Journal of Applied & Environmental Microbiology
ISSN (Print): 2373-6747 ISSN (Online): 2373-6712 Website: http://www.sciepub.com/journal/jaem Editor-in-chief: Sankar Narayan Sinha
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Journal of Applied & Environmental Microbiology. 2014, 2(3), 74-80
DOI: 10.12691/jaem-2-3-3
Open AccessArticle

Regulation of Operative Biomarkers Production by Treating with Marine Actinomycetes L-Asparaginase in HepG2 Cell Line

Senthil Kumar. M1, , Selvam. K2, Singaravel. R3 and Krishnaveni. N4

1Research and development centre, Bharathiar University, Coimbatore, India

2Department of Botany, Periyar University, Salem, India

3Central Institute of Brackish water Aquaculture (CIBA), Chennai, India

4Department of Microbiology, PSG college of Arts and Science, Coimbatore, India

Pub. Date: March 23, 2014

Cite this paper:
Senthil Kumar. M, Selvam. K, Singaravel. R and Krishnaveni. N. Regulation of Operative Biomarkers Production by Treating with Marine Actinomycetes L-Asparaginase in HepG2 Cell Line. Journal of Applied & Environmental Microbiology. 2014; 2(3):74-80. doi: 10.12691/jaem-2-3-3

Abstract

A unique novel extracelluar glutaminase free L-asparaginase obtained from marine Streptomyces radiopugnans MS1 is an important therapeutic enzyme used in the treatment of Hepatocellular carcinoma. A full-length gene of L-asparaginase was cloned from Streptomyces radiopugnans MS1 and over expressed in Escherichia coli BL21 (DE3). The recombinant L-asparaginase from E.coli was purified by Model 491 Prep Cell which gave 136.8% higher yield with final specific activity of 742.8 IU/mg than the indigenous L-asparaginase from S. radiopugnans MS1. The molecular weight of L-asparaginase was found to be approximately 33.3 kDa by SDS–PAGE analysis. L-asparaginase was tested for its efficacy and cytotoxicity to HepG2 cell line with IC50 0.45 IU/mg and selectivity index (SI) at 10.8 which is about eleven time’s higher selectivity over the lymphocyte cells. HepG2 cells were the most sensitive, showing apoptosis with a higher incidence subsequent to L-asparaginase treatment. A multiplexed flow cytometric bead-based assay to analyze the release of cytokines and quantitative real-time PCR (qRT-PCR) to evaluate gene expression were performed. The obtained cytokine pattern showed that, at the increasing rate of two molecules concentrations, two pro-inflammatory cytokines such as VEGF and IL-8 were decreased whereas the anti-inflammatory cytokine such as IL-4 and IL-10 were increased. This is the first report of the cloning and functional expression of a glutaminase free L-asparaginase gene from noval marine Actinomycetes species. This study has endowed confirmation for the mechanism of L-asparaginase anticancer activity against Hepatocellular carcinoma which was compared with the commercial L-asparaginase. Results indicated that L-asparaginase could be utilized as an effective influential substance for cancer treatment.

Keywords:
glutaminase free L-asparaginase Streptomyces radiopugnans MS1 HepG2 cells and cytokines

Creative CommonsThis work is licensed under a Creative Commons Attribution 4.0 International License. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/

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