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Title 

Characterization of arazyme, an exocellular metalloprotease isolated from Serratia proteamaculans culture medium

 

아라자임의 특성

Authors 

P A BersanettiHo Yong ParkKyung Sook BaeKwang Hee SonD H ShinI Y HirataM A JulianoA K CarmonaL Juliano

Publisher 

Elsevier

Issue Date 

2005

Citation 

Enzyme and Microbial Technology, vol. 37, no. 6, pp. 574-581

Keywords 

fluorescent peptidesserralysinserratia proteamaculansarazymebacterium cultureserratia

Abstract 

We investigated the biochemical properties of a 51.5 kDa metalloprotease (arazyme, 3.4.24.40) secreted into the culture medium by Aranicola proteolyticus, a symbiotic bacterium of the spider Nephila clavata. The enzyme was purified to apparent homogeneity by ion exchange chromatography in a Resource Q column (FPLC system). The substrate specificity requirements of purified arazyme were examined using fluorescence resonance energy transfer (FRET) peptides derived from the lead sequence Abz-KLRFSKQ-EDDnp (Abz = ortho-aminobenzoic acid; EDDnp = ethylenediaminedinitrophenyl). Three series of peptides were assayed to map the S2, S1 and S′1 subsites: Abz-KXRFSKQ-EDDnp, Abz-KLXFSKQ-EDDnp and Abz-KLRXSKQ-EDDnp (X are natural amino acids). The results indicated that S 1 subsite has a broad specificity, being Gly the preferred amino acid for this subsite followed by positively charged residues (Arg and His). The S2 and S′1 subsites accommodated better hydrophobic residues with aliphatic or aromatic side chains (Leu, Phe). The pH effect on hydrolysis of Abz-KLFFSKQ-EDDnp indicated that optimal hydrolysis occurred at pH 8.0 or higher. The effect of NaCl on the arazyme activity depends on the substrate, but in general the activity was reduced with this salt. The temperature did not affect the enzyme from 10 to 45°C, after which activity decreased sharply. Arazyme presented high hydrolytic activity on substance P and peptides related to bradykinin. In addition, arazyme activity was resistant to the treatment by pepsin, trypsin and chymotrypsin. In conclusion, arazyme has a broad hydrolytic profile and works in very aggressive conditions, which justify its potential use in therapeutics and biotechnological applications.

ISSN 

0141-0229

Link 

http://dx.doi.org/10.1016/j.enzmictec.2005.01.041

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2017-04-19


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