Facoltà/Dipartimento Dip.Scienze Agrarie,Alimentari e Ambientali
Anno Accademico 2018
Data dell'esame finale 2019-12-05
Titolo italiano Heterologous protein expression of the bovine lipoprotein lipase in Aspergillus nidulans for milk preservation.
Titolo inglese Heterologous protein expression of the bovine lipoprotein lipase in Aspergillus nidulans for milk preservation.
Abstract in inglese The Lipoprotein Lipase (LPL) is a highly conserved protein with a key role in triglycerides (TGs) metabolism. The enzyme catalyses the hydrolysis of TGs into monoacylglycerols and fatty acids (FFA) in the bloodstream and inside mammocytes. In humans, defective LPL causes a rare and potentially fatal disease named Familial Chylomicronemia Syndrome (FCS). LPL is also found in cow’s milk, where it could be responsible for rancidity, acting on fat globules. Bovine LPL (bLPL) is usually purified from cow’s milk albeit with very low yields. In order to deeply investigate the role and potential biotechnological applications of bLPL, it is essential to establish a system for the recombinant production of this protein in its active form. To this purpose a new system based on Aspergillus nidulans is here presented. A. nidulans is a filamentous fungus, of the phylum Ascomycota. Aspergillus genome has been sequenced and the availability of data allows gene manipulation of this microbial genus for heterologous protein expression. Moreover, A. nidulans is cheap to grow, with the possibility of utilizing several selection markers and inducible gene promoters. In this work AlcA plasmid is used as vector for the recombinant expression of bLPL, under the control of the ethanol-inducible Alcohol dehydrogenase promoter. Obtained Aspergillus recombinant strains were analysed for the presence of bLPL sequence within the genome and assayed for their viability after induction of the protein expression. Selected transformants were then tested for their capability to express bLPL through proteomic analyses. Lastly, affinity chromatography was applied to the produced bLPL to establish the purification procedure.
Appare nelle tipologie: Laurea specialistica, magistrale, ciclo unico
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