional, Zanfardino, A., additional, Crescenzi, O., additional, Arciello, A., additional, Varcamonti, M., additional, Veldhuizen, E.J.A., additional, Di Donato, A., additional, Notomista, E., additional, and Pizzo, E., additional
- Published
- 2017
- Full Text
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4. Identification of a new small bioactive peptide from Lactobacillus gasseri supernatant
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Zanfardino, A., primary, Criscuolo, G., additional, Di Luccia, B., additional, Pizzo, E., additional, Ciavatta, M.L., additional, Notomista, E., additional, Carpentieri, A., additional, Pezzella, A., additional, and Varcamonti, M., additional
- Published
- 2017
- Full Text
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5. Inhalable dry powders for chemically-modified human Cationic AntiMicrobial Peptides (CAMPs): moving toward in vivo application
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Notomista, E, primary and Ungaro, F, additional
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- 2015
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6. STRUCTURAL AND DYNAMIC CHARACTERIZATION OF TWO MUTANTS OF ONCONASE, A CANCER CHEMOTERAPEUTIC
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A. DI FIORE, CARANNANTE, A., NOTOMISTA, E., DI DONATO, MERLINO, ANTONELLO, SICA, FILOMENA, MAZZARELLA, LELIO, A., DI FIORE, Merlino, Antonello, Sica, Filomena, Mazzarella, Lelio, Carannante, A., Notomista, E., and Di, Donato
- Subjects
onconase ,anticancer agent - Published
- 2004
7. EFFECT OF POINT MUTATION OF THE HIGHLY RIGID STRUCTURE OF ONCONASE
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A. DI FIORE, CARANNANTE, A., NOTOMISTA, E., DI DONATO, MERLINO, ANTONELLO, SICA, FILOMENA, MAZZARELLA, LELIO, A., DI FIORE, Merlino, Antonello, Sica, Filomena, Mazzarella, Lelio, Carannante, A., Notomista, E., and Di, Donato
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Cancer chemiotherapy ,Onconase variant ,thermal stability - Abstract
This study provides the molecular bases to explain the lower catalytic activity of ONC compared to RNase A.
- Published
- 2004
8. Evolution of Bacterial Multicomponent Monooxygenases
- Author
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Notomista E., LAHM A., TRAMONTANO A., DI DONATO, ALBERTO, Notomista, E., Lahm, A., DI DONATO, Alberto, and Tramontano, A.
- Published
- 2003
9. Expression and purification of the recombinant subunits of toluene/ o-xylene monooxygenase and reconstitution of the active complex
- Author
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CAFARO V, SCOGNAMIGLIO R, VIGGIANI A, PASSARO I, NOTOMISTA E, DAL PIAZ F, AMORESANO A, CASBARRA A, PUCCI P. DI DONATO A., IZZO, VIVIANA, Cafaro, V, Scognamiglio, R, Viggiani, A, Izzo, Viviana, Passaro, I, Notomista, E, DAL PIAZ, F, Amoresano, A, Casbarra, A, and Pucci, P. DI DONATO A.
- Published
- 2002
10. Bioaugmentation of Novosphingobium sp. PP1Y in natural and artificial soils contaminated by PAHs and heavy metals
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Cafaro, V., Notomista, E., Izzo, Viviana, Troncone, L., Donadio, G., Tedesco, P., Di Donato, A., Cafaro, Valeria, Notomista, Eugenio, Izzo, V., Troncone, Luca, Donadio, Giuliana, Tedesco, P., and DI DONATO, Alberto
- Subjects
Novosphingobium sp. PP1Y ,PAHs and Heavy metals - Abstract
Polycyclic Aromatic Hydrocarbons (PAHs) are among the most widespread organic pollutants in the environment. PAHs are of great concern because some of them show strong mutagenicity and carcinogenity. Bioremediation, which involves the use of microorganisms to reduce the concentration of various chemical pollutants, has become nowadays one of the most promising approach for environmental clean-up. Unfortunately, the presence of heavy metals in environments contaminated by organic compounds often hamper the environmental restoration. Several species of bacteria have been isolated from polluted environments able to degrade recalcitrant compounds. We have recently characterized a novel Sphingomonadales, Novosphingobium sp. PP1Y, isolated from surface seawater samples collected inside the harbour of Pozzuoli (Naples, Italy). The strain PP1Y not only uses a surprisingly large number of mono and polycyclic aromatic compounds as the sole carbon and energy sources but it shows a very interesting and effective adaptation to grow on complex mixtures of aromatic compounds dissolved in oil phases like diesel-oil and gasoline. Novosphingobium sp. PP1Y showed the ability to form “biofilm” on several types of hydrophobic surfaces and, in water/oil systems. It is well known that biofilm formation make bacteria able to adhere to growing substrates and to confer the resistance towards heavy metals and organic pollutants. We investigated the biodegradation abilities of Novosphingobium sp. PP1Y in natural soil and compost, artificially contaminated by PAHs and heavy metals such as nickel, copper and zinc, in small-scale experiments. N. sp. PP1Y showed the ability to quickly remove phenanthrene from the compost after addition of the organic compound in crystal form or dissolved in oil phase. The bacterium was also able to metabolize phenanthrene in the presence of heavy metals, showing the most significant delay in phenanthrene removal only at high nickel concentration. When a mixture of PAHs in paraffin containing naphthalene, phenanthrene and pyrene, was added to the compost, the bacterium was able to remove all the three compounds, although with different rates. Furthermore, the addition of N. sp. PP1Y to a natural soil contaminated by heavy metals and PAHs, already endowed with a microbial community adapted to use PAHs in the presence of heavy metals, halved the time of natural self-purifying of the soil, thus suggesting the ability of N. sp. PP1Y to compete with the adapted endogenous microbial community.In conclusion Novosphingobium sp. PP1Y is a good candidate for the bioremediation of organic compound contaminated soils in the presence of heavy metals.
- Published
- 2012
11. Design and synthesis of new linezolid analogues
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DE ROSA, Margherita, Soriente, Annunziata, Francesca, Carbone, Notomista, E., Varcamonti, M., and Saturnino, Carmela
- Published
- 2012
12. Effective expression and purification of recombinant onconase, an antitumor protei
- Author
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NOTOMISTA E., CAFARO V., FUSIELLO R., BRACALE A., D'ALESSIO G., DI DONATO, ALBERTO, Notomista, E., Cafaro, V., Fusiello, R., Bracale, A., D'Alessio, G., and DI DONATO, Alberto
- Published
- 1999
13. The catalytic potential of bacterial multicomponent monooxygenases ToMO and PH for the synthesis of antioxidants tyrosol and hydroxytyrosol
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Izzo, Viviana, Notomista, E., Pezzella, A., D’Auria, M., Donadio, G., and Di Donato, A.
- Published
- 2009
14. Adaptation of Novosphingobium sp.PP1Y to grow on complex mixture of aromatic compounds dissolved in oil phases
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Notomista, E., Cafaro, V., Izzo, Viviana, Troncone, L., Smaldone, G., D’Urso, N., Garzillo, F., Varcamonti, F., and Di Donato, A.
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- 2009
15. Conformational analysis of putative regulatory subunit D of the toluene-o-xylene monooxygenase from Pseudomonas stutzeri OX1
- Author
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Sconamiglio, R., Notomista, E., Barbieri, Paola, Pucci, P., DAL PIAZ, F., Tramontano, A., and DI DONATO, A.
