Role of the antioxidant defense system during the production of lignocellulolytic enzymes by fungi

Fungi are used for the production of several compounds and the efficiency of biotechnological processes is directly related to the metabolic activity of these microorganisms. The reactions catalyzed by lignocellulolytic enzymes are oxidative and generate reactive oxygen species (ROS). Excess of ROS can cause serious damages to cells, including cell death. Thus, the objective of this work was to evaluate the lignocellulolytic enzymes produced by Pleurotus sajor-caju CCB020, Phanerochaete chrysosporium ATCC 28326, Trichoderma reesei RUT-C30, and Aspergillus niger IZ-9 grown in sugarcane bagasse and two yeast extract (YE) concentrations and characterize the antioxidant defense system of fungal cells by the activities of superoxide dismutase (SOD) and catalase (CAT). Pleurotus sajor-caju exhibited the highest activities of laccase and peroxidase in sugarcane bagasse with 2.6 g of YE and an increased activity of manganese peroxidase in sugarcane bagasse with 1.3 g of YE was observed. However, P. chrysosporium showed the highest activities of exoglucanase and endoglucanase in sugarcane bagasse with 1.3 g of YE. Lipid peroxidation and variations in SOD and CAT activities were observed during the production of lignocellulolytic enzymes and depending on the YE concentrations. The antioxidant defense system was induced in response to the oxidative stress caused by imbalances between the production and the detoxification of ROS

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Role of the antioxidant defense system during the production of lignocellulolytic enzymes by fungi.

Fungi are used for the production of several compounds and the efficiency of biotechnological processes is directly related to the metabolic activity of these microorganisms. The reactions catalyzed by lignocellulolytic enzymes are oxidative and generate reactive oxygen species (ROS). Excess of ROS can cause serious damages to cells, including cell death. Thus, the objective of this work was to evaluate the lignocellulolytic enzymes produced by Pleurotus sajor-caju CCB020, Phanerochaete chrysosporium ATCC 28326, Trichoderma reesei RUT-C30, and Aspergillus niger IZ-9 grown in sugarcane bagasse and two yeast extract (YE) concentrations and characterize the antioxidant defense system of fungal cells by the activities of superoxide dismutase (SOD) and catalase (CAT). Pleurotus sajor-caju exhibited the highest activities of laccase and peroxidase in sugarcane bagasse with 2.6 g of YE and an increased activity of manganese peroxidase in sugarcane bagasse with 1.3 g of YE was observed. However, P. chrysosporium showed the highest activities of exoglucanase and endoglucanase in sugarcane bagasse with 1.3 g of YE. Lipid peroxidation and variations in SOD and CAT activities were observed during the production of lignocellulolytic enzymes and depending on the YE concentrations. The antioxidant defense system was induced in response to the oxidative stress caused by imbalances between the production and the detoxification of ROS.

Conditions promoting effective very high gravity sugarcane juice fermentation.

BACKGROUND: Applying very high gravity (VHG) fermentation conditions to the sugarcane juice (SCJ) bioethanol industry would improve its environmental and economic sustainability without the need for major infrastructure changes or investments. It could enable a decrease in the consumption of biological and natural resources (cane/land, water and energy) while maintaining acceptable production parameters. The present study attempts to demonstrate and characterise an effective industrially relevant SCJ-VHG fermentation process. RESULTS: An industry-like SCJ-VHG bioethanol production process with 30 and 35 °Bx broth was employed to investigate the effects of both the yeast strain used and nitrogen source supplementation on process yield, process productivity, biomass viability, glycerol concentration and retention-associated gene expression. Process performance was shown to be variably affected by the different process conditions investigated. Highest process efficiency, with a 17% (w/v) ethanol yield and only 0.2% (w/v) sugar remaining unfermented, was observed with the Saccharomyces cerevisiae industrial strain CAT-1 in 30 °Bx broth with urea supplementation. In addition, efficient retention of glycerol by the yeast strain was identified as a requisite for better fermentation and was consistent with a higher expression of glycerol permease STL1 and channel FPS1. Urea was shown to promote the deregulation of STL1 expression, overcoming glucose repression. The consistency between Fps1-mediated ethanol secretion and ethanol in the extracellular media reinforces previous suggestions that ethanol might exit the cell through the Fps1 channel. CONCLUSIONS: This work brings solid evidence in favour of the utilisation of VHG conditions in SCJ fermentations, bringing it a step closer to industrial application. SCJ concentrated up to 30 °Bx maintains industrially relevant ethanol production yield and productivity, provided the broth is supplemented with a suitable nitrogen source and an appropriate industrial bioethanol-producing yeast strain is used. In addition, the work contributes to a better understanding of the VHG-SCJ process and the variable effects of process parameters on process efficiency and yeast strain response.

Purification, characterization and structural determination of chitinases produced by Moniliophthora perniciosa.

The enzyme chitinase from Moniliophthora perniciosa the causative agent of the witches' broom disease in Theobroma cacao, was partially purified with ammonium sulfate and filtration by Sephacryl S-200 using sodium phosphate as an extraction buffer. Response surface methodology (RSM) was used to determine the optimum pH and temperature conditions. Four different isoenzymes were obtained: ChitMp I, ChitMp II, ChitMp III and ChitMp IV. ChitMp I had an optimum temperature at 44-73ºC and an optimum pH at 7.0-8.4. ChitMp II had an optimum temperature at 45-73ºC and an optimum pH at 7.0-8.4. ChitMp III had an optimum temperature at 54-67ºC and an optimum pH at 7.3-8.8. ChitMp IV had an optimum temperature at 60ºC and an optimum pH at 7.0. For the computational biology, the primary sequence was determined in silico from the database of the Genome/Proteome Project of M. perniciosa, yielding a sequence with 564 bp and 188 amino acids that was used for the three-dimensional design in a comparative modeling methodology. The generated models were submitted to validation using Procheck 3.0 and ANOLEA. The model proposed for the chitinase was subjected to a dynamic analysis over a 1 ns interval, resulting in a model with 91.7% of the residues occupying favorable places on the Ramachandran plot and an RMS of 2.68.