Produção de pectina liase e poligalacturonase pela linhagem recombinante Penicillium griseoroseum T20

• Autor(a) principal: Gonçalves, Daniel Bonoto
• Data de Publicação: 2008
• Tipo de documento: Dissertação
• Idioma: por
• Título da fonte: LOCUS Repositório Institucional da UFV
• Texto Completo: http://locus.ufv.br/handle/123456789/5357
• topic: Penicillium griseoroseum; Pectina liase; Poligalacturonase; Penicillium griseoroseum; pectin lyase; Polygalacturonase; CNPQ::CIENCIAS AGRARIAS::AGRONOMIA::FITOSSANIDADE::MICROBIOLOGIA AGRICOLA

Filamentous fungi are recognized as excellent producers of extracellular enzymes and the genetically modified strains have made possible the production of pectinases with greater specificity and purity, better use of raw materials and lower production of waste. The production of pectin lyase (PL) and polygalacturonase (PG) by genetically modified strain Penicillium griseoroseum T20 has been studied. The response surface methodology (RSM) was used to optimize PL and PG production. The parameters sucrose concentration and cultivation time were evaluated. The highest PL production in Erlenmeyer flasks with 200 mL of culture medium was achieved after 87.7 h in sucrose 15.7 g/L and the highest PL estimated activity was 2428 U/ml. The higher PG production in Erlenmeyer flasks with 200 mL of culture medium was achieved after 83.8 hours and the highest PG estimated activity was 9465 U/ml. The sucrose concentration showed no significant influence on the production of this enzyme. The optimization in Erlenmeyer flasks was followed by the scaleup to 10 L bioreactor. The aeration conditions were evaluated and the highest PL and PG activity was observed in 1.0 L of air per minute. Under this condition, the strain showed low protease activity, just in the final period of cultivation, and β-glucosidase activity was not detected. The protein profile of the recombinant strain T20 showed the presence of two distinct bands of with approximately 38 and 36 kDa. These bands corresponded to PL and PG, respectively. The mycelial growth of the strain P. griseoroseum T20 has been studied by RSM, and the maximum mycelial dry weight observed was 8.63 g/L, on 30 g/L sucrose and 120 hours of cultivation. The mycelial morphology suggested a link between the occurrence of free and dispersed hyphae and the production of PL and PG. The kinetic parameters of fermentation were determined and compared between the scales of PL and PG production. The maximum total protein was 9.1 and 9.5 mg/L in cultures of 200 ml and 10 L, respectively. The specific PL (PspePLp) and PG (PspePGp) activities in relation to total protein were 353 and 613 U/g at 200 mL cultivation and 305 and 1106 U/g at 10 L cultivation, respectively. The maximum yield of PL (PdPL) and PG (PdPG) observed were 33.4 and 73.3 U/mL.h at 200 mL cultivation and 24.1 and 289 U/L.mL.h at 10 L. The performance parameters of PL (RPL/S) and PG (RPG/S) were 214 and 352 at 200 ml cultivation and 87.4 and 1049 at 10 L, respectively. The PL and PG production between P. griseoroseum T20 and P. griseoroseum wide type strain was compared. Increases of more than 400 times in the PL production and at least 14 times in the PG production were observed. The results suggest the great potential for industrial application of this strain for the production of PL and PG.