BIMA (BIOTECNOLOGIE INDUSTRIALI, MOLECOLARI E AMBIENTALI)

Research:

ANTARCTIC MARINE BACTERIA: FROM ECOPHYSIOLOGY STUDIES TO INDUSTRIAL APPLICATIONS

BIOPLASTICS FROM WASTES: CIRCULAR ECONOMY FOR A SUSTAINABLE FUTURE

DEVELOPMENT OF IMPROVED BIOCATALYSTS AND BIOSYSTEMS TO PRODUCE ANTIOXIDANTS, ETHANOL, BIOPOLYMERS AND FINE/BULK CHEMICALS FOR INDUSTRIAL APPLICATIONS AND WASTE VALORIZATION.

POST-TRANSLATIONAL MODIFICATIONS

IN VIVO AND IN VITRO INVESTIGATION OF FUNCTIONAL PROTEIN INTERACTIONS

PROTEOMICS FOR THE INVESTIGATION OF CELL MECHANISMS AND PROTEIN COMPLEXES

REGULATION OF BIOFILM DEVELOPMENT IN GRAM NEGATIVE BACTERIA: NEW ANTIMICROBIAL STRATEGIES

SELF-ASSEMBLING PROTEIN BIOSURFACTANTS: NEW TOOLS FOR GREEN APPLICATIONS

IDENTIFICATION OF BIOMOLECULES FOR ARCHAEOMETRIC INVESTIGATIONS AND AS A TOOL FOR DIAGNOSTIC IN RESTORATION OF HISTORICAL MANUFACTS

ANTARCTIC MARINE BACTERIA: FROM ECOPHYSIOLOGY STUDIES TO INDUSTRIAL APPLICATIONS

Antarctic marine bacteria are being explored either for their molecular mechanisms of adaptation to stressing growth conditions or for the exotic chemical repertoire expressed by indigenous microbiota. The number of reports on successful isolation of novel molecules from these bacteria endowed with antimicrobial, anti-fouling, anti-cancer and biotechnological-relevant activities is steadily increasing.

The Antarctic marine bacterium Pseudoalteromonas haloplanktis TAC125 is also considered one of the most promising non conventional cell-factories for the recombinant production of difficult proteins. Our research activities are focused on the use of cold-loving bacteria either as a source of bioactive compounds to be used in several industrial and biomedical application, or as host for the recombinant protein production. These two research lines are pursued by up to date methodologies, spanning from molecular biology techniques, protein purification and characterization methods, small molecule purification and characterization. The group has also a deep expertise in the use of automatic bioreactor for the production of bioactive products.

Main collaborations:

Marco Fondi, Dept of Biology University of Florence, Italy; Alessandro Weisz, Dipartimento di Medicina, Chirurgia e Odontoiatria 'Scuola Medica Salernitana' Università degli Studi di Salerno; Victor Faundez, Department of Cell Biology, Emory University, Atlanta (USA); Charlotte Kilstrup-Nielsen, Dipartimento Di Biotecnologie E Scienze Della Vita, Università degli Studi dell'Insubria, Varese; Maria Michela Corsaro, Dipartimento Scienze Chimiche, Federico II; Rosanna Papa e Lausa Selan, Dipartimento di Sanità Pubblica e Malattie Infettive", Università di Roma "La Sapienza".

 

BIOPLASTICS FROM WASTES: CIRCULAR ECONOMY FOR A SUSTAINABLE FUTURE

The Circular Economy (CE) purpose can be described by the "3R" principle: Reduce, Reuse, and Recycle of materials and energy: following this principle, wastes should be regarded as valuable resources. Within the CE frame, this research line addresses the valorization of waste materials for the sustainable production of bioplastics. In particular, the research is focused on the designing of microbial processes for the production of microbial biopolymers, the Polyhydroxyalkanoates (PHA). Strain engineering is the key to design ad hoc cell factories for the production of tailored polymers from different raw materials. In view of process sustainability, greener alternatives to commonly used organic solvents are currently being tested for PHA recovery. The research activity also focuses on the exploitation of enzymatic tools for polymer functionalisation, providing biopolymers for different applications (food packaging, medical engineering and drug delivery). Furthermore, the exploitation of the PHA polymers for the synthesis of nanoparticles or in the formulation of composite materials is under investigation.

Main collaboration:

Cinzia Pezzella, Department of Agricultural Sciences, Federico II University, Napoli; Mario Malinconico, CNR-Institute for Polymers, Composites and Biomaterials IPCB, Via C. Flegrei 34, 80078 Naples; Georg Guebitz, Austrian Centre of Industrial Biotechnology GmbH, Division Enzymes & Polymers, Konrad Lorenz Strasse 20, 3430, Tulln an der Donau, Austria.

 

 

DEVELOPMENT OF IMPROVED BIOCATALYSTS AND BIOSYSTEMS TO PRODUCE ANTIOXIDANTS, ETHANOL, BIOPOLYMERS AND FINE/BULK CHEMICALS FOR INDUSTRIAL APPLICATIONS AND WASTE VALORIZATION.

