In-depth analysis of the Aspergillus niger glucoamylase (glaA) promoter performance using high-throughput screening and controlled bioreactor cultivation techniques

Ganzlin, Markus; Rinas, Ursula

doi:10.1016/j.jbiotec.2008.04.005

Cited by 45 publications

(26 citation statements)

References 33 publications

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“…As the effect of a number of carbon sources in the performance of the GlaA expression system has been recently deeply investigated [40], various organic and inorganic nitrogen sources were tested in this work. The best McoB producer out of 15 strains tested in plate assays was grown in 100 mL of minimal medium with the following nitrogen sources: urea, NH 4 CL, casamino acids (CAA), NaNO 3 and (NH 4 ) 2 tartrate.…”

Section: Resultsmentioning

confidence: 99%

“…On the other hand, although no high titres of McoB were achieved in the culture conditions studied, a further screening of transformants has allowed the isolation of strains able to produce around 10 times higher levels to the ones presented in Figure 5 (Tamayo-Ramos et al ., manuscript in preparation). The use of strategies based on efforts done so far to boost yields of recombinant proteins (in particular laccases) [40-44], should result in new increases of McoB production levels.…”

Section: Resultsmentioning

confidence: 99%

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The Aspergillus niger multicopper oxidase family: analysis and overexpression of laccase-like encoding genes

Tamayo-Ramos

Barends²,

Verhaert³

et al. 2011

Microb Cell Fact

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BackgroundMany filamentous fungal genomes contain complex groups of multicopper oxidase (MCO) coding genes that makes them a good source for new laccases with potential biotechnological interest. A bioinformatics analysis of the Aspergillus niger ATCC 1015 genome resulted in the identification of thirteen MCO genes. Ten of them were cloned and homologously overexpressed.ResultsA bioinformatic analysis of the A. niger ATCC 1015 genome revealed the presence of 13 MCO genes belonging to three different subfamilies on the basis of their phylogenetic relationships: ascomycete laccases, fungal pigment MCOs and fungal ferroxidases. According to in silico amino acid sequence analysis, the putative genes encoding for functional extracellular laccases (mcoA, mcoB, mcoC, mcoD, mcoE, mcoF, mcoG, mcoI, mcoJ and mcoM) were placed under the control of the glaA promoter and overexpressed in A. niger N593. Enzyme activity plate assays with several common laccase substrates showed that all genes are actually expressed and code for active MCOs. Interestingly, expressed enzymes show different substrate specificities. In addition, optimization of fungal pigment MCOs extracellular production was investigated. The performance of the widely used glucoamylase signal sequence (ssGlaA) in McoA secretion was studied. Results obtained suggest that ssGlaA do not yield higher levels of secreted McoA when compared to its native secretion signal. Also, McoB synthesis was investigated using different nitrogen sources in minimal medium liquid cultures. Higher yields of extracellular McoB were achieved with (NH4)2 tartrate.ConclusionsAspergillus niger is a good source of new laccases. The different substrate specificity observed in plate assays makes them interesting to be purified and biochemically compared. The homologous signal sequence of McoA has been shown to be a good choice for its extracellular overexpression. From the nitrogen sources tested (NH4)2 tartrate has been found to be the most appropriate for McoB production in A. niger.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

The Aspergillus niger multicopper oxidase family: analysis and overexpression of laccase-like encoding genes

Tamayo-Ramos

Barends²,

Verhaert³

et al. 2011

Microb Cell Fact

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“…Pastry waste collected from local Starbucks contains 44.6% of starch and 7% of protein [8]. Starch has been shown as an inducer for GA synthesis by some Aspergillus producers [9,10,11]; thus, the significant amount of starch in pastry waste could be desirable for GA production. The smaller amount of protein, 7%, on the other hand could provide a source of nitrogen to promote the fungal growth and facilitate GA production.…”

Section: Introductionmentioning

confidence: 99%

Production of Fungal Glucoamylase for Glucose Production from Food Waste

2013

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The feasibility of using pastry waste as resource for glucoamylase (GA) production via solid state fermentation (SSF) was studied. The crude GA extract obtained was used for glucose production from mixed food waste. Our results showed that pastry waste could be used as a sole substrate for GA production. A maximal GA activity of 76.1 ± 6.1 U/mL was obtained at Day 10. The optimal pH and reaction temperature for the crude GA extract for hydrolysis were pH 5.5 and 55 °C, respectively. Under this condition, the half-life of the GA extract was 315.0 minutes with a deactivation constant (kd) 2.20 × 10−3 minutes−1. The application of the crude GA extract for mixed food waste hydrolysis and glucose production was successfully demonstrated. Approximately 53 g glucose was recovered from 100 g of mixed food waste in 1 h under the optimal digestion conditions, highlighting the potential of this approach as an alternative strategy for waste management and sustainable production of glucose applicable as carbon source in many biotechnological processes.

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“…The present study and their report share certain similarities in that the flux variation in carbon metabolism has an influence on enzyme production. With respect to the transcriptional response to glucoamylase production, an indepth characterization of the A. niger glucoamylase (glaA) promoter has been reported, clearly demonstrating that maltodextrin is a good inducer of glaA transcription, and that a low-maltose feeding protocol enabled control without accumulation of extracellular glucose, resulting in increased induction [5]. In terms of the supply of carbon substrates to the mycelia, indigestible dextrin batch culture has certain advantages.…”

Section: Enzymatic Hydrolysis Of Indigestible Dextrinmentioning

confidence: 99%

Indigestible dextrin stimulates glucoamylase production in submerged culture of Aspergillus kawachii

Sugimoto

Horaguchi

Shoji

2011

J Ind Microbiol Biotechnol

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Submerged batch cultures of Aspergillus kawachii grown on indigestible dextrin were investigated for potential improvements in glucoamylase (GA) production. In flask culture, specific GA productivities per dry weight biomass using dextrin and indigestible dextrin were 11.0 and 56.1 mU/mg-DW, respectively. Indigestible dextrin was a poor substrate for enzymatic hydrolysis. Rates of glucose formation from dextrin and indigestible dextrin by enzymatic hydrolysis were 0.477 and 0.100 mg-glucose/ml/h, respectively. For this reason, residual glucose concentrations in batch cultures grown on indigestible dextrin remained below 1.32 mg/ml where glucose-limiting conditions were easily maintained. Batch culture using indigestible dextrin had the same residual glucose profile as dextrin fed-batch culture, and nearly the same GA activity was obtained after 42.5 h of growth. However, between 42.5 and 66 h, the GA production rate of the indigestible dextrin batch culture (11.5 mU/ml/h) was higher than that of the dextrin fed-batch culture (6.5 mU/ml/h). During this period, a high amount of residual maltooligosaccharide was detected in the culture supernatant grown on indigestible dextrin. The high GA productivity observed in the indigestible dextrin batch culture may have resulted from the absence of glucose and the simultaneous presence of maltooligosaccharides throughout growth.

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In-depth analysis of the Aspergillus niger glucoamylase (glaA) promoter performance using high-throughput screening and controlled bioreactor cultivation techniques

Cited by 45 publications

References 33 publications

The Aspergillus niger multicopper oxidase family: analysis and overexpression of laccase-like encoding genes

The Aspergillus niger multicopper oxidase family: analysis and overexpression of laccase-like encoding genes

Production of Fungal Glucoamylase for Glucose Production from Food Waste

Indigestible dextrin stimulates glucoamylase production in submerged culture of Aspergillus kawachii

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