期刊论文详细信息
The Journal of General and Applied Microbiology
Purification and characterization of a novel extracellular neutral metalloprotease from Cerrena albocinnamomea
Keisuke Kubota1  Shigeki Hamada1  Masanobu Sagisaka2 
[1] Faculty of Agriculture and Life Science, Hirosaki University;Faculty of Science and Technology, Hirosaki University
关键词: Cerrena albocinnamomea;    fibrinogenolysis;    metalloprotease;    neutral protease;   
DOI  :  10.2323/jgam.2016.07.006
学科分类:微生物学和免疫学
来源: Applied Microbiology, Molecular and Cellulrar Biosciences Research Foundation
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【 摘 要 】

We selected a fungus secreting a neutral protease from soil and identified it as the basidiomycete fungus Cerrena albocinnamomea according to its ITS-5.8S rDNA and 28S rDNA-D1/D2 sequences. A major extracellular protease isolated from C. albocinnamomea was purified approximately 44-fold through two purification steps. SDS-PAGE analyses of the purified protease revealed a single band, and its molecular mass of 39,756 Da was determined using MALDI-TOF-MS. The enzyme was optimally active at approximately pH 7.0 and 45°C. The Km and Vmax values for the hydrolysis of azocasein were 2.46 mg/mL and 989 units/min/mg protein, respectively. The enzyme was stable at pH 3.6–8.6 for 16 h and at temperatures ≤35°C for 1 h. Enzymatic activity was completely inhibited by Cu2+ and Zn2+ and markedly by EDTA and phosphoramidon. The N-terminal amino acid sequence ASYRVLPIT is highly similar to those of the members of the metalloprotease family M36, such as keratinase and elastinase. However, the protease did not detectably hydrolyze keratin or elastin. In contrast, the protease hydrolyzed fibrinogen, although there were no significant sequence similarities to the N-terminal amino acid sequences of other fibrinolytic enzymes. These results suggest that the purified protease represents a new neutral metalloprotease with fibrinogenolytic activity.

【 授权许可】

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