title: Purification and biochemical characterization of a novel protease from Penicillium digitatum - Use in bioactive peptides production.
abstract: This work reports the production of a novel serine protease enzyme (P. dig-protease) from the fungus Penicillium digitatum. The protease was purified from the culture supernatant to homogeneity using ammonium sulfate precipitation, Sephadex G-150 gel filtration and carboxymethyl-sepharose ion exchange chromatography with a 13-fold increase in specific activity. The apparent molecular weight of P.dig-protease was estimated to be 120 kDa by native high performance liquid chromatography (HPLC), sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) revealed a single polypeptide at about 30 kDa that indicates a tetrameric protein. The proteolytic activity was inhibited by phenylmethylsulfonyl fluoride suggesting a serine-protease enzyme. P.dig-protease stability was investigated over broad range of pH, temperature, salt concentrations, surfactants and metal ions. The purified P.dig-protease was used for the production of bioactive peptides. Red scorpionfish (Scorpaena notata) muscle was hydrolyzed with P.dig-protease in order to obtain peptides with biological activities. Interestingly, the hydrolysate revealed the presence of antioxidant and angiotensin-I converting enzyme inhibitor peptides.
authors.affiliation: Laboratory of Protein Engineering and Bioactive Molecules (LIP-MB), National Institute of Applied Sciences and Technology, University of Carthage, Tunis, Cedex, Tunisia.
authors.forename: Neyssene
authors.initials: N
authors.lastname: Aissaoui
authors.forename: Ferid
authors.initials: F
authors.lastname: Abidi
authors.forename: Safa
authors.initials: S
authors.lastname: Mahat
authors.forename: M Nejib
authors.initials: MN
authors.lastname: Marzouki
mesh_terms: D000818:Animals; D011232:Chemical Precipitation; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008670:Metals / Q000378:metabolism; D008970:Molecular Weight; D009124:Muscle Proteins / Q000378:metabolism; D010407:Penicillium / Q000201:enzymology*; D010455:Peptides / Q000378:metabolism*; D010473:Perciformes; D010664:Phenylmethylsulfonyl Fluoride / Q000378:metabolism; D011480:Protease Inhibitors / Q000378:metabolism; D055503:Protein Multimerization; D012492:Salts / Q000378:metabolism; D057057:Serine Proteases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D013501:Surface-Active Agents / Q000378:metabolism; D013696:Temperature
chemical_list: D008670:Metals; D009124:Muscle Proteins; D010455:Peptides; D011480:Protease Inhibitors; D012492:Salts; D013501:Surface-Active Agents; D010664:Phenylmethylsulfonyl Fluoride; D057057:Serine Proteases
keywords: Angiotensin-I converting enzyme inhibitory activity; Antioxidant peptides; Penicillium digitatum; Protease purification; Protein hydrolysate