title: Purification and characterization of a keratinolytic metalloprotease from Chryseobacterium sp. kr6.
abstract: The Chryseobacterium sp. kr6 strain has been described as a highly keratinolytic bacterium showing effective feather-degrading and de-hairing activities. A keratinase Q1 enzyme was purified from Chryseobacterium sp. kr6 culture by Phenyl Sepharose and Superose 12HR chromatography. This enzyme showed a specific activity of 967U/mg for keratin azure. Electrophoresis under denaturing conditions showed a monomeric protein with approximately 64kDa. The enzyme showed pH and temperature optima of 8.5 and 50 degrees C, respectively. The inhibitory effect of EDTA, EGTA and 1,10-phenanthroline characterized Q1 enzyme as a Zn-metalloprotease. Its activity was increased by three-fold in the presence of Ca(2+). ESI-MS/MS analysis of peptides generated from a tryptic digestion revealed sequence homology which may characterize the Q1 keratinase as a member of the M14 metalloprotease family, with a consensus glycosylation region similar to proteins from Chryseobacerium meningosepticum.
authors.affiliation: EMBRAPA (Empresa Brasileira de Pesquisa Agropecuária) Tabuleiros Costeiros - UEP Rio Largo, caixa postal 2013, 57061-970 Maceió/AL, Brazil. ariffel@esalq.usp.br
authors.forename: Alessandro
authors.initials: A
authors.lastname: Riffel
authors.forename: Adriano
authors.initials: A
authors.lastname: Brandelli
authors.forename: Cláudia de M
authors.initials: Cde M
authors.lastname: Bellato
authors.forename: Gustavo H M F
authors.initials: GH
authors.lastname: Souza
authors.forename: Marcos N
authors.initials: MN
authors.lastname: Eberlin
authors.forename: Flavio C A
authors.initials: FC
authors.lastname: Tavares
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D045247:Chryseobacterium / Q000201:enzymology* / Q000235:genetics*; D003001:Cloning, Molecular; D005241:Feathers / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D045726:Metalloproteases / Q000235:genetics / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010447:Peptide Hydrolases / Q000235:genetics / Q000302:isolation & purification*; D011200:Poultry; D017386:Sequence Homology, Amino Acid; D013696:Temperature
chemical_list: D045726:Metalloproteases; D010447:Peptide Hydrolases; C035855:keratinase