- Published
- 2001
16. Purification and characterization of the subunits B and D of the complex of toluene/o-xylene monooxygenase (Tomo) from Pseudomonas stutzeri OX1
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Sconamiglio, R., Notomista, E., Barbieri, Paola, Pucci, P., DAL PIAZ, F., DI DONATO, A., Comitato Scientifico del Convegno, Scognamiglio, Roberta, Notomista, Eugenio, Barbieri, P., Pucci, Pietro, Dal Piaz, F., and DI DONATO, Alberto
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Toluene-o-xylene monooxygenase ,subunits ,enzymatic complex ,Pseudomona - Published
- 1999
17. Expression and partial purification of the complex of toluene/o-xylene monooxygenase (Tomo) from Pseudomonas stutzeri OX1
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Notomista, E., Fusiello, S., Barbieri, Paola, DI DONATO, A., Comitato Scientifico del Convegno, Notomista, Eugenio, Fusiello, R., Barbieri, P., and DI DONATO, Alberto
- Subjects
Toluene-o-xylene monooxygenase ,Pseudomona ,protein expression - Published
- 1999
18. De Novo Sequencing and Assembly of the Whole Genome of Novosphingobium sp. Strain PP1Y
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D'Argenio, V., primary, Petrillo, M., additional, Cantiello, P., additional, Naso, B., additional, Cozzuto, L., additional, Notomista, E., additional, Paolella, G., additional, Di Donato, A., additional, and Salvatore, F., additional
- Published
- 2011
- Full Text
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19. Solution structure of the transmembrane proximal region of the hepatis C virus E1 glycoprotein
- Author
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Spadaccini, R., primary, D'Errico, G., additional, D'Alessio, V., additional, Notomista, E., additional, Bianchi, A., additional, Merola, M., additional, and Picone, D., additional
- Published
- 2010
- Full Text
- View/download PDF
20. Structural and dynamic studies of onconase mutants
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Merlino, A., primary, Mazzarella, L., additional, Carannante, A., additional, Di Fiore, A., additional, Di Donato, A., additional, Notomista, E., additional, and Sica, F., additional
- Published
- 2005
- Full Text
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21. X-ray structure of (C87S,des103-104) onconase
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Merlino, A., primary, Mazzarella, L., additional, Carannante, A., additional, Di Fiore, A., additional, Di Donato, A., additional, Notomista, E., additional, and Sica, F., additional
- Published
- 2005
- Full Text
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22. Enthalpic and entropic consequences of the removal of disulfide bridges in ribonuclease A
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Graziano, G., Catanzano, F., and Notomista, E.
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- 2000
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23. Effective expression and purification of recombinant onconase, an antitumor protein
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Notomista, E., Cafaro, V., Fusiello, R., Bracale, A., D'Alessio, G., and Donato, A. Di
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- 1999
- Full Text
- View/download PDF
24. Protein engineering of ribonucleases
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Cafaro, V., Bracale, A., Formiggini, F., Notomista, E., D'Alessio, G., and Donato, A. Di
- Published
- 1998
- Full Text
- View/download PDF
25. Mining for encrypted peptide antibiotics in the human proteome
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Marcelo D. T. Torres, Marcelo C. R. Melo, Cesar de la Fuente-Nunez, Orlando Crescenzi, Eugenio Notomista, Torres, M. D. T., Melo, M. C. R., Crescenzi, O., Notomista, E., and de la Fuente-Nunez, C.
- Subjects
Proteomics ,Proteome ,medicine.drug_class ,Antibiotics ,Antimicrobial peptides ,Biomedical Engineering ,Medicine (miscellaneous) ,Bioengineering ,Computational biology ,Biology ,medicine.disease_cause ,Mice ,Antibiotic resistance ,Immune system ,Anti-Bacterial Agent ,Human proteome project ,medicine ,Animal ,Microbial Sensitivity Test ,Pathogenic bacteria ,Antimicrobial ,biology.organism_classification ,Computer Science Applications ,Peptide ,Bacteria ,Biotechnology ,Human - Abstract
The emergence of drug-resistant bacteria calls for the discovery of new antibiotics. Yet, for decades, traditional discovery strategies have not yielded new classes of antimicrobial. Here, by mining the human proteome via an algorithm that relies on the sequence length, net charge, average hydrophobicity and other physicochemical properties of antimicrobial peptides, we report the identification of 2,603 encrypted peptide antibiotics that are encoded in proteins with biological function unrelated to the immune system. We show that the encrypted peptides kill pathogenic bacteria by targeting their membrane, modulate gut and skin commensals, do not readily select for bacterial resistance, and possess anti-infective activity in skin abscess and thigh infection mouse models. We also show, in vitro and in the two mouse models of infection, that encrypted antibiotic peptides from the same biogeographical area display synergistic antimicrobial activity. Our algorithmic strategy allows for the rapid mining of proteomic data and opens up new routes for the discovery of candidate antibiotics. The human proteome can be algorithmically mined to identify thousands of encrypted peptides, encoded in proteins with biological function unrelated to the immune system, that display antibacterial activity in vivo.
- Published
- 2022
26. Impact of a Single Point Mutation on the Antimicrobial and Fibrillogenic Properties of Cryptides from Human Apolipoprotein B
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Rocco Di Girolamo, Giovanni Smaldone, Angela Arciello, Mariano Rumolo, Pompea Del Vecchio, Angela Cesaro, Eugenio Notomista, Rosa Gaglione, Rosario Oliva, Emilia Pedone, Maria De Luca, Luigi Petraccone, Gaglione, R., Smaldone, G., Cesaro, A., Rumolo, M., De Luca, M., Di Girolamo, R., Petraccone, L., Del Vecchio, P., Oliva, R., Notomista, E., Pedone, E., and Arciello, A.
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Circular dichroism ,bioactive cryptides ,medicine.drug_class ,Antibiotics ,single point mutation ,Pharmaceutical Science ,Peptide ,Article ,in vitro fibrillogenesis ,Pharmacy and materia medica ,Drug Discovery ,medicine ,chemistry.chemical_classification ,Point mutation ,In vitro fibrillogenesi ,Antimicrobial ,Phenotype ,RS1-441 ,Biochemistry ,chemistry ,anti-biofilm activity ,Molecular Medicine ,Medicine ,Bioactive cryptide ,Antibacterial activity ,Wound healing - Abstract
Host defense peptides (HDPs) are gaining increasing interest, since they are endowed with multiple activities, are often effective on multidrug resistant bacteria and do not generally lead to the development of resistance phenotypes. Cryptic HDPs have been recently identified in human apolipoprotein B and found to be endowed with a broad-spectrum antimicrobial activity, with anti-biofilm, wound healing and immunomodulatory properties, and with the ability to synergistically act in combination with conventional antibiotics, while being not toxic for eukaryotic cells. Here, a multidisciplinary approach was used, including time killing curves, differential scanning calorimetry, circular dichroism, ThT binding assays, and transmission electron microscopy analyses. The effects of a single point mutation (Pro → Ala in position 7) on the biological properties of ApoB-derived peptide r(P)ApoBLPro have been evaluated. Although the two versions of the peptide share similar antimicrobial and anti-biofilm properties, only r(P)ApoBLAla peptide was found to exert bactericidal effects. Interestingly, antimicrobial activity of both peptide versions appears to be dependent from their interaction with specific components of bacterial surfaces, such as LPS or LTA, which induce peptides to form β-sheet-rich amyloid-like structures. Altogether, obtained data indicate a correlation between ApoB-derived peptides self-assembling state and their antibacterial activity.
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- 2021
27. Encapsulating properties of sulfobutylether-β-cyclodextrin toward a thrombin-derived antimicrobial peptide
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Serena Cozzolino, Roland Winter, Pompea Del Vecchio, Filomena Battista, Eugenio Notomista, Rosario Oliva, Luigi Petraccone, Oliva, R., Battista, F., Cozzolino, S., Notomista, E., Winter, R., Del Vecchio, P., and Petraccone, L.
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chemistry.chemical_classification ,Circular dichroism ,Chemistry ,Antimicrobial peptides ,Peptide ,02 engineering and technology ,021001 nanoscience & nanotechnology ,Condensed Matter Physics ,Antimicrobial ,01 natural sciences ,Combinatorial chemistry ,Fluorescence ,DSC ,010406 physical chemistry ,0104 chemical sciences ,Differential scanning calorimetry ,Membrane ,Liposomes ,Isothermal titration calorimetry ,Cyclodextrin ,Physical and Theoretical Chemistry ,Antimicrobial peptide ,0210 nano-technology ,Lipid bilayer - Abstract
The pharmacological application of antimicrobial peptides (AMPs) is seriously limited as they are not chemically and physically stable. Their encapsulation could represent a way to protect AMPs improving their pharmacological properties. In this study, the complex between the sulfobutylether-b-cyclodextrin (SBE-b-CD) and the antimicrobial peptide (P)GKY20 and its effect on a lipid bilayer have been characterized by a combination of calorimetric (ITC, DSC) and spectroscopic (fluorescence, Circular Dichroism) techniques. The results obtained indicate that the (P)GKY20 form a 1:1 inclusion complex with the anionic SBE-b-CD. Our ITC experiments revealed that the interaction process is entropically driven and that the enthalpy change only slightly contributes to the free energy change. Finally, differential scanning calorimetry data revealed that the peptide, even in the presence of SBE-b-CD, is still able to perturb the bacterial model membrane composed of DPPC/DPPG (8/2 mol mol-1). This work demonstrated that, in principle, SBE-b-CD could be used as efficient encapsulating agent for the (P)GKY20 or other antimicrobial peptides, rendering possible their pharmacological applications.
