title: Purification and biochemical characterization of a detergent-stable keratinase from a newly thermophilic actinomycete Actinomadura keratinilytica strain Cpt29 isolated from poultry compost.
abstract: An extracellular thermostable keratinase (KERAK-29) was purified and biochemically characterized from a thermophilic actinomycete Actinomadura keratinilytica strain Cpt29 newly isolated from Algerian poultry compost. The isolate exhibited high keratinase production when grown in chicken feather meal media (24,000 U/ml). Based on matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF/MS) analysis, the purified enzyme is a monomer with a molecular mass of 29,233.10-Da. The data revealed that the 25 N-terminal residue sequence displayed by KERAK-29 was TQADPPSWGLNNIDRQTAFTKATSI, which showed high homology with those of Streptomyces proteases. This keratinase was completely inhibited by phenylmethanesulfonyl fluoride (PMSF) and diiodopropyl fluorophosphates (DFP), which suggests that it belongs to the serine protease family. Using keratin azure as a substrate, the optimum pH and temperature values for keratinase activity were pH 10 and 70°C, respectively. KERAK-29 was stable between 20 and 60°C and pH 3 and 10 for 5 and 120 h, respectively, and its thermoactivity and thermostability were enhanced in the presence of 5 mM Mn(2+). Its catalytic efficiency was higher than that of the KERAB keratinase from Streptomyces sp. strain AB1. KERAK-29 was also noted to show high keratinolytic activity and significant stability in the presence of detergents, which made it able to accomplish the entire feather-biodegradation process on its own. The ability of the A. keratinilytica strain Cpt29 to grow and produce substantial levels of keratinase using feather as a substrate could open new promising opportunities for the valorization of keratin-containing wastes and reduction of its impacts on the environment.
authors.affiliation: Laboratory of Applied Biochemistry and Microbiology (LABM), Faculty of Science of Annaba (FSA), Badji Mokhtar-Annaba University, P.O. Box 12, 23000 Annaba, Algeria.
authors.forename: Amina
authors.initials: A
authors.lastname: Habbeche
authors.affiliation: Laboratory of Applied Biochemistry and Microbiology (LABM), Faculty of Science of Annaba (FSA), Badji Mokhtar-Annaba University, P.O. Box 12, 23000 Annaba, Algeria.
authors.forename: Boudjema
authors.initials: B
authors.lastname: Saoudi
authors.affiliation: Laboratory of Microorganisms and Biomolecules (LMB), Centre of Biotechnology of Sfax (CBS), University of Sfax, Road of Sidi Mansour Km 6, P.O. Box 1177, Sfax 3018, Tunisia.
authors.forename: Bassem
authors.initials: B
authors.lastname: Jaouadi
authors.affiliation: Laboratory of Applied Biochemistry and Microbiology (LABM), Faculty of Science of Annaba (FSA), Badji Mokhtar-Annaba University, P.O. Box 12, 23000 Annaba, Algeria.
authors.forename: Soumaya
authors.initials: S
authors.lastname: Haberra
authors.affiliation: Laboratory of Applied Biochemistry and Microbiology (LABM), Faculty of Science of Annaba (FSA), Badji Mokhtar-Annaba University, P.O. Box 12, 23000 Annaba, Algeria.
authors.forename: Bilal
authors.initials: B
authors.lastname: Kerouaz
authors.affiliation: Laboratory of Applied Biochemistry and Microbiology (LABM), Faculty of Science of Annaba (FSA), Badji Mokhtar-Annaba University, P.O. Box 12, 23000 Annaba, Algeria.
authors.forename: Mokhtar
authors.initials: M
authors.lastname: Boudelaa
authors.affiliation: Laboratory of Natural Products Chemistry and Biomolecules (LNPCB), University of Saàd Dahlab (USD) of Blida, P.O. Box 270, 09000 Blida, Algeria.
authors.forename: Abdelmalek
authors.initials: A
authors.lastname: Badis
authors.affiliation: Laboratory of Applied Biochemistry and Microbiology (LABM), Faculty of Science of Annaba (FSA), Badji Mokhtar-Annaba University, P.O. Box 12, 23000 Annaba, Algeria. Electronic address: ali.ladjama@univ-annaba.dz.
authors.forename: Ali
authors.initials: A
authors.lastname: Ladjama
mesh_terms: D000192:Actinomycetales / Q000201:enzymology*; D000595:Amino Acid Sequence; D000818:Animals; D003902:Detergents / Q000494:pharmacology; D004795:Enzyme Stability / Q000187:drug effects; D005241:Feathers / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007633:Keratins / Q000378:metabolism; D007700:Kinetics; D008345:Manganese / Q000494:pharmacology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010447:Peptide Hydrolases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D010802:Phylogeny; D011200:Poultry / Q000382:microbiology*; D057057:Serine Proteases / Q000737:chemistry / Q000378:metabolism; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology; D012988:Soil Microbiology; D013302:Streptomyces / Q000201:enzymology; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D003902:Detergents; D015842:Serine Proteinase Inhibitors; D008345:Manganese; D007633:Keratins; D010447:Peptide Hydrolases; D057057:Serine Proteases; C035855:keratinase
keywords: Actinomadura; KERAK-29; Keratin hydrolysis; Keratinase; Protease; Streptomyces