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Listeriolysin O (LLO) is an important extracellular toxin of Listeria monocytogenes which degrades the phospholipid membranes of the host cells’ phagosomes at low pH during the intracellular survival. In contrast to the natural form, the mutant LLO carrying two replacements of amino acid E247M and D320K possesses stable activity at pH 7.4. In this study, we have constructed vectors that carry the mhlyA gene encoding double-mutated LLOE247M/D320K express in B. subtilis 1012. The target gene was fused to the sequence of LysRN-6xHis-TEV site to enhance the recombinant protein expression in B. subtilis and to ease the acquisition of protein by Ni2+-based affinity chromatography. As results, we have obtained the purified protein LLOE247M/D320K with high purity. The activity measurement with 3 HU hemolytic toxins in the pH range from 5.0 to 8.5 suggested that the activity of LLOE247M/D320K was much better than that of natural LLO at neutral pH and slightly alkaline. These results not only provided important scientific foundations for mutant LLO bases expression in B. subtilis but also supplied purified materials for researche and medical applications based on this protein.

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Issue: Vol 19 No 3 (2016)
Page No.: 20-31
Published: Sep 30, 2016
Section: Natural Sciences - Research article

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Copyright: The Authors. This is an open access article distributed under the terms of the Creative Commons Attribution License CC-BY 4.0., which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

 How to Cite
Ngo, H., Nguyen, A., Huynh, P., Nguyen, P., Tran, T., Nguyen, H., & Phan, T. (2016). Expression and purification of listeriolysin O from Listeria monocytogenes harbouring E247M and D320K mutations in Bacillus subtilis. Science and Technology Development Journal, 19(3), 20-31.

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