- Published
- 2019
28. Environment‐sensitive fluorescent labelling of peptides by luciferin analogues
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Marialuisa Siepi, Rosario Oliva, Antonio Masino, Rosa Gaglione, Angela Arciello, Rosita Russo, Antimo Di Maro, Anna Zanfardino, Mario Varcamonti, Luigi Petraccone, Pompea Del Vecchio, Marcello Merola, Elio Pizzo, Eugenio Notomista, Valeria Cafaro, Siepi, M., Oliva, R., Masino, A., Gaglione, R., Arciello, A., Russo, R., Di Maro, A., Zanfardino, A., Varcamonti, M., Petraccone, L., Del Vecchio, P., Merola, M., Pizzo, E., Notomista, E., and Cafaro, V.
- Subjects
Membrane‐binding peptide ,QH301-705.5 ,Fluorescent Dye ,Antitumor peptide ,Catalysis ,Article ,Inorganic Chemistry ,Luciferin ,Peptide labeling ,membrane-binding peptide ,Physical and Theoretical Chemistry ,Biology (General) ,Molecular Biology ,fluorescent peptide ,environment-sensitive fluorophore ,peptide labeling ,luciferin ,antimicrobial peptide ,antitumor peptide ,RGD peptide ,QD1-999 ,Fluorescent peptide ,Spectroscopy ,Fluorescent Dyes ,Organic Chemistry ,General Medicine ,Hydrogen-Ion Concentration ,Computer Science Applications ,Chemistry ,Luciferins ,Environment‐sensitive fluorophore ,Peptides ,Antimicrobial peptide - Abstract
Environment-sensitive fluorophores are very valuable tools in the study of molecular and cellular processes. When used to label proteins and peptides, they allow for the monitoring of even small variations in the local microenvironment, thus acting as reporters of conformational variations and binding events. Luciferin and aminoluciferin, well known substrates of firefly luciferase, are environment-sensitive fluorophores with unusual and still-unexploited properties. Both fluorophores show strong solvatochromism. Moreover, luciferin fluorescence is influenced by pH and water abundance. These features allow to detect local variations of pH, solvent polarity and local water concentration, even when they occur simultaneously, by analyzing excitation and emission spectra. Here, we describe the characterization of (amino)luciferin-labeled derivatives of four bioactive peptides: the antimicrobial peptides GKY20 and ApoBL, the antitumor peptide p53pAnt and the integrin-binding peptide RGD. The two probes allowed for the study of the interaction of the peptides with model membranes, SDS micelles, lipopolysaccharide micelles and Escherichia coli cells. Kd values and binding stoichiometries for lipopolysaccharide were also determined. Aminoluciferin also proved to be very well-suited to confocal laser scanning microscopy. Overall, the characterization of the labeled peptides demonstrates that luciferin and aminoluciferin are previously neglected environment-sensitive labels with widespread potential applications in the study of proteins and peptides.
- Published
- 2021
29. Antimicrobial D-amino acid oxidase-derived peptides specify gut microbiota
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Murtas, Giulia, Sacchi, Silvia, Tedeschi, Gabriella, Maffioli, Elisa, Notomista, Eugenio, Cafaro, Valeria, Abbondi, Monica, Mothet, Jean-Pierre, Pollegioni, Loredano, University of Insubria, Varese, University of Milan, University of Naples Federico II, Laboratoire Lumière, Matière et Interfaces (LuMIn), CentraleSupélec-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS)-Ecole Normale Supérieure Paris-Saclay (ENS Paris Saclay), Murtas, G., Sacchi, S., Tedeschi, G., Maffioli, E., Notomista, E., Cafaro, V., Abbondi, M., Mothet, J. -P., and Pollegioni, L.
- Subjects
Male ,D-Amino acids ,Protein Conformation ,d-Amino acid ,D-amino acid oxidase ,Sequence Homology ,Gut flora ,Flavoprotein ,Microbiota selection ,Novel function ,Mice ,0302 clinical medicine ,Intestine, Small ,Amino Acids ,chemistry.chemical_classification ,0303 health sciences ,Oxidase test ,biology ,Anti-Bacterial Agents ,Amino acid ,Amino Acid ,Pore Forming Cytotoxic Protein ,Biochemistry ,Molecular Medicine ,Original Article ,Female ,Human ,Pore Forming Cytotoxic Proteins ,D-Amino-Acid Oxidase ,Proteases ,Antimicrobial peptides ,Gram-Positive Bacteria ,03 medical and health sciences ,Cellular and Molecular Neuroscience ,[SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN] ,Anti-Bacterial Agent ,Gram-Negative Bacteria ,Animals ,Humans ,Secretion ,Amino Acid Sequence ,Rats, Wistar ,Molecular Biology ,030304 developmental biology ,Pharmacology ,Animal ,Cell Biology ,biology.organism_classification ,Rats ,Gastrointestinal Microbiome ,Mice, Inbred C57BL ,chemistry ,Rat ,030217 neurology & neurosurgery ,Bacteria - Abstract
The flavoenzyme d-amino acid oxidase (DAAO) is deputed to the degradation of d-enantiomers of amino acids. DAAO plays various relevant physiological roles in different organisms and tissues. Thus, it has been recently suggested that the goblet cells of the mucosal epithelia secrete into the lumen of intestine, a processed and active form of DAAO that uses the intestinal d-amino acids to generate hydrogen peroxide (H2O2), an immune messenger that helps fighting gut pathogens, and by doing so controls the homeostasis of gut microbiota. Here, we show that the DAAO form lacking the 1–16 amino acid residues (the putative secretion signal) is unstable and inactive, and that DAAO is present in the epithelial layer and the mucosa of mouse gut, where it is largely proteolyzed. In silico predicted DAAO-derived antimicrobial peptides show activity against various Gram-positive and Gram-negative bacteria but not on Lactobacilli species, which represent the commensal microbiota. Peptidomic analysis reveals the presence of such peptides in the mucosal fraction. Collectively, we identify a novel mechanism for gut microbiota selection implying DAAO-derived antimicrobial peptides which are generated by intestinal proteases and that are secreted in the gut lumen. In conclusion, we herein report an additional, ancillary role for mammalian DAAO, unrelated to its enzymatic activity.
- Published
- 2021
30. Human Cryptic Host Defence Peptide GVF27 Exhibits Anti-Infective Properties against Biofilm Forming Members of the Burkholderia cepacia Complex
- Author
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Bosso, Andrea, Gaglione, Rosa, Di Girolamo, Rocco, Veldhuizen, Edwin J A, García-Vello, Pilar, Fusco, Salvatore, Cafaro, Valeria, Monticelli, Maria, Culurciello, Rosanna, Notomista, Eugenio, Arciello, Angela, Pizzo, Elio, Immunologie, dI&I I&I-3, Bosso, A., Gaglione, R., Di Girolamo, R., Veldhuizen, E. J. A., Garcia-Vello, P., Fusco, S., Cafaro, V., Monticelli, M., Culurciello, R., Notomista, E., Arciello, A., Pizzo, E., Immunologie, and dI&I I&I-3
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Immunomodulatory activitie ,Anti-biofilm agents ,Pharmaceutical Science ,Host defence peptides ,Immunomodulatory activities ,Anti-biofilm agent ,Host defence peptide ,Synergy ,host defence peptides ,antimicrobial peptides ,anti-biofilm agents ,LPS neutralisation ,agglutinating activity ,immunomodulatory activities ,synergy ,Drug Discovery ,Antimicrobial peptides ,Molecular Medicine ,Antimicrobial peptide ,Agglutinating activity - Abstract
Therapeutic solutions to counter Burkholderia cepacia complex (Bcc) bacteria are challenging due to their intrinsically high level of antibiotic resistance. Bcc organisms display a variety of potential virulence factors, have a distinct lipopolysaccharide naturally implicated in antimicrobial resistance. and are able to form biofilms, which may further protect them from both host defence peptides (HDPs) and antibiotics. Here, we report the promising anti-biofilm and immunomodulatory activities of human HDP GVF27 on two of the most clinically relevant Bcc members, Burkholderia multivorans and Burkholderia cenocepacia. The effects of synthetic and labelled GVF27 were tested on B. cenocepacia and B. multivorans biofilms, at three different stages of formation, by confocal laser scanning microscopy (CLSM). Assays on bacterial cultures and on human monocytes challenged with B. cenocepacia LPS were also performed. GVF27 exerts, at different stages of formation, anti-biofilm effects towards both Bcc strains, a significant propensity to function in combination with ciprofloxacin, a relevant affinity for LPSs isolated from B. cenocepacia as well as a good propensity to mitigate the release of pro-inflammatory cytokines in human cells pre-treated with the same endotoxin. Overall, all these findings contribute to the elucidation of the main features that a good therapeutic agent directed against these extremely leathery biofilm-forming bacteria should possess.
