Thiol-dependent serine alkaline proteases from Bacillus sp. HR-08 and KR-8102: isolation, production, and characterization.
Two Bacillus sp. strains, HR-08 and KR-8102, isolated from soil of the west and north parts of Iran were screened on gelatin agar medium for their ability to produce alkaline protease. The enzymes were active in a wide pH range (6.0-11.0) and stable in the alkaline range (7.0-12.0). The optimum temperatures for the protease from HR-08 and KR-8102 were 65 and 50 degrees C, respectively. The irreversible thermoinactivation of HR-08 and KR-8102 proteases showed that the stability of HR-08 enzyme was higher than that of KR-8102 and the half-lives of these enzymes were 95 and 32 min at 50 degrees C, respectively. In the presence of 10 mM Ca(2+), HR-08 retained 100, 90, and 20% of its initial activity after heating for 30 min at 50, 60, and 70 degrees C, respectively. Enzymes were inhibited by phenylmethylsulfonyl fluoride and iodoacetate. After inhibition by iodoacetate, both enzymes were reactivated by dithiothreitol. These data show that the enzymes seem to be thiol-dependent serine alkaline proteases. The enzymes especially from HR-08 were stable in the presence of H(2)O(2), surfactants, and local detergents; their activities were enhanced in the presence of 5 mM Fe(2+); and the presence of 5 mM metal ions such as Mg(2+), Cu(2+), and Mn(2+) produced almost no effect.
Moradian
Fatemeh
F
Department of Biochemistry, Faculty of Sciences, Tarbiat Modarres University, PO Box 14115-175, Tehran, Iran.
Khajeh
Khosro
K
Naderi-Manesh
Hossein
H
Ahmadvand
Rahim
R
Sajedi
Reza H
RH
Sadeghizadeh
Majid
M
D001407:Bacillus / Q000201:enzymology* / Q000302:isolation & purification; D003902:Detergents; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007492:Iran; D012697:Serine Endopeptidases / Q000302:isolation & purification* / Q000378:metabolism; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology; D012988:Soil Microbiology; D013438:Sulfhydryl Compounds / Q000378:metabolism; D013696:Temperature
D003902:Detergents; D015842:Serine Proteinase Inhibitors; D013438:Sulfhydryl Compounds; D012697:Serine Endopeptidases; C032923:microbial serine proteinases