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Home > Production and Characterization of Extracellular Protease Produced by a Thermophilic Bacterium Strain TLS33
 
Production and Characterization of Extracellular Protease Produced by a Thermophilic Bacterium Strain TLS33
Paper Type
Opinion
Title
Production and Characterization of Extracellular Protease Produced by a Thermophilic Bacterium Strain TLS33
Author
Supachok Sinchaikul, Hataitip Sritanaudomchai and Suree Phutrakul
Email
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Abstract:
Extracellular protease was produced by a thermophilic bacterium strain TLS33 isolated from Thephanom hot spring in Chiang Mai, Thailand. The bacterium was grown and secreted high protease activity when cultured in a medium containing 0.1% w/v yeast extract and 0.25% w/v skim milk in 80% v/v base mixture pH 7.2 at 65oC for 48 h. The protease has been characterized as neutral protease and its optimum pH and temperature were 7.0 and 70-80oC respectively. It was stable up to 75oC after preincubated over a range of temperature for 1 h and its thermostability was assessed at 80oC in the presence of 60 mM CaCl2. Thermostability of the protease was markedly increased by the addition of CaCl2. The protease could hydrolyse a number of substrates including casein, soybean, hemoglobin, BSA, lysozyme, skim milk, ovalbumin and the azocasein (synthetic substrate). It was completely inactivated by 40 mM EDTA which could be classified as a neutral metalloprotease. The inhibition effects of PMSF and thiol reagents suggest that the hydroxy and thiol groups also have a role in enzume integrity. Thus, the metal ion composed in the protease molecule was studied using the holoenzyme, apoenzyme, and Zn2+-apoenzyme preparation. The apoenzyme containing no metal ions had low residual activity (33.12%) and its activity was restored up to 65% both by the addition of 1 mM ZnCl2 in the assay medium and the dialysis of apoenzyme against 0.02 M Tris-HCl buffer pH 7.0 containing 5 mM ZnCl2 which name as Zn2+-apoenzyme. Regarding to stabilization, 1 mM CaCl2 imparted the stability on the apoenzyme and Zn2+-apoenzyme over the 90 min. pre-incubation period at 70oC. The results indicated that Zn2+ enhanced the protease activity whereas the stability of the enzyme was increased by Ca2+. The enzyme could be characterized as a thermostable metalloprotease containing Zn2+ in its molecule.
Start & End Page
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Received Date
1997-12-20
Accepted Date
1998-10-02
Full Text
None
Correspondence:
Author Name
Supachok Sinchaikul - Department of Chemistry, Faculty of Science, Chiang Mai University, Chiang Mai 50200, Thailand.
Hataitip Sritanaudomchai - Department of Chemistry, Faculty of Science, Chiang Mai University, Chiang Mai 50200, Thailand.
Suree Phutrakul - Department of Chemistry, Faculty of Science, Chiang Mai University, Chiang Mai 50200, Thailand.
Keyword:
Keyword
Volume
Vol.25 No.2 (DECEMBER 1998)
 




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