- Published
- 2022
31. Enzymes as a Reservoir of Host Defence Peptides
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Alberto Di Donato, Elio Pizzo, Antimo Di Maro, Valeria Cafaro, Andrea Bosso, Eugenio Notomista, Bosso, Andrea, Di Maro, Antimo, Cafaro, Valeria, Di Donato, Alberto, Notomista, Eugenio, Pizzo, Elio, Bosso, A., Di Maro, A., Cafaro, V., Di Donato, A., Notomista, E., and Pizzo, E.
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Hydrolases ,Protein Conformation ,Proteolysis ,Hydrolase ,proteolysi ,Computational biology ,Biology ,immunomodulation ,03 medical and health sciences ,Enzyme activator ,Protein structure ,medicine ,Humans ,Structure–activity relationship ,030304 developmental biology ,chemistry.chemical_classification ,0303 health sciences ,Antimicrobial Cationic Peptide ,Innate immune system ,medicine.diagnostic_test ,030306 microbiology ,Drug discovery ,Hydrolysis ,structure-activity relationship ,mimetic peptide ,Biocatalysi ,General Medicine ,Host defence ,Hydrolysi ,Immunity, Innate ,Enzyme Activation ,Enzyme ,chemistry ,Oxidoreductase ,Biocatalysis ,Oxidoreductases ,Antimicrobial Cationic Peptides ,Human - Abstract
Host defence peptides (HDPs) are powerful modulators of cellular responses to various types of insults caused by pathogen agents. To date, a wide range of HDPs, from species of different kingdoms including bacteria, plant and animal with extreme diversity in structure and biological activity, have been described. Apart from a limited number of peptides ribosomally synthesized, a large number of promising and multifunctional HDPs have been identified within protein precursors, with properties not necessarily related to innate immunity, consolidating the fascinating hypothesis that proteins have a second or even multiple biological mission in the form of one or more bio-active peptides. Among these precursors, enzymes constitute certainly an interesting group, because most of them are mainly globular and characterized by a fine specific internal structure closely related to their catalytic properties and also because they are yet little considered as potential HDP releasing proteins. In this regard, the main aim of the present review is to describe a panel of HDPs, identified in all canonical classes of enzymes, and to provide a detailed description on hydrolases and their corresponding HDPs, as there seems to exist a striking link between these structurally sophisticated catalysts and their high content in cationic and amphipathic cryptic peptides.
- Published
- 2020
32. Similarities and differences for membranotropic action of three unnatural antimicrobial peptides
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Rosario Oliva, Luigi Petraccone, Pompea Del Vecchio, Angelina Lombardi, Marco Chino, Eugenio Notomista, Flavia Nastri, Oliva, R., Chino, M., Lombardi, A., Nastri, F., Notomista, E., Petraccone, L., and Del Vecchio, P.
- Subjects
Pore Forming Cytotoxic Proteins ,Circular dichroism ,Gram-negative bacteria ,antimicrobial peptide ,Stereochemistry ,Gram-positive bacteria ,Antimicrobial peptides ,Peptide ,Microbial Sensitivity Tests ,010402 general chemistry ,01 natural sciences ,Biochemistry ,Structural Biology ,Drug Discovery ,Molecular Biology ,Pharmacology ,chemistry.chemical_classification ,biology ,Calorimetry, Differential Scanning ,010405 organic chemistry ,Chemistry ,Circular Dichroism ,Organic Chemistry ,General Medicine ,biology.organism_classification ,0104 chemical sciences ,Amino acid ,Anti-Bacterial Agents ,lipoteichoic acid ,Spectrometry, Fluorescence ,liposome ,Molecular Medicine ,Lipoteichoic acid ,fluorescence ,differential scanning calorimetry ,Bacteria ,unnatural amino acids - Abstract
Previously, we described the design and synthesis of three nine-residue AMPs, P9Nal(SS), P9Trp(SS), and P9Nal(SR), showing high stability in serum and broad spectrum antimicrobial activity. The peptides P9Trp(SS) and P9Nal(SR) differ from P9Nal(SS) for the replacement of the two 2Nal residues with Trp residues and for the replacement of the two Cys (StBu) with Cys (tBu) residues, respectively. These changes led to peptides with a lower hydrophobicity respect to the P9Nal(SS). Interestingly, the three peptides have very similar activity against Gram-negative bacteria. Instead, they exhibit a significant difference towards Gram-positive bacteria, being P9Nal(SS) the most active. In order to evaluate the impact of amino acids substitution on membranotropic activity and rationalize the observed effects in vivo, here, we report the detailed biophysical characterization of the interaction between P9Nal(SR) and P9Trp(SS) and liposomes by combining differential scanning calorimetry, circular dichroism, and fluorescence spectroscopy. The comparison with the results for the previously characterized P9Nal(SS) peptide reveals similarities and differences on the interaction process and perturbation activities. It was found that the three peptides can penetrate at different extent inside the bilayer upon changing their conformation and inducing lipid domains formation, revealing that the formation of lipid domains is fundamental for the activity against Gram-negative bacteria. On the contrary, the dissimilar activity against Gram-positive bacteria well correlate with the different affinity of peptides for the lipoteichoic acid, a component selectively present in the cell wall of Gram-positive bacteria.
- Published
- 2020
33. Characterisation of EFV12 a bio-active small peptide produced by the human intestinal isolate Lactobacillus gasseri SF1109
- Author
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Andrea Carpentieri, Gerardino D'Errico, Anna Zanfardino, Giuseppe Vitiello, B Di Luccia, Eliodoro Pizzo, Mario Varcamonti, Alessandro Pezzella, M. Di Napoli, Eugenio Notomista, Di Napoli, M., Di Luccia, B., Vitiello, G., D'Errico, G., Carpentieri, A., Pezzella, A., Pizzo, E., Notomista, E., Varcamonti, M., and Zanfardino, A.
- Subjects
0301 basic medicine ,Microbiology (medical) ,p38 mitogen-activated protein kinases ,Lactobacillus gasseri ,01 natural sciences ,Microbiology ,law.invention ,03 medical and health sciences ,Probiotic ,law ,medicine ,Extracellular ,Receptor ,biology ,010405 organic chemistry ,Kinase ,Chemistry ,Antimicrobial ,biology.organism_classification ,0104 chemical sciences ,030104 developmental biology ,Mechanism of action ,Biochemistry ,medicine.symptom ,LPS binding ,Antimicrobial peptide - Abstract
EFV12 is a small bioactive peptide produced by Lactobacillus gasseri SF1109, a human intestinal isolate with probiotic features. In this study, EFV12 antimicrobial and anti-inflammatory properties are characterised. In particular, we propose a possible mechanism of action for EFV12 involving bacterial membranes targeting. Moreover, we show that this small peptide is able to bind lipopolysaccharides (LPS) and to counteract its inflammatory insult preventing LPS action on Toll-like receptor 4, thus interfering with extracellular signal-regulated kinase, p38 and Jun N-terminal kinase, mitogen-activated protein kinases signalling pathways. Altogether these observations suggest that the bioactive peptide EFV12 is a good candidate to promote L. gasseri induced gut homeostasis and counteracting intestinal pathogens.
