Purification and biochemical characterization of a novel thermostable protease from the oyster mushroom Pleurotus sajor-caju strain CTM10057 with industrial interest
Abstract Background Proteases are hydrolytic enzymes that catalyze peptide linkage cleavage reactions at the level of proteins and peptides with different degrees of specificity. This group draws the attention of industry. More than one protease in three is a serine protease. Classically, they are a...
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| Format: | Article |
| Language: | English |
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BMC
2019-07-01
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| Series: | BMC Biotechnology |
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| Online Access: | http://link.springer.com/article/10.1186/s12896-019-0536-4 |
| _version_ | 1818250634974986240 |
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| author | Maroua Omrane Benmrad Sondes Mechri Nadia Zaraî Jaouadi Mouna Ben Elhoul Hatem Rekik Sami Sayadi Samir Bejar Nabil Kechaou Bassem Jaouadi |
| author_facet | Maroua Omrane Benmrad Sondes Mechri Nadia Zaraî Jaouadi Mouna Ben Elhoul Hatem Rekik Sami Sayadi Samir Bejar Nabil Kechaou Bassem Jaouadi |
| author_sort | Maroua Omrane Benmrad |
| collection | DOAJ |
| description | Abstract Background Proteases are hydrolytic enzymes that catalyze peptide linkage cleavage reactions at the level of proteins and peptides with different degrees of specificity. This group draws the attention of industry. More than one protease in three is a serine protease. Classically, they are active at neutral to alkaline pH. The serine proteases are researched for industrial uses, especially detergents. They are the most commercially available enzyme group in the world market. Overall, fungi produced extracellular proteases, easily separated from mycelium by filtration. Results A new basidiomycete fungus CTM10057, a hyperproducer of a novel protease (10,500 U/mL), was identified as Pleurotus sajor-caju (oyster mushroom). The enzyme, called SPPS, was purified to homogeneity by heat-treatment (80 °C for 20 min) followed by ammonium sulfate precipitation (35–55%)-dialysis, then UNO Q-6 FPLC ion-exchange chromatography and finally HPLC-ZORBAX PSM 300 HPSEC gel filtration chromatography, and submitted to biochemical characterization assays. The molecular mass was estimated to be 65 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), Native-PAGE, casein-zymography, and size exclusion by HPLC. A high homology with mushroom proteases was displayed by the first 26 amino-acid residues of the NH2-terminal aminoacid sequence. Phenylmethanesulfonyl fluoride (PMSF) and diiodopropyl fluorophosphates (DFP) strongly inhibit SPPS, revealing that it is a member of the serine-proteases family. The pH and temperature optima were 9.5 and 70 °C, respectively. Interestingly, SPPS possesses the most elevated hydrolysis level and catalytic efficiency in comparison with SPTC, Flavourzyme® 500 L, and Thermolysin type X proteases. More remarkably, a high tolerance towards organic solvent tolerance was exhibited by SPPS, together with considerable detergent stability compared to the commercial proteases Thermolysin type X and Flavourzyme® 500 L, respectively. Conclusions This proves the excellent proprieties characterizing SPPS, making it a potential candidate for industrial applications especially detergent formulations. |
| first_indexed | 2024-12-12T15:55:31Z |
| format | Article |
| id | doaj.art-495facd170eb44e9917057a555d6848b |
| institution | Directory Open Access Journal |
| issn | 1472-6750 |
| language | English |
| last_indexed | 2024-12-12T15:55:31Z |
| publishDate | 2019-07-01 |
| publisher | BMC |
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| series | BMC Biotechnology |
| spelling | doaj.art-495facd170eb44e9917057a555d6848b2022-12-22T00:19:30ZengBMCBMC Biotechnology1472-67502019-07-0119111810.1186/s12896-019-0536-4Purification and biochemical characterization of a novel thermostable protease from the oyster mushroom Pleurotus sajor-caju strain CTM10057 with industrial interestMaroua Omrane Benmrad0Sondes Mechri1Nadia Zaraî Jaouadi2Mouna Ben Elhoul3Hatem Rekik4Sami Sayadi5Samir Bejar6Nabil Kechaou7Bassem Jaouadi8Laboratory of Microbial Biotechnology and Engineering Enzymes (LMBEE), Centre of Biotechnology of Sfax (CBS), University of SfaxLaboratory of Microbial Biotechnology and Engineering Enzymes (LMBEE), Centre of Biotechnology of Sfax (CBS), University of SfaxLaboratory of Microbial Biotechnology and Engineering Enzymes (LMBEE), Centre of Biotechnology of Sfax (CBS), University of SfaxLaboratory of Microbial Biotechnology and Engineering Enzymes (LMBEE), Centre of Biotechnology of Sfax (CBS), University