Novel feruloyl esterases and glucuronyl esterases were developed to replace chemical processes for the production of antioxidants for the cosmetics with bioconversions based on esterification reactions. In order to produce biofuels and high‐added value bioproducts with applications in chemical and plastic industries, a toolkit of cellulases, hemicellulases and auxiliary enzymes with better performances in hydrolysis of pretreated lignocellulose was developed. The improved biocatalysts are developed by both isolation from new microorganims and genetic engineering and metagenomics approaches. Faraco's labs are equipped with an automated workstation consisting of the robots Colony Picker QPix450 (Molecular Devices) and Biomek® NX for automatic manipulation and analysis of thousands of microorganisms and enzymes.

Main collaboration:

Collaborations in the frame of the project "Enhanced bioconversion of agricultural residues through cascading use" (BIOrescue), "H2020-BBI-PPP-2015-2-1", contract number 720708

 

 

POST-TRANSLATIONAL MODIFICATIONS

Posttranslational modification of proteins refers to the chemical changes proteins may undergo after translation. Such modifications are mostly catalyzed by enzymes that recognize specific target sequences in specific proteins. The most common modifications are the covalent addition or removal of low-molecular-weight groups, such as acetylation, glycation (nonenzymatic conjugation with carbohydrates), glycosylation (enzymatic conjugation with carbohydrates), phosphorylation, oxidation, etc. Posttranslational modifications play a fundamental role in regulating the folding of proteins, their targeting to specific subcellular compartments, their interaction with ligands or other proteins, and their functional state. A very powerful way to study posttranslational modifications is by ‘proteomics', an extremely rapid and sensitive methodology for the systematic identification of proteins. This involves separation of proteins (two-dimensional gel electrophoresis, affinity chromatography etc) followed by mass spectrometry. The technique reveals very precisely the site and nature of posttranslational modifications.

Main Collaborations:

1) Prof. J.C. Samuelson, Boston University School of Medicine

2) Dr. Chiara Guerrera, Necker  Faculté de Médecine, Université de Paris

 

 

IN VIVO AND IN VITRO INVESTIGATION OF FUNCTIONAL PROTEIN INTERACTIONS

The investigation of functional protein interactions in vivo encompasses the identification of protein components of protein complexes as well as the proteins involved in functional networks. Moreover, in vitro analyses of protein complexes three-dimensional structure constitutes a fundamental prerequisite for the elucidation of their biological functions at molecular level.

The investigation of protein complexes and functional protein networks in vivo is carried out by using functional or differential proteomics approaches, respectively. Both approaches, although are relied on different experimental workflows, are addresses to the comprehension of cell mechanisms (differentiation, proliferation protein trafficking, chromatin remodeling, viral infection etc) both in physiological and in pathological conditions, leading to the definition of the molecular basis of genetic (LSDs, Wilson Disease, Anemia), chronic (diabetes) infectious (Covid-19), neurodegenerative (Huntington, Alzheimer, Parkinson, etc) and oncologic (glioma, sarcoma, thyroid tumors) diseases.

Protein-ligands interactions (proteins, lipids, DNA/RNA, drugs) are also in vitro investigated   by direct ESI-MS analyses, for the definition of complexes stoichiometry and binding modalities. Moreover, by using limited proteolysis and cross-lynking experiments, coupled to mass spectrometry, it is possible to map protein-ligand interacting regions. This approach has been successfully applied in the investigation of amylodogenic state of several proteins involved in amyloidosis, leading to a model for the description of their aggregation process and to probe, by epitope mapping, the antigens-antibodies interaction region.

Main collaboration:

Prof. V. Bellotti, University of Pavia and Visiting Researcher at The Centre for Amyloidosis and Acute Phase Proteins, Royal Free and University College Medical School, UCL, London, UK; TIGEM (Telethon Institute of Genetics and Medicine); dell'IRCCS-Casa Sollievo della Sofferenza (FG); Prof.ssa C. Zuccato, Università di Milano; Dompè, Dr.ssa Iaconis; CEINGE COVID-19 Network.

 

PROTEOMICS IN CULTURAL HERITAGE

The term proteomics for cultural heritage refer to the study of proteins in ancient samples such as archaeological objects and works of art by applying proteomic strategies. In case of archaeological objects and artwork, it can reveal ancient human habits, commercial exchanges, but also important information to understand manufacturing processes, the technique used by an artist, and therefore it provides essential information for art historians. It may also support the choice of the most appropriate conservation conditions or restoration procedures. It demands for the adaptation of standard protein chemistry to ancient and degraded proteins to understand the degradation processes and the development of more and more sensitive analytical procedures.

