期刊
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
卷 91, 期 -, 页码 198-207出版社
ELSEVIER
DOI: 10.1016/j.ijbiomac.2016.05.034
关键词
Termitomyces clypeatus; Alkaline serine protease; HepG2 cell apoptosis
资金
- Council of Scientific and Industrial Research, New Delhi
Termitomyces clypeatus is an edible mushroom, prized for its therapeutic values and as producer of industrially important enzymes. However, the biomedical efficacies of anticancer proteases have not been reported yet. The present study aimed to purify and characterize a serine protease (AkP) from T. clypeatus for investigating cytotoxic potency on HepG2, Hep3B, and compared the effect on normal hepatic L-02 cells. Purification and biochemical characterization of AkP were evaluated by three stage chromatography, 1D/2D-SDS-PAGE, 1D zymography, far-UV CD spectral analysis, N-terminal sequencing, MALDI TOF/MS-MS analysis and enzyme kinetics studies. AkP could cleave the growth promoting cell surface proteoglycans of HepG2, corroborated by RP-HPLC analysis. AkP (IC50: 75 +/- 1.18 nM) mediated anti-proliferative activity solely on HepG2 cells through the induction of apoptosis. Augmentation of apoptosis was attributed to up-regulation of p53 and Bax protein expression succeeded by caspase-3 activation. Serine protease inhibitor phenyl methane sulfonyl fluoride (PMSF) inhibited both its proteolytic activity and cytotoxicity on HepG2. These findings demonstrate that AkP could be an effective biomolecule for killing of cancer cells by p53 restoration and surface proteoglycans cleavage. (C) 2016 Elsevier B.V. All rights reserved.
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