Biochemical and Functional Characterization of a Metalloprotease from the Thermophilic Fungus<i>Thermoascus aurantiacus</i>

Merheb-Dini, Carolina; Cabral, Hamilton; Leite, Rodrigo Simões Ribeiro; Zanphorlin, L.M.; Okamoto, Débora N.; Bonilla-Rodriguez, Gustavo Orlando; Juliano, Luiz; Arantes, Eliane Candiani; Gomes, Eleni; Silva, Roberto da

doi:10.1021/jf9017977

Cited by 34 publications

(25 citation statements)

References 39 publications

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“…The fungal suspensions were obtained by gently scraping the surface of the culture medium and putting it into 30 mL of nutrient solution (0.1% ammonium sulfate, 0.1% magnesium sulfate heptahydrate, and 0.1% ammonium nitrite, Merheb-Dini et al [16]). The fungi were inoculated in the agroindustrial residues by transfer of 5 mL of the microbial suspension.…”

Section: Methodsmentioning

confidence: 99%

Production and Catalytic Properties of Amylases fromLichtheimia ramosaandThermoascus aurantiacusby Solid-State Fermentation

Oliveira

Silvestre

Garcia

et al. 2016

The Scientific World Journal

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The present study compared the production and the catalytic properties of amylolytic enzymes obtained from the fungi Lichtheimia ramosa (mesophilic) and Thermoascus aurantiacus (thermophilic). The highest amylase production in both fungi was observed in wheat bran supplemented with nutrient solution (pH 4.0) after 96 hours of cultivation, reaching 417.2 U/g of dry substrate (or 41.72 U/mL) and 144.5 U/g of dry substrate (or 14.45 U/mL) for L. ramosa and T. aurantiacus, respectively. The enzymes showed higher catalytic activity at pH 6.0 at 60°C. The amylases produced by L. ramosa and T. aurantiacus were stable between pH 3.5–10.5 and pH 4.5–9.5, respectively. The amylase of L. ramosa was stable at 55°C after 1 hour of incubation, whereas that of T. aurantiacus maintained 60% of its original activity under the same conditions. Both enzymes were active in the presence of ethanol. The enzymes hydrolyzed starch from different sources, with the best results obtained with corn starch. The enzymatic complex produced by L. ramosa showed dextrinizing and saccharifying potential. The enzymatic extract produced by the fungus T. aurantiacus presented only saccharifying potential, releasing glucose monomers as the main hydrolysis product.

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Section: Methodsmentioning

confidence: 99%

Production and Catalytic Properties of Amylases fromLichtheimia ramosaandThermoascus aurantiacusby Solid-State Fermentation

Oliveira

Silvestre

Garcia

et al. 2016

The Scientific World Journal

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“…(Thys and Brandelli, 2006), Penicillium spp. (El-Gendy, 2010), Thermoascus aurantiacus (Merheb-Dini et al, 2009), and Streptomyces septatus TH-2 (Hatanaka et al, 2005) produce enzymes with a neutral pH optimum.…”

Section: Discussionmentioning

confidence: 99%

Analysis of the Specificity and Biochemical Characterization of Metalloproteases Isolated from Eupenicillium javanicum Using Fluorescence Resonance Energy Transfer Peptides

Neto

Oliveira

et al. 2017

Front. Microbiol.

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Enzymes have important features that may facilitate their application in industrial processes and have been used as alternatives to chemical catalysts. In particular, proteases can be isolated from microorganisms, which provide important sources of advantageous enzymes for industrial processes. For example, Eupenicillium javanicum is a filamentous fungus that has been shown to express industrially applicable enzymes and chemical components, such as antifungal compounds. The biotechnological potential of E. javanicum and proteases made us search a novel protease from this microorganism. The macromolecule was isolated, the main biochemical properties was evaluated, and the specificity of the protease subsites was determined. The protease was produced under solid-state bioprocess with wheat bran and isolated by two chromatography steps with yield of 27.5% and 12.4-fold purification. The molecular mass was estimated at 30 kDa. The N-terminal sequence of the first 20 amino acid residues was AVGAGYNASVALALEKALNN. The enzyme presented higher proteolytic activity at pH 6.0 and 60°C. The protease is stable at wide range of pH values and temperatures and in the presence of surfactants. The “primed” side of the catalytic site showed the highest catalytic efficiency of the enzyme isolated from E. javanicum. The S′1 subsite is responsible for catalyzing the protease reaction with substrates with tyrosine in P′1. These findings provide important insights into the biochemical characterization of a highly active protease from E. javanicum and may facilitate the development of industrial processes involving this protease.

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“…In D. mulleri ( Figure 9A), Fe +2 increased fivefold the EST-4 activity compared to the control. Knowing that ions can bind to amino acids and influence protein structural conformation, directly affecting the catalytic performance of the enzyme (Merheb-Dini et al 2009), we were able to propose that Fe +2 bound to EST-4 of D. mulleri and improved the enzyme activity probably because it made the enzyme structure better organized. Na + and Ba +2 also showed a positive modulation effect, increasing around 50% of the EST-4 activity.…”

Section: Effect Of Chemicals On the Est-4 Activitymentioning

confidence: 91%

Purification and characterization of a specific late-larval esterase from two species of the Drosophila repleta group: contributions to understand its evolution

et al. 2014

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Background: After duplication, one copy of an original gene can become redundant and decay toward a pseudogene status or functionally diverge. Here, we performed the purification and biochemical characterization of EST-4 (a late larval β-esterase) from two Drosophila repleta group species, Drosophila mulleri and Drosophila arizonae, in order to establish comparative parameters between these enzymes in these species and to contribute to better understand their evolution. Results: In D. mulleri, EST-4 had an optimal activity in temperatures ranging from 40°to 45°C and at pH 7.5, maintaining stability in alkaline pH (8.0 to 10.0). It was classified as serine esterase as its activity was inhibited by PMSF. No ion negatively modulated EST-4 activity, and iron had the most positive modulating effect. In D. arizonae, it showed similar optimum temperature (40°C), pH (8.0), and was also classified as a serine esterase, but the enzymatic stability was maintained in an acidic pH (5.5 to 6.5). Fe +2 had the opposite effect found in D. mulleri, that is, negative modulation. Al +3 almost totally inhibited the EST-4 activity, and Na + and Cu +2 had a positive modulation effect. Kinetic studies, using ρ-nitrophenyl acetate as substrate, showed that EST-4 from D. mulleri had higher affinity, while in D. arizonae, it showed higher V max and catalytic efficiency in optimal reaction conditions. Conclusions: EST-4 from D. mulleri and D. arizonae are very closely related and still maintain several similar features; however, they show some degree of differentiation. Considering that EST-4 from D. mulleri has more conspicuous gel mobility difference among all EST-4 studied so far and a lower catalytic efficiency was observed here, we proposed that after duplication, this new copy of the original gene became redundant and started to decay toward a pseudogene status in this species, which probably is not occurring in D. arizonae.

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Biochemical and Functional Characterization of a Metalloprotease from the Thermophilic FungusThermoascus aurantiacus

Cited by 34 publications

References 39 publications

Production and Catalytic Properties of Amylases fromLichtheimia ramosaandThermoascus aurantiacusby Solid-State Fermentation

Production and Catalytic Properties of Amylases fromLichtheimia ramosaandThermoascus aurantiacusby Solid-State Fermentation

Analysis of the Specificity and Biochemical Characterization of Metalloproteases Isolated from Eupenicillium javanicum Using Fluorescence Resonance Energy Transfer Peptides

Purification and characterization of a specific late-larval esterase from two species of the Drosophila repleta group: contributions to understand its evolution

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