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Membranolytic Effects of KT2 on Gram-Negative Escherichia coli Evaluated by Atomic Force Microscopy

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Abstract

KT2 is a cationic antimicrobial peptide belonging to Crocodylus siamensis leucrocin I analogs. The mode of action of this compound taken at lethal concentration includes translocation into bacterial cells where binding to DNA is presumed to occur. However, the effects of KT2 on bacterial membrane have not been completely elucidated to date. In this study, a LIVE/DEAD staining technique was used to estimate the appropriate time of peptide-bacteria interaction. The results indicated more than 90% of Escherichia coli population was killed at density of ∼5 × 108 CFU/mL within 30 min after treatment with KT2 at MIC and 10 × MIC. The effects of KT2 on bacterial cells were investigated by the atomic force microscopy (AFM). At near MICs, the peptide induced heavy indentation of the bacterial surface as well as cellular collapse. Conversely, at concentrations of several times the MIC the potential to kill bacteria was greatly increased as judged by the induction of multiple membrane buds on the cell surface. Therefore, the collected results indicate that KT2 can cause different effects on bacterial surface which are positively correlated in magnitude and severity with peptide concentration via membranolytic effects.

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ACKNOWLEDGMENTS

Additionally, we would like to thank the Protein and Proteomics Research Center for Commercial and Industrial Purposes (ProCCI), Department of Biochemistry, Faculty of Science, Khon Kaen University, Thailand as well as the School of Natural System, College of Science and Engineering, Kanazawa University, Kakuma-machi, Japan for providing essential laboratory facilities.

Moreover, we would like to thank Mr. Anuchit Ruangvittayanon and Dr. Worawikunya Kiatponglarp for the AFM assessment, as well as the Synchrotron Light Research Institute (Public Organization), Thailand, for providing access to the AFM equipment.

Funding

This work was financially supported by the Research and Researchers for Industries (RRI), the Thailand Research Fund, and Sriracha Moda Co., Ltd., Chon Buri, Thailand.

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Authors and Affiliations

  1. Protein and Proteomics Research Center for Commercial and Industrial Purposes (ProCCI), Faculty of Science, Khon Kaen University, 40002, Khon Kaen, Thailand

    T. Theansungnoen, N. Jangpromma, P. Anwised, S. Daduang & S. Klaynongsruang

  2. Department of Biochemistry, Faculty of Science, Khon Kaen University, 40002, Khon Kaen, Thailand

    T. Theansungnoen & S. Klaynongsruang

  3. Department of Integrated Science, Forensic Science Program, Faculty of Science, Khon Kaen University, 40002, Khon Kaen, Thailand

    N. Jangpromma

  4. Division of Pharmacognosy and Toxicology, Faculty of Pharmaceutical Sciences, Khon Kaen University, 40002, Khon Kaen, Thailand

    S. Daduang

  5. Faculty of Natural System, Institute of Science and Engineering, Kanazawa University, 920-1192, Kanazawa, Japan

    Y. Fukumori & A. Taoka

  6. Bio-AFM Frontier Research Center, College of Science and Engineering, Kanazawa University, 920-1192, Kanazawa, Japan

    A. Taoka

  7. School of Cosmetic Science, Mae Fah Luang University, 57100, Chiang Rai, Thailand

    T. Theansungnoen

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  1. T. Theansungnoen
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  2. N. Jangpromma
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  7. S. Klaynongsruang
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Correspondence to S. Klaynongsruang.

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The authors declare that they have no conflict of interest. This article does not contain any studies involving animals or human participants performed by any of the authors.

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Theansungnoen, T., Jangpromma, N., Anwised, P. et al. Membranolytic Effects of KT2 on Gram-Negative Escherichia coli Evaluated by Atomic Force Microscopy. Appl Biochem Microbiol 55, 495–505 (2019). https://doi.org/10.1134/S0003683819050144

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