Fibrinolytic serine protease isolation from Bacillus amyloliquefaciens An6 grown on Mirabilis jalapa tuber powders.
In this study, Mirabilis jalapa tuber powder (MJTP) was used as a new complex organic substrate for the growth and production of fibrinolytic enzymes by a newly isolated Bacillus amyloliquefaciens An6. Maximum protease activity (1,057 U/ml) with casein as a substrate was obtained when the strain was grown in medium containing (grams per liter) MJTP 30, yeast extract 6, CaCl(2) 1, K(2)HPO(4) 0.1, and K(2)HPO(4) 0.1. The strain was also found to grow and produce extracellular proteases in a medium containing only MJTP, indicating that it can obtain its carbon, nitrogen, and salts requirements directly from MJTP. The B. amyloliquefaciens An6 fibrinase (BAF1) was partially purified, and fibrinolytic activity was assayed in a test tube with an artificial fibrin clot. The molecular weight of the partially purified BAF1 fibrinolytic protease was estimated to be 30 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis and gel filtration. The optimum temperature and pH for the caseinolytic activity were 60 degrees C and 9.0, respectively. The enzyme was highly stable from pH 6.0 to 11.0 and retained 62% of its initial activity after 1 h incubation at 50 degrees C. However, the enzyme was inactivated at higher temperatures. The activity of the enzyme was totally lost in the presence of phenylmethylsulfonyl fluoride, suggesting that BAF1 is a serine protease.
Laboratoire de Génie Enzymatique et de Microbiologie, Ecole Nationale d'Ingénieurs de Sfax, Route de soukra Km 3, 3038 Sfax, Tunisia.
Rym
R
Agrebi
Noomen
N
Hmidet
Mohamed
M
Hajji
Nawrez
N
Ktari
Anissa
A
Haddar
Nahed
N
Fakhfakh-Zouari
Moncef
M
Nasri
D000818:Animals; D001407:Bacillus / Q000201:enzymology* / Q000254:growth & development*; D002417:Cattle; D003470:Culture Media / Q000378:metabolism; D004795:Enzyme Stability; D005342:Fibrinolysis; D006863:Hydrogen-Ion Concentration; D039282:Mirabilis / Q000378:metabolism*; D035281:Plant Tubers / Q000378:metabolism*; D011208:Powders; D057057:Serine Proteases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology; D020860:Subtilisin / Q000302:isolation & purification / Q000378:metabolism; D013696:Temperature
D003470:Culture Media; D011208:Powders; D015842:Serine Proteinase Inhibitors; D057057:Serine Proteases; D020860:Subtilisin