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Pakistan Journal of Biological Sciences
  Year: 2016 | Volume: 19 | Issue: 3 | Page No.: 106-114
DOI: 10.3923/pjbs.2016.106.114
 
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Recombinant Engineering of L-Methioninase Using Two Different Promoter and Expression Systems and in vitro Analysis of Its Anticancer Efficacy on Different Human Cancer Cell Lines

M.M. Muharram

Abstract:
Recombinant methioninase (rMETase) is an enzyme that has antitumor activity. In this work, METase gene from Pseudomonas putida ATTCC 8209 was cloned to pT7-7 plasmid (yielded, PT7-METase-R7 clone) and expressed in E. coli strain BL21 (DE3). A protein band with a molecular massof 42 kDa was visualized by SDS-PAGE. The applied protocol yielded a total protein of 3.13 g with a recovery of 66.89% and a specific activity of 18.59 U mg–1 which considered as a low yield. However, when the METase gene was cloned to the vector (pTrc99A, clone: pTrc99A-MET-3) cells of E. coli JM109 yielded a total protein of 32.63 g with a recovery of 41.62% and a specific activity of 54.86 U mg–1 which revealed that the enhancement of METase gene expression by trc promoter was more than the T7 RNA polymerase promoter. The t1/2 of the rMETase was 2 h asanalyzed in mice by IV injection. Antitumor efficacy of rMETase was studied in five human cancer cell lines. At 1 U mL–1 the growth rate of treated colon cancer cell lines, Colo205 and SW620, with rMETase was 46 and 32% relative to control, respectively. With the ovarian cancer cell line (A2780) rMETase produced an inhibition effect of 54% at 1.5 U mL–1. In addition, the growth rate was reduced to 45 and 53% with the skin cancer cell line (A375) and the breast cancer cell line (MCF-7), respectively. These results indicate the feasibility of rMETase for use as a potent antitumor agent.
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How to cite this article:

M.M. Muharram , 2016. Recombinant Engineering of L-Methioninase Using Two Different Promoter and Expression Systems and in vitro Analysis of Its Anticancer Efficacy on Different Human Cancer Cell Lines. Pakistan Journal of Biological Sciences, 19: 106-114.

DOI: 10.3923/pjbs.2016.106.114

URL: https://scialert.net/abstract/?doi=pjbs.2016.106.114

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