Curci, Nicola (2021) Enzymes discovery and characterization: microbial hydrolases for sustainable bioconversion and novel applications. [Tesi di dottorato]


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Item Type: Tesi di dottorato
Resource language: English
Title: Enzymes discovery and characterization: microbial hydrolases for sustainable bioconversion and novel applications
Date: 13 July 2021
Number of Pages: 186
Institution: Università degli Studi di Napoli Federico II
Department: Biologia
Dottorato: Biotecnologie
Ciclo di dottorato: 33
Coordinatore del Corso di dottorato:
Cobucci-Ponzano, BeatriceUNSPECIFIED
Date: 13 July 2021
Number of Pages: 186
Keywords: Enzymes discovery, enzymes characterization; biotechnology.
Settori scientifico-disciplinari del MIUR: Area 05 - Scienze biologiche > BIO/10 - Biochimica
Date Deposited: 22 Jul 2021 16:07
Last Modified: 07 Jun 2023 10:44

Collection description

The development of sustainable processes and the society needs to shift towards a circular bioeconomy have increased the demand for biocatalysts in biotechnology to support industrial processes. The discovery of novel enzymes with higher activity and stability than those of catalysts already available paves the way for improving current industrial bioprocesses and developing novel applications. Microbial enzymes represent the bulk of the enzymes market, being more active and stable, producing high yield, and microorganisms represent an easily exploitable source by modern metagenomics technique. Among the more sought-after enzymes for industrial and biotechnological purposes, glycoside hydrolases (GHs) and lipolytic enzymes have increased their impact in this field. The present thesis describes different strategies to identify novel enzymes and the evaluation of several GH activities for their potential exploitation in biotechnological applications. Chapter 2 reports the exploration of microbial consortia populating two mud pools in Pisciarelli solfatara using a shotgun metagenomic approach, that led to the identification of 586 putative CAZymes. In this work, I focused on the characterization of a GH109 with a previously unreported β-N-acetylglucosaminide/β-glucoside specificity. By using a genome mining approach, chapter 3 describes the identification of a novel thermophilic esterase (EstGtA3) from the thermophilic bacterium G. thermodenitrificans. The characterization revealed that the enzyme is active at 60°C and in a wide range of pH. Moreover, EstGtA3 showed an activating effect in n-hexane and other denaturing agents, making this enzyme suitable for biotechnological applications. More advanced studies on possible industrial applications of some GHs are reported in chapters 4 and 5. In chapter 4 the characterization of the mechanism of action of three thermostable GHs (LacS, XylS, and SsαFuc) from the hyperthermophilic archaeon S. solfataricus on xyloglucan oligosaccharides shows excellent operational stability at 65°C and pH 5.5 of the three enzymes. SsαFuc was able to remove all fucose residues, while LacS and XylS showed a strong synergy for the hydrolysis of these substrates. The last chapter, in collaboration with Novozymes A/S company, describes the characterization of three novel GH109 (NAg68, NAg69, and NAg71). Interestingly, these enzymes showed the ability to remove the immunogenic determinant N-acetylgalactosamine from the erythrocyte on the surface of group A blood, converting it into the universal donor group 0. In particular, NAg71 requires less enzyme concentration for conversion than the already available enzymes used for this biotechnological application.


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