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Title 

Purification and characterization of two novel fibrinolytic proteases from mushroom, Fomitella fraxinea

Authors 

Jong Suk LeeH S BaikS S Park

Publisher 

The Korean Society for Applied Microbiology

Issue Date 

2006

Citation 

Journal of Microbiology and Biotechnology, vol. 16, no. 2, pp. 264-271

Keywords 

characterizationfibrinolytic proteasefomitella fraxineametalloproteasepurificationserine proteasefomitella fraxinea protease 1fomitella fraxinea protease 2mushroom

Abstract 

Two fibrinolytic enzymes were purified from the culture supernatant of Fomitella fraxinea mycelia by ion-exchange and gel filtration chromatographies, and were designated as F. fraxenia proteases 1 and 2 (FFP1 and FFP2). The apparent molecular masses of the enzymes were estimated to be 32 kDa and 42 kDa, respectively, by SDS-PAGE and gel filtration chromatography. Both enzymes had the same optimal temperature (40°C), but different pH optima (10.0 and 5.0 for FFP1 and FFP2, respectively). FFP1 was relatively stable at pH 7.0-9.0 and temperature below 30°C, whereas FFP2 was very stable in the pH range of 4-11 and temperature below 40°C. FFP1 activity was completely inhibited by phenylmethylsulfonyl fluoride (PMSF) and aprotinin, indicating that this enzyme is a serine protease. The activity of FFP2 was enhanced by the addition of Co2+ and Zn2+ and inhibited by Cu2+, Ni2+, and Hg2+. Furthermore, FFP2 activity was strongly inhibited by EDTA and 1,10-phenanthroline, implying that the enzyme is a metalloprotease. Both enzymes readily hydrolyzed fibrinogen, preferentially digesting the Aα- and Bβ-chains of fibrinogen over γ-chain. FFP1 showed broad substrate specificity for synthetic substrates, but FFP2 did not. Km, and Vmax values of FFP1 for a synthetic substrate, N-succinyl-Ala-Ala-Pro-Phe-pNA, were 0.213 mM and 39.68 units /ml, respectively. The first 15 amino acids of the N-terminal sequences of both enzymes were APXXPXGPWGPQRIS and ARPP(G)VDGQ(R,I)SK(L)ETLPE, respectively.

ISSN 

1017-7825

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2017-04-19


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