- Published
- 2020
34. Antimicrobial peptide Temporin-L complexed with anionic cyclodextrins results in a potent and safe agent against sessile bacteria
- Author
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Ettore Novellino, Elio Pizzo, Francesco Merlino, Alfonso Carotenuto, Agnese Miro, Milo Malanga, Eugenio Notomista, Andrea Bosso, Paolo Grieco, Federica De Lise, Valeria Cafaro, Rosa Gaglione, Francesca Ungaro, Fabiana Quaglia, Diego Brancaccio, Brancaccio, D., Pizzo, E., Cafaro, V., Notomista, E., De Lise, F., Bosso, A., Gaglione, R., Merlino, F., Novellino, E., Ungaro, F., Grieco, P., Malanga, M., Quaglia, F., Miro, A., and Carotenuto, A.
- Subjects
Models, Molecular ,medicine.drug_class ,Antibiotic resistance ,Alginates ,Cell Survival ,Antimicrobial peptides ,Antibiotics ,Pharmaceutical Science ,Peptide ,02 engineering and technology ,Antibiofilm agent ,Bacterial Physiological Phenomena ,030226 pharmacology & pharmacy ,Cell Line ,03 medical and health sciences ,0302 clinical medicine ,Anti-Infective Agents ,medicine ,Humans ,chemistry.chemical_classification ,Cyclodextrins ,biology ,Bacteria ,Chemistry ,021001 nanoscience & nanotechnology ,Antimicrobial ,biology.organism_classification ,Combinatorial chemistry ,Temporin ,Yeast ,Solubility ,Biofilms ,0210 nano-technology ,Antimicrobial peptide ,Antimicrobial Cationic Peptides - Abstract
Concern over antibiotic resistance is growing, and new classes of antibiotics, particularly against Gram-negative bacteria, are needed. Antimicrobial peptides (AMPs) have been proposed as a new class of clinically useful antimicrobials. Special attention has been devoted to frog-skin temporins. In particular, temporin L (TL) is strongly active against Gram-positive, Gram-negative bacteria and yeast strains. With the aim of overcoming some of the main drawbacks preventing the widespread clinical use of this peptide, i.e. toxicity and unfavorable pharmacokinetics profile, we designed new formulations combining TL with different types of cyclodextrins (CDs). TL was associated to a panel of neutral or negatively charged, monomeric and polymeric CDs. The impact of CDs association on TL solubility, as well as the transport through bacterial alginates was assessed. The biocompatibility on human cells together with the antimicrobial and antibiofilm properties of TL/CD systems was explored.
- Published
- 2019
35. Human cytomegalovirus pUL10 interacts with leukocytes and impairs TCR‐mediated T‐cell activation
- Author
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Diego Piccioli, Luisanna Zedda, Eugenio Notomista, Francesca Schiavetti, Mary Schaefer, Mirko Cortese, Andrea Carfi, Michela Gentile, Stefano Calo, Elena Cattaneo, Luca Bruno, Marcello Merola, Giuseppina Monda, Domenico Maione, Yasushi Uematsu, Bruno, Luca, Cortese, Mirko, Monda, Giuseppina, Gentile, Michela, Caló, Stefano, Schiavetti, Francesca, Zedda, Luisanna, Cattaneo, Elena, Piccioli, Diego, Schaefer, Mary, Notomista, Eugenio, Maione, Domenico, Carfí, Andrea, Merola, Marcello, Uematsu, Yasushi, Bruno, L., Cortese, M., Monda, G., Gentile, M., Calo, S., Schiavetti, F., Zedda, L., Cattaneo, E., Piccioli, D., Schaefer, M., Notomista, E., Maione, D., Carfi, A., Merola, M., and Uematsu, Y.
- Subjects
CD4-Positive T-Lymphocytes ,0301 basic medicine ,Human cytomegalovirus ,Allergy ,Glycosylation ,T-Lymphocytes ,T cell ,Immunology ,Receptors, Antigen, T-Cell ,chemical and pharmacologic phenomena ,CD8-Positive T-Lymphocytes ,Biology ,Lymphocyte Activation ,Cell Line ,03 medical and health sciences ,Immune system ,Immunity ,medicine ,Humans ,Immunology and Allergy ,Amino Acid Sequence ,Cell Proliferation ,Cell Membrane ,T-cell receptor ,Membrane Proteins ,virus diseases ,hemic and immune systems ,Cell Biology ,medicine.disease ,Recombinant Proteins ,HEK293 Cells ,030104 developmental biology ,medicine.anatomical_structure ,Immunology, virology, HCMV, T cells, viral proteins, pUL10 ,Cytokines ,Capsid Proteins ,Tumor immunology ,Signal Transduction - Abstract
Human cytomegalovirus (HCMV) is known to exert suppressive effects on the host immune system through expression of various viral genes, thus directly and indirectly affecting antiviral immunity of the infected individuals. We report here that HCMV UL10 encodes a protein (pUL10) with immunosuppressive properties. UL10 has been classified as a member of the HCMV RL11 gene family. Although pUL10 is known to be dispensable for viral replication in cultured cells, its amino-acid sequence is well conserved among different HCMV isolates, suggesting that the protein has a crucial role in viral survival in the host environment. We show that pUL10 is cleaved from the cell surface of fibroblasts as well as epithelial cells and interacts with a cellular receptor ubiquitously expressed on the surface of human leukocytes, demonstrated by ex vivo cell-based assays and flow cytometric analyses on both lymphoid cell lines and primary blood cells. Furthermore, preincubation of peripheral blood mononuclear cells with purified pUL10 ectodomain results in significantly impaired proliferation and substantially reduced pro-inflammatory cytokine production, in particular in CD4 + T cells upon in vitro T-cell stimulation. The inhibitory effect of pUL10 is also observed on antigen receptor-mediated intracellular tyrosine phosphorylation in a T-cell line. Based on these observations, we suggest that pUL10 is a newly identified immunomodulatory protein encoded by HCMV. Further elucidation of interactions between pUL10 and the host immune system during HCMV may contribute to finding ways towards new therapies for HCMV infection.
- Published
- 2016
36. Human Cryptic Host Defence Peptides: identification and analysis of their antimicrobial, anti-biofilm and immunomodulatory properties
- Author
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NOTOMISTA, EUGENIO, BOSSO, ANDREA, PANE, KATIA, ZANFARDINO, ANNA, VARCAMONTI, MARIO, ARCIELLO, ANGELA, CATANIA, MARIA ROSARIA, CAFARO, VALERIA, DI DONATO, ALBERTO, PIZZO, ELIODORO, Emilia, Pedone, Edwin, Veldhuizen, César De La Fuente núñez, Notomista E., Bosso A., Pane K, Zanfardino A., Varcamonti M., Pedone E., Arciello A., Veldhuizen E., de la Fuente-Núñez C., Catania MR., Cafaro V., Di Donato A., Pizzo E., Notomista, Eugenio, Bosso, Andrea, Pane, Katia, Zanfardino, Anna, Varcamonti, Mario, Emilia, Pedone, Arciello, Angela, Edwin, Veldhuizen, César De La Fuente, Núñez, Catania, MARIA ROSARIA, Cafaro, Valeria, DI DONATO, Alberto, and Pizzo, Eliodoro
- Subjects
Cryptic Host Defence Peptides, biofilm, CAMP - Published
- 2017
37. RHA-P: a novel bacterial α-L-rhamnosidase of biotechnological relevance from Novosphingobium sp. PP1Y
- Author
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MENSITIERI, FRANCESCA, DE LISE, FEDERICA, CAFARO, VALERIA, Maria, Lumacone, STRAZZULLI, ANDREA, NOTOMISTA, EUGENIO, MORACCI, Marco, DI DONATO, ALBERTO, Viviana, Izzo, Mensitieri F., De Lise F., Cafaro V., Lumacone M., Strazzulli A., Notomista E., Moracci M., Di Donato A., Izzo V., Mensitieri, Francesca, DE LISE, Federica, Cafaro, Valeria, Maria, Lumacone, Strazzulli, Andrea, Notomista, Eugenio, Moracci, Marco, DI DONATO, Alberto, and Viviana, Izzo
- Abstract
α-L-Rhamnosidases (α-RHAs, EC 3.2.1.40) are glycosyl hydrolases (GHs) hydrolyzing terminal α-L-rhamnose residues from different substrates such as heteropolysaccharides, glycosylated proteins and natural flavonoids. Although the possibility to hydrolyze rhamnose from natural flavonoids has boosted the use of these enzymes in several biotechnological applications over the past decades, to date only few bacterial rhamnosidases have been fully characterized and only one crystal structure of a rhamnosidase of the GH106 family has been described. In our previous work, an α-L-rhamnosidase belonging to this family, named RHA-P, was isolated from the marine microorganism Novosphingobium sp. PP1Y. The initial biochemical characterization highlighted the biotechno- logical potential of RHA-P for bioconversion applications. In this work, further functional and structural char- acterization of the enzyme is provided. The recombinant protein was obtained fused to a C-terminal His-tag and, starting from the periplasmic fractions of induced recombinant cells of E. coli strain BL21(DE3), was purified through a single step purification protocol. Homology modeling of RHA-P in combination with a site directed mutagenesis analysis confirmed the function of residues D503, E506, E644, likely located at the catalytic site of RHA-P. In addition, a kinetic characterization of the enzyme on natural flavonoids such as naringin, rutin, hesperidin and quercitrin was performed. RHA-P showed activity on all flavonoids tested, with a catalytic ef- ficiency comparable or even higher than other bacterial α-RHAs described in literature. The results confirm that RHA-P is able to hydrolyze both α-1,2 and α-1,6 glycosidic linkages, and suggest that the enzyme may locate different polyphenolic aromatic moities in the active site.
