Production and partial characterization of serine and metallo peptidases secreted by Aspergillus fumigatus Fresenius in submerged and solid state fermentatio

Abstract

Enzyme production varies in different fermentation systems. Enzyme expression in different fermentation systems yields important information for improving our understanding of enzymatic production induction. Comparative studies between solid-state fermentation (SSF) using agro-industrial waste wheat bran and submerged fermentation (SmF) using synthetic media were carried out to determinate the best parameters for peptidase production by the fungus Aspergillus fumigatus Fresen. Variables tested include: the concentration of carbon and protein nitrogen sources, the size of the inoculum, the pH of the media, temperature, and the length of the fermentation process. The best peptidase production during SSF was obtained after 96 hours using wheat bran at 30 °C with an inoculum of 1 × 106 spores and yielded 1500 active units (U/mL). The best peptidase production using SmF was obtained after periods of 72 and 96 hours of fermentation in media containing 0.5% and 0.25% of casein, respectively, at a pH of 6.0 and at 30 °C and yielded 40 U/mL. We also found examples of catabolite repression of peptidase production under SmF conditions. Biochemical characterization of the peptidases produced by both fermentative processes showed optimum activity at pH 8.0 and 50 °C, and also showed that their proteolytic activity is modulated by surfactants. The enzymatic inhibition profile using phenylmethylsulfonyl fluoride (PMSF) in SmF and SSF indicated that both fermentative processes produced a serine peptidase. Additionally, the inhibitory effect of the ethylene-diaminetetraacetic acid (EDTA) chelating agent on the peptidase produced by SmF indicated that this fermentative process also produced a metallopeptidase.

Bacterial and Fungal Proteolytic Enzymes: Production, Catalysis and Potential Applications

“…The production of metallopeptidase from fungi has been reported in several studies (e.g., Fernandez et al [36] and Silva et al [29] with A. fumigatus, Neto et al [37] with Eupenicillium javanicum, Merheb-Dini et al [13] with the fungus T. aurantiacus, and Jashni et al [38] with Fusarium oxysporum f. sp. lycopersici.…”

“…Several studies have showed the production of serine peptidase from bacteria and fungi, for example, Silva et al [3] with Penicillium corylophilum and Penicillium waksmanii, Silva et al [29] and Silva et al [4] with A. fumigatus, Graminho et al [30] with P. waksmanii, Biaggio et al [15] with A. terreus, Ida et al [31] with Aspergillus fischeri and Penicillium citrinum, a thermostable peptidase from Geobacillus stearothermophilus [32], alkaline serine peptidase from Bacillus lehensis [33], subtilisin from B. licheniformis and Bacillus amyloliquefaciens [24], and keratinases from Chryseobacterium L99 sp. nov. [34] and Bacillus halodurans PPKS-2 [35].…”

“…These peptidases exhibit a wide catalytic variety for proteolysis, such as collagenase and elastase (bacteria and fungi), hemorrhagic toxin of snake venoms, and bacterial thermolysin [24]. Metallopeptidases exhibit inhibition by chelating agents, such as ethylenediaminetetraacetic acid (EDTA) and ethyleneglycoltetraacetic acid (EGTA), whose catalysis is influenced by the removal of divalent ions [13,29,37].…”

“…Two different systems, SmB and SSB, are used in microbial processes. In SmB, the variations in the composition of the culture medium, pH, and carbon and nitrogen sources affect the yield and diversity in peptidase expression [29,69]. SmB has various advantages such as ease of control of physical and chemical variables (pH, temperature, and oxygenation of the culture medium) and better distribution of medium components [29].…”

Solid-State Fermentation as a Novel Paradigm for Organic Waste Valorization: A Review

“…Similarly, peptidase production by Aspergillus fumigatus in SSF using wheat bran and SmF using casein as a substrate was compared. An improved yield of peptidase production, approximately 30 times higher, was obtained through SSF compared to SmF [115]. In another comparative study among SSF, slurry state fermentation (SlSF) and SmF, revealed that maximum β-xylosidase was obtained in SSF (33.7 U·mL −1 ) followed by SlSF (24.9 U·mL −1 ) and SmF (5.5 U·mL −1 ) [129].…”