Gel mobility shift scanning of pectin-inducible promoter from Penicillium griseoroseum reveals the involvement of a CCAAT element in the expression of a polygalacturonase gene

Ribon, Andréa de Oliveira Barros; Ribeiro, João Batista; Gonçalves, Daniel Bonoto; Queiroz, Marisa Vieira de; Araújo, Elza Fernandes de

doi:10.1590/s1415-47572009005000021

Cited by 3 publications

(2 citation statements)

References 22 publications

(25 reference statements)

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“…Worldwide interest in pectinolytic enzymes has increased particularly because of their efficacy in improving fruit juice quality and high industrial value [8]. Polygalacturonases (PG) are among the most extensively studied pectinolytic enzymes [9]. PG catalyse the hydrolytic cleavage of the polygalacturonic acid chain by the introduction of a water molecules across each oxygen bridge [10].…”

Section: Introductionmentioning

confidence: 99%

Purification of polygalacturonases produced by Aspergillus niger using an aqueous two-phase system

Maciel

Ottoni

Herculano

et al. 2014

Fluid Phase Equilibria

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The partitioning and purification of polygalacturonases (PG) produced by Aspergillus niger URM 5162 were investigated in aqueous two-phase systems (ATPS), formed by polyethylene glycol and phosphate salts (PEG/phosphate). To evaluate the effect of the 4 independent variables-molar mass of polyethylene glycol (PEG) (400-8000 g/mol-M PEG), PEG concentration (12.5-17.5%, w/w-C PEG), phosphate concentration (15-25%, w/w, C PHOS) and pH (6.0-8.0)-on the 4 response variables: partition coefficient (K), activity yield (Y), purification factor (PF) and selectivity (S), a factorial design (2 4) was used. The endopolygalacturonases (endo-PG) and exo-polygalacturonase (exo-PG) were preferentially partitioned in the top phase. For endo-and exo-PG, the highest values for the response variables K (1.23 and 2.40), Y (74.04% and 17.97%), PF (8.18 and 1.98) and S (24.68 and 48.07), respectively, were obtained for a C PEG of 12.5% (w/w), M PEG of 8000 g/mol, and C PHOS of 25% (w/w) at pH 6.0. These conditions were considered the most suitable for the purification of PG produced by A. niger URM 5162. Furthermore, the most important independent variables for endo-and exo-PG were C PHOS and M PEG , respectively. All independent variables studied and their interactions significantly influenced the response variables. According to these results, the PEG/phosphate system is a useful cost-effective alternative for purification of PG produced by A. niger URM 5162.

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Section: Introductionmentioning

confidence: 99%

Purification of polygalacturonases produced by Aspergillus niger using an aqueous two-phase system

Maciel

Ottoni

Herculano

et al. 2014

Fluid Phase Equilibria

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“…In this context, our group isolated and characterized two genes encoding PL (plg1 and plg2) and two genes encoding PG (pgg2 and pgg1) in P. griseoroseum (Ribon et al 2002a,b;Bazzolli et al 2006) and studied their regulation (Bazzolli et al 2008;Ribon et al 2009). Genes plg1 and pgg2 are single copies in the genome of P. griseoroseum and make the greatest contribution to the production of PL and PG.…”

Section: Introductionmentioning

confidence: 99%

Improved pectinase production in Penicillium griseoroseum recombinant strains

Teixeira

Gonçalves

Queiroz

et al. 2011

Journal of Applied Microbiology

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Aims: To obtain recombinant strains of Penicillium griseoroseum that produce high levels of pectin lyase (PL) and polygalacturonase (PG) simultaneously. Methods and Results: A strain with high production of PL was transformed with the plasmid pAN52pgg2, containing the gene encoding PG of P. griseoroseum, under control of the gpd promoter gene from Aspergillus nidulans. Southern blot analysis demonstrated that all strain had at least one copy of pAN52pgg2 integrated into the genome. The recombinant strain P. griseoroseum T20 produced levels of PL and PG that were 266‐ and 27‐fold greater, respectively, than the wild‐type strain. Furthermore, the extracellular protein profile of recombinant T20 showed two protein bands of c. 36 and 38 kDa, associated with PL and PG, respectively. Conclusions: This recombinant strain T20 produces PL and PG using carbon sources of low costs, and an enzyme preparation that is free of cellulolytic and proteolytic activities. Significance and Impact of the Study: PL and PG play an important role in the degradation of pectin. Owing to their use in the juice and wines industries, there is a growing interest in the inexpensive production of these enzymes. This work describes an efficient system of protein expression and secretion using the fungus P. griseoroseum.

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Genome organization and assessment of high copy number and increased expression of pectinolytic genes from Penicillium griseoroseum: a potential heterologous system for protein production

Teixeira

Nogueira

Queiroz

et al. 2014

Journal of Industrial Microbiology and Biotechnology

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The fungus Penicillium griseoroseum has the potential for application on an industrial scale as a host for the production of homologous and heterologous proteins, mainly because it does not produce some mycotoxins or secrete proteases under the growth conditions for pectinase production. However, for the fungus to be used effectively as an expression heterologous system, an understanding of the organization of its genome, as well as the mechanisms of gene expression and protein production, is required. In the present study, the size of the P. griseoroseum genome was estimated to be 29.8-31.5 Mb, distributed among four chromosomes. An analysis of plg1 and pgg2 pectinolytic genes expression and copy number in recombinant multi-copy strains of P. griseoroseum demonstrated that an increase in the number of gene copies could increase enzyme production, but the transcription could be affected by the gene integration position. Placing a copy of the plg1 gene under the control of the gpd promoter of Aspergillus nidulans yielded a 200-fold increase in transcription levels compared to the endogenous gene, and two copies of the pgg2 gene produced an 1100-fold increase compared with the endogenous gene. These results demonstrated that transcription, translation, and protein secretion in the fungus P. griseoroseum respond to an increased number of gene copies in the genome. The processing capacity and efficiency of protein secretion in P. griseoroseum are consistent with our premise that this fungus can be used for the industrial-scale production of several enzymes.

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Gel mobility shift scanning of pectin-inducible promoter from Penicillium griseoroseum reveals the involvement of a CCAAT element in the expression of a polygalacturonase gene

Cited by 3 publications

References 22 publications

Purification of polygalacturonases produced by Aspergillus niger using an aqueous two-phase system

Purification of polygalacturonases produced by Aspergillus niger using an aqueous two-phase system

Improved pectinase production in Penicillium griseoroseum recombinant strains

Genome organization and assessment of high copy number and increased expression of pectinolytic genes from Penicillium griseoroseum: a potential heterologous system for protein production

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