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Open Access Research article

Biotechnical paving of recombinant enterocin A as the candidate of anti-Listeria agent

Xiaoyuan Hu123, Ruoyu Mao12, Yong Zhang12, Da Teng12, Xiumin Wang12, Di Xi12, Jianzhong Huang3* and Jianhua Wang12*

Author Affiliations

1 Key Laboratory of Feed Biotechnology, Ministry of Agriculture, Beijing 100081, China

2 Gene Engineering Laboratory, Feed Research Institute, Chinese Academy of Agricultural Sciences, 12 Zhongguancun Nandajie St., Haidian District, Beijing 100081, P. R. China

3 College of Life Science, Engineering Research Center of Industrial Microbiology, Fujian Normal University, Fuzhou 350108, China

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BMC Microbiology 2014, 14:220  doi:10.1186/s12866-014-0220-8

Published: 28 August 2014

Abstract

Background

Enterocin A is a classic IIa bacteriocin isolated firstly from Enterococcus faecium CTC492 with selective antimicrobial activity against Listeria strains. However, the application of enterocin A as an anti-Listeria agent has been limited due to its very low native yield. The present work describes high production of enterocin A through codon optimization strategy and its character study.

Results

The gene sequence of enterocin A was optimized based on preferential codon usage in Pichia pastoris to increase its expression efficiency. The highest anti-Listeria activity reached 51,200 AU/ml from 180 mg/l of total protein after 24 h of induction in a 5-L fermenter. Recombinant enterocin A (rEntA), purified by gel filtration chromatography, showed very strong activity against Listeria ivanovii ATCC 19119 with a low MIC of 20 ng/ml. In addition, the rEntA killed over 99% of tested L. ivanovii ATCC19119 within 4 h when exposed to 4?×?MIC (80 ng/ml). Moreover, it showed high stability under a wide pH range (2¿10) and maintained full activity after 1 h of treatment at 80°C within a pH range of 2¿8. Its antimicrobial activity was enhanced at 25 and 50 mM NaCl, while 100¿400 mM NaCl had little effect on the bactericidal ability of rEntA.

Conclusion

The EntA was successfully expressed in P. pastoris, and this feasible system could pave the pre-industrial technological path of rEntA as a competent candidate as an anti-Listeria agent. Furthermore, it showed high stability under wide ranges of conditions, which could be potential as the new candidate of anti-Listeria agent.

Keywords:
Enterocin A; Listeria ivanovii; Antimicrobial activity; Pichia pastoris; Expression