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Mechanism of action of the tri-hybrid antimicrobial peptide LHP7 from lactoferricin, HP and plectasin on Staphylococcus aureus

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Abstract

The tri-hybrid peptide-LHP7 has the potent activity against Gram-positive and Gram-negative as well as fungi, but its mechanism of action has remained elusive. The effluences of LHP7 on the Staphylococcus aureus cell membrane and targets of intracellular action were investigated. LHP7 exhibited an inhibitory effect on the S. aureus growth, similar to those achieved by plectasin, vancomycin and gramicidin. The membrane integrity studies confirmed that LHP7 disrupted the cell membrane, indicating a membrane permeabilizing killing action. A marginal decline in the intensity fluorescence indicated no significant depolarization of the membrane potential following LHP7 treatment. Furthermore, electron microscopy showed that cell shrinkage, cell wall thickening, cellular content leakage, and cell disruption were observed in the cells treated with LHP7. A gel retardation assay showed that LHP7 bound to the genomic DNA of S. aureus or plasmid DNA at a mass ratio of 2.5–10 (peptide/DNA). Circular dichroism indicated that LHP7 inserted into the groove of DNA. The cell cycle analysis showed that after the treatment with LHP7 for 30 and 60 min, the proportion of cells in I-phase increased from 8.71 to 12.09 % and from 8.71 to 15.68 %, indicating that LHP7 induced arrest of cells in the I-phase. These results would conduce to elucidate its underlying antibacterial mechanism.

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Acknowledgments

The authors wish to acknowledge technicians Ms Tong Zhao, Ms Jingnan Liang, Mr Chunli Li and Ms Xiaolan Zhang from the Core Facility at the Institute of Microbiology at the Chinese Academy of Sciences (CAS) for their technical support with the flow cytometric, TEM, SEM and the CLSM analyses. In addition, all other works of this paper was run in Gene Engineering Laboratory, Feed Research Institute, Chinese Academy of Agricultural Sciences. This study was supported by the National Natural Science Foundation of China (No. 31372346 and No. 31302004), the Project of the National Support Program for Science and Technology in China (No. 2013BAD10B02 and No. 2011BAD26B02), the Special Fund for Agro-scientific Research in the Public Interest in China (No. 20140410) and the 2013 AMP Direction of National Innovation Program of Agric Sci & Technol in CAAS.

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Authors and Affiliations

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

    Di Xi, Xiumin Wang, Da Teng, Ruoyu Mao, Yong Zhang, Xiaojie Wang & Jianhua Wang

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

    Di Xi, Xiumin Wang, Da Teng, Ruoyu Mao, Yong Zhang, Xiaojie Wang & Jianhua Wang

  3. Biochemical Engineering Laboratory, School of Life Science, Northeast Agricultural University, Harbin, 150030, China

    Di Xi

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  1. Di Xi
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Correspondence to Jianhua Wang.

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Di Xi and Xiumin Wang authors are contributed equally to this paper.

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Xi, D., Wang, X., Teng, D. et al. Mechanism of action of the tri-hybrid antimicrobial peptide LHP7 from lactoferricin, HP and plectasin on Staphylococcus aureus . Biometals 27, 957–968 (2014). https://doi.org/10.1007/s10534-014-9768-x

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  • DOI: https://doi.org/10.1007/s10534-014-9768-x

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