- Published
- 2017
38. Cryptic antimicrobial peptides, a still unexploited source of bioactive peptides
- Author
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PANE, KATIA, CAFARO, VALERIA, AVITABILE, ANGELA, DE GREGORIO, ELIANA, VOLLARO, ADRIANA, CATANIA, MARIA ROSARIA, CRESCENZI, ORLANDO, ARCIELLO, ANGELA, ZANFARDINO, ANNA, VARCAMONTI, MARIO, CONTURSI, PATRIZIA, DI DONATO, ALBERTO, PIZZO, ELIODORO, NOTOMISTA, EUGENIO, Antimo Di Maro, Pane K., Cafaro V., Avitabile A., De Gregorio E., Vollaro A., Catania MR., Crescenzi O., Arciello A., Di Maro A., Zanfardino A., Varcamonti M., Contursi P., Di Donato A., Pizzo E., Notomista E., Pane, Katia, Cafaro, Valeria, Avitabile, Angela, DE GREGORIO, Eliana, Vollaro, Adriana, Catania, MARIA ROSARIA, Crescenzi, Orlando, Arciello, Angela, Antimo Di, Maro, Zanfardino, Anna, Varcamonti, Mario, Contursi, Patrizia, DI DONATO, Alberto, Pizzo, Eliodoro, and Notomista, Eugenio
- Subjects
Cryptic antimicrobial peptides, CAMP, antimicrobial - Published
- 2017
39. Novel human antimicrobial peptides from ApoB are endowed with promising anti-inflammatory properties
- Author
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6. R. Gaglione, K. Pane, E. Dell’Olmo, A. Bosso, M. Chino F. Itri, V. Cafaro, E. Pizzo, E. J. A. Veldhuizen, A. Lombardi, R. Piccoli, E. Notomista, A. Arciello, Italian Federation of Life Sciences, Gaglione, 6. R., Pane, K., Dell’Olmo, E., Bosso, A., Itri, M. Chino F., Cafaro, V., Pizzo, E., Veldhuizen, E. J. A., Lombardi, A., Piccoli, R., Notomista, E., and Arciello, A.
- Published
- 2016
40. A new active antimicrobial peptide from PD-L4, a type 1 ribosome inactivating protein of Phytolacca dioica L.: A new function of RIPs for plant defence?
- Author
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Eliodoro Pizzo, Antonella M.A. Di Giuseppe, Andrea Bosso, Mario Varcamonti, Anna Zanfardino, Eugenio Notomista, Nicola Landi, Antimo Di Maro, Sara Ragucci, Pizzo, Eliodoro, Zanfardino, Anna, Di Giuseppe, Antonella M. A, Bosso, Andrea, Landi, Nicola, Ragucci, Sara, Varcamonti, Mario, Notomista, Eugenio, Di Maro, Antimo, Pizzo, E., Zanfardino, A., Di Giuseppe, A. M. A., Bosso, A., Landi, N., Ragucci, S., Varcamonti, M., Notomista, E., and DI MARO, Antimo
- Subjects
Protein Conformation ,Antimicrobial peptides ,Molecular Sequence Data ,Biophysics ,Ribosome Inactivating Proteins ,Phytolacca dioica ,Peptide ,Biochemistry ,Protein structure ,Anti-Infective Agents ,Structural Biology ,Cationic antimicrobial peptide ,Genetics ,Humans ,Amino Acid Sequence ,Molecular Biology ,Peptide sequence ,Phytolacca ,Plant Proteins ,chemistry.chemical_classification ,biology ,Bacteria ,Ribosome-inactivating protein ,Cell Biology ,Ribosome inactivating protein ,biology.organism_classification ,Antimicrobial ,Peptide Fragments ,Plant Leaves ,chemistry ,Caco-2 Cells ,Antimicrobial peptide - Abstract
We investigated the antimicrobial activity of PD-L4, a type 1 RIP from Phytolacca dioica. We found that this protein is active on different bacterial strains both in a native and denatured/alkylated form and that this biological activity is related to a cryptic peptide, named PDL440-65, identified by chemical fragmentation. This peptide showed the same antimicrobial activity of full-length protein and possessed, similarly to several antimicrobial peptides, an immunomodulatory effect on human cells. It assumes an alpha-helical conformation when interact with mimic membrane agents as TFE and likely bacterial membranes are a target of this peptide. To date PDL440-65 is the first antimicrobial peptide identified in a type 1 RIP. We investigated the antimicrobial activity of PD-L4, a type 1 RIP from Phytolacca dioica. We found that this protein is active on different bacterial strains both in a native and denatured/alkylated form and that this biological activity is related to a cryptic peptide, named PDL4(40-65), identified by chemical fragmentation. This peptide showed the same antimicrobial activity of full-length protein and possessed, similarly to several antimicrobial peptides, an immunomodulatory effect on human cells. It assumes an alpha-helical conformation when interact with mimic membrane agents as TFE and likely bacterial membranes are a target of this peptide. To date PDL4(40-65) is the first antimicrobial peptide identified in a type 1 RIP. (C) 2015 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
- Published
- 2015
41. Isolation of an Escherichia coli K4 kfoC mutant over-producing capsular chondroitin
- Author
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Mario De Rosa, Eugenio Notomista, Donatella Cimini, Chiara Schiraldi, Mario Varcamonti, Odile Francesca Restaino, Maurilio De Felice, Anna Zanfardino, Zanfardino, A., Restaino, Odile Francesca, Notomista, E., Cimini, Donatella, Schiraldi, Chiara, DE ROSA, Mario, De Felice, M., Varcamonti, M., Zanfardino, Anna, Restaino, Of, Notomista, Eugenio, Cimini, D, Schiraldi, C, De Rosa, M, DE FELICE, Maurilio, and Varcamonti, Mario
- Subjects
Bacterial capsule ,Mutant ,Mutation, Missense ,lcsh:QR1-502 ,Mutagenesis (molecular biology technique) ,Bioengineering ,medicine.disease_cause ,Applied Microbiology and Biotechnology ,lcsh:Microbiology ,Microbiology ,chemistry.chemical_compound ,medicine ,Escherichia coli ,Chondroitin ,Bacterial Capsules ,chemistry.chemical_classification ,biology ,Organisms, Genetically Modified ,Escherichia coli Proteins ,Research ,Anti-Inflammatory Agents, Non-Steroidal ,Chondroitin Sulfates ,Wild type ,Gene Expression Regulation, Bacterial ,biology.organism_classification ,carbohydrates (lipids) ,Enzyme ,chemistry ,Biochemistry ,Hexosyltransferases ,Bacteria ,Biotechnology - Abstract
Background Chondroitin sulphate is a complex polysaccharide having important structural and protective functions in animal tissues. Extracted from animals, this compound is used as a human anti-inflammatory drug. Among bacteria, Escherichia coli K4 produces a capsule containing a non-sulphate chondroitin and its development may provide an efficient and cheap fermentative production of the polysaccharide. Results A random N-methyl-N'-nitro-N-nitrosoguanidine mutagenesis was performed on E. coli K4 to isolate mutants showing an increased production of chondroitin. Several mutants were isolated, one of which, here named VZ15, produced about 80% more chondroitin than the wild type E. coli. We found that the mutant has a missense mutation in the codon 313 of kfoC, the gene encoding chondroitin polymerase (K4CP), with a change from arginine to glutamine. A docking analysis to explain the increased productivity of the K4CP enzyme is presented. Conclusion The enhanced chondroitin production by the E. coli K4 mutant reported here shows the validity of the strain improvement strategy for more cost-friendly fermentative processes in the production of this pharmaceutically important but so-far expensive polysaccharide.
