Purification and characterization of a novel protease from culture filtrates of a Streptomyces sp.
A fibrinolytic protease has been isolated from Streptomyces sp. culture filtrate by successive chromatography on Mono S and Sephadex G50. The purified protease had a molecular mass of 33 kDa and had an isoelectric point of 6.7. It showed a sharp pH optimum at 7.8 with maximal protease activity between 35 degrees C and 50 degrees C. Its amino acid composition and amino-terminal sequence (17 residues) were determined. The protein exhibited marked hydrolytic activity toward the substrates N-Succ-(Ala)2-Pro-Phe-pNA (K(m) = 0.77 mM, Vmax = 24.2 mumol mg-1 min-1) and N-Succ-(Ala)2-Pro-Leu-pNA (K(m) = 0.92 mM, Vmax = 7.7 mumol mg-1 min-1). It was totally inhibited by alpha 1-antitrypsin, D-Phe-Pro-Arg-chloromethylketone and sodium dodecyl sulfate but was insensitive to EDTA, dithiothreitol, phenylmethylsulfonyl fluoride, soybean trypsin inhibitor, pepstatin or elastatinal. In this respect, this protease differed in its physico-chemical and biochemical properties from other extracellular proteases previously found in bacteria and fungi. The results suggest that it has properties of chymotrypsin-like serine-type proteases.
Bono
F
F
Haemobiology Research Department, Sanofi Recherche, Toulouse, France.
Savi
P
P
Tuong
A
A
Maftouh
M
M
Pereillo
J M
JM
Capdevielle
J
J
Guillemot
J C
JC
Maffrand
J P
JP
Herbert
J M
JM
D000595:Amino Acid Sequence; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D010450:Endopeptidases / Q000737:chemistry* / Q000302:isolation & purification*; D005342:Fibrinolysis; D007526:Isoelectric Point; D008969:Molecular Sequence Data; D011480:Protease Inhibitors / Q000378:metabolism; D013302:Streptomyces / Q000201:enzymology*; D013379:Substrate Specificity
D011480:Protease Inhibitors; D010450:Endopeptidases