Main collaboration:

Prof. Ilaria Bonaduce, Department of Chemistry and Industrial Chemistry, University of Pisa, Pisa, Italy;

Prof. Enrico Cappellini, University of Copenhagen, Copenhagen, Denmark

Prof. Caroline Tokarski, Institute of Chemistry and Biology of Membranes & Nanoobjects, University of Bordeaux, France

Prof. Matthew Collins, University of Copenhagen, Copenhagen, Denmark

REGULATION OF BIOFILM DEVELOPMENT IN GRAM NEGATIVE BACTERIA: NEW ANTIMICROBIAL STRATEGIES

Biofilm formation in static conditions was investigated by differential proteomics approaches. In E.coli the involvement of the N-acetylneuraminase protein NanA was demonstrated; particularly the inhibition of NanA substantially decreases the formation of the biofilm. Similarly M. smegmatis biofilm formation is linked to the overexpression of the bifunctional enzyme GlmU involved in bacterial cell wall synthesis. The results obtained indicate that inhibition of NanA or GlmU strongly decreases the ability of bacteria to form biofilms, suggesting that the two enzymes could be specific targets of new therapeutic strategies.

The effect of Temporin-L (TL) on biofilm formation in Pseudomonas fluorescens in dynamic conditions was examined using Confocal Laser Scanning Microscopy and quantified via image analysis showing that TL exerted antibiofilm activity, impairing biofilm formation. Moreover, TL also affected mature biofilm as large portion of preformed biofilm architecture was clearly perturbed with a significative decrease of all the biofilm surface properties and the overall biomass.

The molecular mechanism of the antimicrobial peptide Temporin-L was investigated by functional proteomics approaches and the putative peptide interactors from bacterial membrane proteins were identified by advanced mass spectrometry procedures. The results demonstrated that Temporina-L interacts with the FtsZ protein, an essential component of the divisoma complex, and with the elongasoma proteins thus inhibiting bacterial cell division.

Main collaborations:

Franz Cemiˇc, Department of Mathematics, Natural Sciences and Computer Science, University of Applied Sciences Giessen, Giessen, Germany; Marco Rinaldi Oggioni, Department of Genetics and Genome Biology, University of Leicester, Leicester, UK; Stefano Guido and Sergio Caserta, Department of Chemical, Materials and Industrial Production Engineering, University of Naples "Federico II"; Alessandra Romanelli, Department of Pharmaceutical Sciences, University of Milan; Laura Selan and Marco Artini, Department of Public Health and Infectious Diseases, Sapienza University of Rome; Gianna Tempera, Department of Biomedical and Biotecnological Sciences, University of Catania; Giovanni Lentini e Maria Maddalena Cavalluzzi, Department of Pharmacy-Pharmaceutical Sciences, University of Bari Aldo Moro, Bari.

 

SELF-ASSEMBLING PROTEIN BIOSURFACTANTS: NEW TOOLS FOR GREEN APPLICATIONS

Filamentous fungi produce proteinaceous biosurfactants, whose activity is intrinsic to the protein molecules themselves, being some hydrophobic aliphatic side chains exposed on one side of their surface, whilst polar or charged residues are confined to the other side. Among fungal biosurfactants, hydrophobins (HFB) are described as the most powerful surface-active proteins known. HFBs self-assemble and can form highly insoluble aggregates similar to amyloid fibrils (functional amyloids). The Janus-faced character of HFBs allows to tune the wettability of surfaces and they can be used for several biotechnological applications, such as surface coating and functionalization. Enzymes, peptides, antibodies and nanomaterials can be immobilized on functionalized surfaces for biosensing and biomedical applications. Up to now three HFBs and other fungal protein biosurfactants were characterized and exploited for the above-mentioned applications.

Main collaboration:

Luca De Stefano, Institute for Microelectronics and Microsystems, CNR, Naples;

Giovanna Cristina Varese, Scientific Head of the Mycotheca Universitatis Taurinensis, Department of Life Sciences, University of Turin;

Alan Le Goff, CNRS, Departement de chimie moleculaire, Grenoble, France.

 

IDENTIFICATION OF BIOMOLECULES FOR ARCHAEOMETRICAL INVESTIGATIONS AND FOR DIAGNOSTIC IN RESTORATION OF HISTORIC-ARTISTIC MANUFACTS

Cultural heritage represents our history, identity and bond to the past. The link between Science and Cultural Heritage is becoming more and more important. The materials used for the creation of art works (such as binders, pigments, thickeners) are complex heterogeneous mixtures of different compounds (for example lipids, oligosaccharides, etc) and their identification and characterization represent an important task. At the same time, the characterization of historical artefacts at a molecular level is becoming an increasingly important aspect for Archaeometric investigations.

Because of their importance, minimum amounts of such samples are available for analysis. Therefore, it is necessary to use broad-spectrum approach for both extraction and analysis of the organic fraction. In this context, mass spectrometry (MS) is a useful methodology especially for the sensitivity and the ability to work with complex matrices.

The approaches should not only prevent damage but also choose the most appropriate way to handle the samples, in order to extrapolate as much information as possible.

Main Collaborations:

Prof. C. Cheung, Department of Classics, Princeton (NJ)

Prof. A. Naso, Dipartimento di Studi umanistici – Federico II, Napoli

Prof. G.Trojsi, Dipartimento di Scienze umanistiche, Suor Orsola Benincasa, Napoli.