- Published
- 2010
42. EXPRESSION AND PURIFICATION OF THE RECOMBINANT SUBUNITS OF TOLUENE/O-XYLENE MONOOXYGENASE AND RECONSTITUTION OF THE ACTIVE COMPLEX
- Author
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CAFARO, VALERIA, NOTOMISTA, EUGENIO, AMORESANO, ANGELA, PUCCI, PIETRO, DI DONATO, ALBERTO, Scognamiglio, R, Viggiani, A, Izzo, V, Passaro, I, DAL PIAZ, F, Casbarra, A, Cafaro, V., Scognamiglio, R., Viggiani, A., Izzo, V., Passaro, I., Notomista, Eugenio, DEL PIAZ, F., Amoresano, A., Casbarra, A., Pucci, P., DI DONATO, Alberto, Notomista, E., DAL PIAZ, F., Cafaro, Valeria, DI DONATO, AND A., Scognamiglio, R, Viggiani, A, Izzo, V, Passaro, I, DAL PIAZ, F, Amoresano, Angela, Casbarra, A, and Pucci, Pietro
- Subjects
bioremediation ,monooxygenase ,recombinant ,electron transfer ,protein expression - Abstract
This paper describes the cloning of the genes coding for each component of the complex of toluene/o-xylene monooxygenase from Pseudomonas stutzeri OX1, their expression, purification and characterization. Moreover, the reconstitution of the active complex from the recombinant subunits has been obtained, and the functional role of each component in the electron transfer from the electron donor to molecular oxygen has been determined. The coexpression of subunits B, E and A leads to the formation of a subcomplex, named H, with a quaternary structure (BEA)2, endowed with hydroxylase activity. Tomo F component is an NADH oxidoreductase. The purified enzyme contains about 1 mol of FAD, 2 mol of iron, and 2 mol of acid labile sulfide per mol of protein, as expected for the presence of one [2Fe-2S] cluster, and exhibits a typical flavodoxin absorption spectrum. Interestingly, the sequence of the protein does not correspond to that previously predicted on the basis of DNA sequence. We have shown that this depends on minor errors in the gene sequence that we have corrected. C component is a Rieske-type ferredoxin, whose iron and acid labile sulfide content is in agreement with the presence of one [2Fe-2S] cluster. The cluster is very sensitive to oxygen damage. Mixtures of the subcomplex H and of the subunits F, C and D are able to oxidize p-cresol into 4-methylcathecol, thus demonstrating the full functionality of the recombinant subunits as purified. Finally, experimental evidence is reported which strongly support a model for the electron transfer. Subunit F is the first member of an electron transport chain which transfers electrons from NADH to C, which tunnels them to H subcomplex, and eventually to molecular oxygen
- Published
- 2002
43. Conformational analysis of putative regulatory subunit D of the toluene/o-xylene-monooxygenase complex from Pseudomonas stutzeri OX1
- Author
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Alberto Di Donato, Eugenio Notomista, Piero Pucci, Paola Barbieri, Roberta Scognamiglio, Fabrizio Dal Piaz, Anna Tramontano, Scognamiglio, R., Notomista, E., Barbieri, P., Pucci, P., DAL PIAZ, F., Tramontano, A., DI DONATO, Alberto, Scognamiglio, R, Notomista, Eugenio, Barbieri, P, Pucci, Pietro, Dal Piaz, F, and Tramontano, A
- Subjects
Models, Molecular ,Protein Conformation ,Protein subunit ,medicine.disease_cause ,Pseudomona ,Biochemistry ,Article ,Protein sequencing ,Protein structure ,Bacterial Proteins ,Pseudomonas ,expression ,Genes, Regulator ,medicine ,Native state ,Limited proteolysi ,Amino Acid Sequence ,limited proteolysis ,mass spectrometry ,monooxygenase ,pseudomonas ,recombinant ,Molecular Biology ,Escherichia coli ,Peptide sequence ,biology ,Chemistry ,Hydrolysis ,biology.organism_classification ,Pseudomonas putida ,Recombinant Proteins ,Pseudomonas stutzeri ,Protein Subunits ,Models, Chemical ,Oxygenases ,Trans-Activators - Abstract
A gene cluster isolated from Pseudomonas stutzeri OX1 genomic DNA and containing six ORFs codes for toluene/o-xylene-monooxygenase. The putative regulatory D subunit was expressed in Escherichia coli and purified. Its protein sequence was verified by mass spectrometry mapping and found to be identical to the sequence predicted on the basis of the DNA sequence. The surface topology of subunit D in solution was probed by limited proteolysis carried out under strictly controlled conditions using several proteases as proteolytic probes. The same experiments were carried out on the homologous P2 component of the multicomponent phenol hydroxylase from Pseudomonas putida CF600. The proteolytic fragments released from both proteins in their native state were analyzed by electrospray mass spectrometry, and the preferential cleavage sites were assessed. The results indicated that despite the relatively high similarity between the sequences of the two proteins, some differences in the distribution of preferential proteolytic cleavages were detected, and a much higher conformational flexibility of subunit D was inferred. Moreover, automatic modeling of subunit D was attempted, based on the known three-dimensional structure of P2. Our results indicate that, at least in this case, standard modeling procedures based on automatic alignment on the structure of P2 fail to produce a model consistent with limited proteolysis experimental data. Thus, it is our opinion that reliable techniques such as limited proteolysis can be employed to test three-dimensional models and highlight problems in automatic model building.
- Published
- 2001
44. Onconase: An Unusually Stable Protein
- Author
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A. Di Donato, Giuseppe Graziano, Eugenio Notomista, Francesca Catanzano, Guido Barone, G D'Alessio, F. Dal Piaz, Notomista, E., Catanzano, F., Graziano, G., DAL PIAZ, F., Barone, G., D'Alessio, G., DI DONATO, Alberto, Notomista, Eugenio, F., Catanzano, G., Graziano, F., DAL PIAZ, G., Barone, G., D'Alessio, and And, A. DI D. O. N. A. T. O.
- Subjects
Protein Denaturation ,Circular dichroism ,Protein Conformation ,RNase P ,Proteolysis ,Mutant ,medicine.disease_cause ,Biochemistry ,Catalysis ,Ribonucleases ,Protein structure ,Enzyme Stability ,medicine ,Animals ,Chymotrypsin ,Denaturation (biochemistry) ,chemistry.chemical_classification ,Mutation ,Calorimetry, Differential Scanning ,medicine.diagnostic_test ,Circular Dichroism ,Egg Proteins ,Rana pipiens ,Hydrogen-Ion Concentration ,Pepsin A ,Peptide Fragments ,Recombinant Proteins ,Spectrometry, Fluorescence ,Enzyme ,Amino Acid Substitution ,chemistry ,Thermodynamics - Abstract
Several members of the RNase A superfamily are endowed with antitumor activity, showing selective cytotoxicity toward tumor cell lines. One of these is onconase, the smallest member of the superfamily, which at present is undergoing phase-III clinical trials as an antitumor drug. Our investigation focused on other interesting features of the enzyme, such as its unusually high denaturation temperature, its low catalytic activity, and its renal toxicity as a drug. We used differential scanning calorimetry, circular dichroism, fluorescence measurements, and limited proteolysis to investigate the molecular determinants of the stability of onconase and of a mutant, (M23L)-ONC, which is catalytically more active than the wild-type enzyme, and fully active as an antitumor agent. The determination of the main thermodynamic parameters of the protein led to the conclusion that onconase is an unusually stable protein. This was confirmed by its resistance to proteolysis. On the basis of this analysis and on a comparative analysis of the (M23L)-ONC variant of the protein, which is less stable and more sensitive to proteolysis, a model was constructed in line with available data. This model supports a satisfactory hypothesis of the molecular basis of onconase stability and low-catalytic activity.