of SfaxLaboratory of Microbial Biotechnology and Engineering Enzymes (LMBEE), Centre of Biotechnology of Sfax (CBS), University of SfaxLaboratory of Environmental Bioprocesses (LEBP), LMI COSYS-Med, Centre of Biotechnology of Sfax (CBS), University of SfaxLaboratory of Microbial Biotechnology and Engineering Enzymes (LMBEE), Centre of Biotechnology of Sfax (CBS), University of SfaxResearch Group of Agro-Food Processing Engineering (GP2A), Laboratory of Applied Fluid Mechanics, Process Engineering and Environment, National School of Engineers of Sfax (ENIS), University of SfaxLaboratory of Microbial Biotechnology and Engineering Enzymes (LMBEE), Centre of Biotechnology of Sfax (CBS), University of SfaxAbstract Background Proteases are hydrolytic enzymes that catalyze peptide linkage cleavage reactions at the level of proteins and peptides with different degrees of specificity. This group draws the attention of industry. More than one protease in three is a serine protease. Classically, they are active at neutral to alkaline pH. The serine proteases are researched for industrial uses, especially detergents. They are the most commercially available enzyme group in the world market. Overall, fungi produced extracellular proteases, easily separated from mycelium by filtration. Results A new basidiomycete fungus CTM10057, a hyperproducer of a novel protease (10,500 U/mL), was identified as Pleurotus sajor-caju (oyster mushroom). The enzyme, called SPPS, was purified to homogeneity by heat-treatment (80 °C for 20 min) followed by ammonium sulfate precipitation (35–55%)-dialysis, then UNO Q-6 FPLC ion-exchange chromatography and finally HPLC-ZORBAX PSM 300 HPSEC gel filtration chromatography, and submitted to biochemical characterization assays. The molecular mass was estimated to be 65 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), Native-PAGE, casein-zymography, and size exclusion by HPLC. A high homology with mushroom proteases was displayed by the first 26 amino-acid residues of the NH2-terminal aminoacid sequence. Phenylmethanesulfonyl fluoride (PMSF) and diiodopropyl fluorophosphates (DFP) strongly inhibit SPPS, revealing that it is a member of the serine-proteases family. The pH and temperature optima were 9.5 and 70 °C, respectively. Interestingly, SPPS possesses the most elevated hydrolysis level and catalytic efficiency in comparison with SPTC, Flavourzyme® 500 L, and Thermolysin type X proteases. More remarkably, a high tolerance towards organic solvent tolerance was exhibited by SPPS, together with considerable detergent stability compared to the commercial proteases Thermolysin type X and Flavourzyme® 500 L, respectively. Conclusions This proves the excellent proprieties characterizing SPPS, making it a potential candidate for industrial applications especially detergent formulations.http://link.springer.com/article/10.1186/s12896-019-0536-4ProteaseOyster mushroomPleurotus sajor-cajuOrganic solventPeptide synthesisDetergent formulations |
| spellingShingle | Maroua Omrane Benmrad Sondes Mechri Nadia Zaraî Jaouadi Mouna Ben Elhoul Hatem Rekik Sami Sayadi Samir Bejar Nabil Kechaou Bassem Jaouadi Purification and biochemical characterization of a novel thermostable protease from the oyster mushroom Pleurotus sajor-caju strain CTM10057 with industrial interest BMC Biotechnology Protease Oyster mushroom Pleurotus sajor-caju Organic solvent Peptide synthesis Detergent formulations |
| title | Purification and biochemical characterization of a novel thermostable protease from the oyster mushroom Pleurotus sajor-caju strain CTM10057 with industrial interest |
| title_full | Purification and biochemical characterization of a novel thermostable protease from the oyster mushroom Pleurotus sajor-caju strain CTM10057 with industrial interest |
| title_fullStr | Purification and biochemical characterization of a novel thermostable protease from the oyster mushroom Pleurotus sajor-caju strain CTM10057 with industrial interest |
| title_full_unstemmed | Purification and biochemical characterization of a novel thermostable protease from the oyster mushroom Pleurotus sajor-caju strain CTM10057 with industrial interest |
| title_short | Purification and biochemical characterization of a novel thermostable protease from the oyster mushroom Pleurotus sajor-caju strain CTM10057 with industrial interest |
| title_sort | purification and biochemical characterization of a novel thermostable protease from the oyster mushroom pleurotus sajor caju strain ctm10057 with industrial interest |
| topic | Protease Oyster mushroom Pleurotus sajor-caju Organic solvent Peptide synthesis Detergent formulations |
| url | http://link.springer.com/article/10.1186/s12896-019-0536-4 |
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