Banca de DEFESA: PRISCILA AMARAL DINIZ

Uma banca de DEFESA de MESTRADO foi cadastrada pelo programa.
STUDENT : PRISCILA AMARAL DINIZ
DATE: 30/05/2022
TIME: 08:30
LOCAL: meet.google.com/ovm-uqeh-xvg
TITLE:

DEVELOPMENT OF A HOMOLOGOUS EXPRESSION SYSTEM IN Penicillium camemberti

KEY WORDS:

biotechnology, Penicillium camemberti, genetic modifications, expression system, filamentous fungi.

PAGES: 61
BIG AREA: Ciências Biológicas
AREA: Bioquímica
SUBÁREA: Bioquímica dos Microorganismos
SUMMARY:

The genetic manipulation of organisms has shown to be one of the most promising advances in biotechnology, highlighting the products development within economic and industrial interest. In order to produce proteins, expression systems can be developed, which are protein production systems generated through organism genetic modification. Although filamentous fungi are microorganisms, which have robust protein secretion system, there is no commercially available expression system in this organism, and there is a scarcity of scientific studies using such systems. In this work, development of a homologous expression system in Penicillium camemberti was initiated. This is a filamentous fungus that has FDA’s GRAS status for lipase production, has post-translational modifications such as glycosylation, phosphorylation and disulfide bonds, secretes large amounts of enzymes in the extracellular environment, in addition to being cultivated in low-cost and have potential for genetic improvement. To study expression system, we selected a galactosidase, alpha-amylase (α-amy), from Penicillium camemberti itself, as well as the components of an ORF, open reading frame, such as the promoter and terminator sequences. For this purpose, fusion polymerase chain reaction (fusion PCR or overlap PCR) technique was used, which allows obtaining several ORFs with different promoters and terminators, making it possible optimize the constructions for the proteins production with industrial interest. To test the level of ORF expression with different constructs, were chosen constitutive and inducible promoters already used in expression vectors in other microorganisms. With the amplification of the promoter, target gene and terminator sequences, the fusion PCR has been optimized. At the same time, in silico analyzes of the alpha-amylase enzyme were performed to obtain data such as physicochemical, structural, cell location and similarity characteristics. Some data obtained were number of amino acids, molecular weight, theoretical isoelectric point, number of negatively charged residues, number of positively charged residues and hydropathicity coefficient. Preliminary studies indicated that alpha-amylase from P. camemberti is an extracellular enzyme that is easily secreted from the intracellular environment, which already indicates that this is an enzyme whose characteristics tend to be promising for large-scale production for various purposes such as in the application of the textile and food industry.

BANKING MEMBERS:
Presidente - 1757978 - DANIEL BONOTO GONCALVES
Externa ao Programa - 2254152 - MAIRA NICOLAU DE ALMEIDA
Externo à Instituição - MOACYR COMAR JUNIOR - UFU