- Published
- 2000
45. Thermal stability of onconase and some mutant forms
- Author
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Giuseppe Graziano, Francesca Catanzano, Eugenio Notomista, Alberto Di Donato, Guido Barone, Graziano, G., Notomista, E., Catanzano, F., Barone, G., and DI DONATO, Alberto
- Subjects
chemistry.chemical_classification ,biology ,Globular protein ,Mutant ,Bovine pancreatic ribonuclease ,Biochemistry ,Catalysis ,Enzyme ,Differential scanning calorimetry ,chemistry ,biology.protein ,Thermal stability ,Denaturation (biochemistry) ,Pancreatic ribonuclease ,Biotechnology - Abstract
Onconase is a small globular protein of the pancreatic ribonuclease superfamily. It is an important molecule because it possesses a selective antitumor activity. The interesting finding is that onconase has a high thermal stability, with a denaturation temperature close to 90d`C at pH 6.0. A detailed comparison between the tertiary structures of onconase and bovine pancreatic ribonuclease has been accomplished in order to identify the molecular determinants of the high stability. The results of differential scanning calorimetry measurements confirm that the mutant forms of onconase, designed to be less stable than the parent enzyme, exhibit lower denaturation temperatures. In particular, the disulfide bridge at the C-terminus of onconase seems to play a pivotal role in stability.
- Published
- 2000
46. Protein engineering of ribonucleases
- Author
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Aurora Bracale, Ennio Notomista, Giuseppe D'Alessio, Alberto Di Donato, Valeria Cafaro, Fabio Formiggini, Cafaro, V., Bracale, A., Formiggini, F., Notomista, E., D'Alessio, G., DI DONATO, Alberto, Scientific Committee of the Meeting, Cafaro, Valeria, Bracale, Aurora, Notomista, Eugenio, and D'Alessio, Giuseppe
- Subjects
Models, Molecular ,RNase P ,Protein subunit ,Mutant ,Antineoplastic Agents ,Biology ,Biochemistry ,RNase PH ,Structure-Activity Relationship ,Cytotoxic T cell ,Animals ,Ribonuclease ,antitumor ,protein engineering ,General Medicine ,Protein engineering ,Ribonuclease, Pancreatic ,Molecular biology ,Recombinant Proteins ,RNase MRP ,Amino Acid Substitution ,Mutation ,biology.protein ,Cattle ,ribonuclease - Abstract
Natural bovine seminal RNase possesses a potent antitumor action. We have mutagenized monomeric bovine pancreatic RNase A, devoid of any cytotoxic action, to insert residues present at corresponding positions in the subunit of dimeric, antitumor, seminal RNase. Like naturally dimeric seminal RNase, the mutant dimeric RNase display selective toxicity for malignant cells, which is absent in the monomeric mutants.
- Published
- 1999
47. Antimicrobial Functionalization of Surfaces by a Chimeric Adhesive Protein.
- Author
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Pitocchi R, Pennacchio A, Zuber F, Ren Q, Notomista E, Campioni S, Nyström G, Giardina P, and Piscitelli A
- Abstract
The main aim of this work is to account for the prevention and control of microbial growth on surfaces of interest in medical technology. Surface modification is often achieved by physiotherapy or chemical treatments that can involve time-consuming steps, hazardous reagents, and harsh conditions. One of the ways to overcome these drawbacks is the use of surface-active proteins such as hydrophobins. They can form stable protein layers on different surfaces, serving as anchoring points for other molecules of interest. The fungal hydrophobin Vmh2, already exploited for its adhesive ability, has been fused with the antimicrobial peptide GKY20, forming the chimeric protein used herein for functionalizing polystyrene (PS) and bacterial cellulose (BC). As a natural biomass, BC has multiple advantages, including biodegradability, low cost, renewability, high purity, and excellent mechanical properties. The chimeric protein has been proven to successfully adhere to both surfaces. A strong decrease in biofilm formation on PS and a good bactericidal effect of BC have been demonstrated. These findings provide evidence of an alternative strategy to obtain functional composites using a green, easy process.
- Published
- 2024
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48. Study of the Antimicrobial Activity of the Human Peptide SQQ30 against Pathogenic Bacteria.
- Author
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Di Napoli M, Castagliuolo G, Pio S, Di Nardo I, Russo T, Antonini D, Notomista E, Varcamonti M, and Zanfardino A
- Abstract
Given the continuous increase in antibiotic resistance, research has been driven towards the isolation of new antimicrobial molecules. Short, charged, and very hydrophobic antimicrobial peptides have a direct action against biological membranes, which are less prone to developing resistance. Using a bioinformatic tool, we chose the SQQ30 peptide, isolated from the human SOGA1 protein. The antimicrobial activity of this peptide against various Gram-negative and Gram-positive bacterial strains and against a fungal strain was studied. A mechanism of action directed against biological membranes was outlined. When administered in combination with the antibiotic ciprofloxacin and with the TRS21 (buforin II), another antimicrobial peptide, SQQ30 can be used with a lower MIC, showing additivity and synergism, respectively. Particularly interesting is the ability of SQQ30 to bind LPS in Gram-negative strains, preventing the eukaryotic cell from releasing inflammatory mediators. Our study indicates SQQ30 as a novel and promising antimicrobial agent.
- Published
- 2024
- Full Text
- View/download PDF
49. Tailoring the stress response of human skin cells by substantially limiting the nuclear localization of angiogenin.
- Author
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Culurciello R, Di Nardo I, Bosso A, Tortora F, Troisi R, Sica F, Arciello A, Notomista E, and Pizzo E
- Abstract
Human angiogenin (hANG) is the most studied stress-induced ribonuclease (RNase). In physiological conditions it performs its main functions in nucleoli, promoting cell proliferation by rDNA transcription, whereas it is strongly limited by its inhibitor (RNH1) throughout the rest of the cell. In stressed cells hANG dissociates from RNH1 and thickens in the cytoplasm where it manages the translational arrest and the recruitment of stress granules, thanks to its propensity to cleave tRNAs and to induce the release of active halves. Since it exists a clear connection between hANG roles and its intracellular routing, starting from our recent findings on heterologous ANG (ANG) properties in human keratinocytes (HaCaT cells), here we designed a variant unable to translocate into the nucleus with the aim of thoroughly verifying its potentialities under stress. This variant, widely characterized for its structural features and biological attitudes, shows more pronounced aid properties than unmodified protein. The collected evidence thus fully prove that ANG stress-induced skills in assisting cellular homeostasis are strictly due to its cytosolic localization. This study opens an interesting scenario for future studies regarding both the strengthening of skin defences and in understanding the mechanism of action of these special enzymes potentially suitable for any cell type., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors.)
- Published
- 2024
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50. The Antimicrobial, Antibiofilm and Anti-Inflammatory Activities of P13#1, a Cathelicidin-like Achiral Peptoid.
- Author
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Cafaro V, Bosso A, Di Nardo I, D'Amato A, Izzo I, De Riccardis F, Siepi M, Culurciello R, D'Urzo N, Chiarot E, Torre A, Pizzo E, Merola M, and Notomista E
- Abstract
Cationic antimicrobial peptides (CAMPs) are powerful molecules with antimicrobial, antibiofilm and endotoxin-scavenging activities. These properties make CAMPs very attractive drugs in the face of the rapid increase in multidrug-resistant (MDR) pathogens, but they are limited by their susceptibility to proteolytic degradation. An intriguing solution to this issue could be the development of functional mimics of CAMPs with structures that enable the evasion of proteases. Peptoids ( N -substituted glycine oligomers) are an important class of peptidomimetics with interesting benefits: easy synthetic access, intrinsic proteolytic stability and promising bioactivities. Here, we report the characterization of P13#1, a 13-residue peptoid specifically designed to mimic cathelicidins, the best-known and most widespread family of CAMPs. P13#1 showed all the biological activities typically associated with cathelicidins: bactericidal activity over a wide spectrum of strains, including several ESKAPE pathogens; the ability to act in combination with different classes of conventional antibiotics; antibiofilm activity against preformed biofilms of Pseudomonas aeruginosa , comparable to that of human cathelicidin LL-37; limited toxicity; and an ability to inhibit LPS-induced proinflammatory effects which is comparable to that of "the last resource" antibiotic colistin. We further studied the interaction of P13#1 with SDS, LPSs and bacterial cells by using a fluorescent version of P13#1. Finally, in a subcutaneous infection mouse model, it showed antimicrobial and anti-inflammatory activities comparable to ampicillin and gentamicin without apparent toxicity. The collected data indicate that P13#1 is an excellent candidate for the formulation of new antimicrobial therapies.
- Published
- 2023
- Full Text
- View/download PDF
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