Characterization of a keratinolytic metalloprotease from Bacillus sp. SCB-3.
A keratinolytic protease-producing microorganism was isolated from soybean paste waste and was identified as a strain of Bacillus sp. The keratinase was purified by polyethylene glycol precipitation and two successive column chromatographies with DEAE-Toyopearl 650C and Sephacryl S-200 HR. The purified enzyme had overall 11 purification folds with an 18% yield. The results of sodium dodecyl sulfate polyacrylamide gel electrophoresis and gel filtration on Sephacryl G-200 indicated that the purified enzyme was monomeric and had a molecular weight of 134 kDa. The optimum temperature and pH were 40 degrees C and 7.0, respectively. This enzyme was completely inhibited by EDTA and EGTA, and it was restored by the addition of Ca2+ and Mg2+. These results suggested that it is a metalloprotease. The stimulated enzyme activity by reducing agents indicated that the reducing condition was important in the expression of the activity.
Lee
H
H
Department of Food Science and Biotechnology, Kyonggi University, Suwon, Korea.
Suh
D B
DB
Hwang
J H
JH
Suh
H J
HJ
D001407:Bacillus / Q000201:enzymology*; D002118:Calcium / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007477:Ions; D015603:Keratinocytes / Q000378:metabolism*; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008666:Metalloendopeptidases / Q000737:chemistry* / Q000378:metabolism; D019163:Reducing Agents / Q000494:pharmacology; D013696:Temperature; D013997:Time Factors
D007477:Ions; D019163:Reducing Agents; D008666:Metalloendopeptidases; D008274:Magnesium; D002118:Calcium