Microbiologia

URI permanente desta comunidadehttps://locus.ufv.br/handle/123456789/11840

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Autor: search.filters.author.Araújo, Elza Fernandes deData inicial: 2001Data final: 2007Assunto: search.filters.subject.Penicillium griseoroseum

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    Characterization, regulation, and phylogenetic analyses of the Penicillium griseoroseum nitrate reductase gene and its use as selection marker for homologous transformation
    (Canadian Journal Of Microbiology, 2004) Pereira, Jorge Fernando; Queiroz, Marisa Vieira de; Lopes, Francis Júlio Fagundes; Rocha, Rodrigo Barros; Daboussi, Marie-Josée; Araújo, Elza Fernandes de
    Penicillium griseoroseum has been studied because of its efficient pectinases production. In this work, the Penicillium griseoroseum nitrate reductase gene was characterized, transcriptionally analyzed in different nitrogen sources, and used to create a phylogenetic tree and to develop a homologous transformation system. The regulatory region contained consensus signals involved in nitrogen metabolism and the structural region was possibly interrupted by 6 introns coding for a deduced protein with 864 amino acids. RT-PCR analysis revealed high amounts of niaD transcript in the presence of nitrate. Transcription was repressed by ammonium, urea, and glutamine showing an efficient turnover of the niaD mRNA. Phylogenetics analysis showed distinct groups clearly separated in accordance with the classical taxonomy. A mutant with a 122-bp deletion was used in homologous transformation experiments and showed a transformation frequency of 14 transformants/microg DNA. All analyzed transformants showed that both single- and double-crossover recombination occurred at the niaD locus. The establishment of this homologous transformation system is an essential step for the improvement of pectinase production in Penicillium griseoroseum.
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    Overexpression of the plg1 gene encoding pectin lyase in Penicillium griseoroseum
    (Journal of Industrial Microbiology & Biotechnology, 2007-11-12) Teixeira, Janaina Aparecida; Ribeiro, João Batista; Teixeira, Janaina Aparecida; Queiroz, Marisa Vieira de; Araújo, Elza Fernandes de
    The pectin lyase (PL) is an industrially important enzyme since it is used for maceration and clarification in the process of fruit juice production in food industries. In order to increase the yields of pectin lyase we cloned the plg1 (pectin lyase 1) from Penicillium griseoroseum gene under the control of the strong constitutive promoter of the glyceraldehyde-3-phosphate dehydrogenase gene (gpdA) and the terminator region of the tryptophan synthetase (trpC) gene from Aspergillus nidulans (plasmid pAN52-Plg1) and transformed this construct into the P. griseoroseum strain PG63. One of the pAN52-Plg1 multi-copy transformants (strain 105) grown in culture medium containing glucose or sugar cane juice showed PL activities of 4,804 or 5,202 U ml−1 respectively, which represented 57- and 132-fold increases. In addition, the apparent specific activity of PL produced by this strain was much higher than the one observed for a commercial pectinase preparation. Evaluation of the extracellular proteins in the culture supernatant of strain 105 by SDS-PAGE showed the presence of a clear and strong band of approximately 40 kDa that probably corresponds to PL. The enzyme yields reported here demonstrate that the system we developed is able to express pectin lyase at levels comparable to, or exceeding, previously reported data.