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Biochemical Properties and Mechanism of Action of Enterocin LD3 Purified from Enterococcus hirae LD3

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Abstract

Enterocin LD3 was purified using activity-guided multistep chromatography techniques such as cation-exchange and gel-filtration chromatography. The preparation’s purity was tested using reverse-phase ultra-performance liquid chromatography. The specific activity was tested to be 187.5 AU µg−1 with 13-fold purification. Purified enterocin LD3 was heat stable up to 121 °C (at 15 psi pressure) and pH 2–6. The activity was lost in the presence of papain, reduced by proteinase K, pepsin and trypsin, but was unaffected by amylase and lipase, suggesting proteinaceous nature of the compound and no role of carbohydrate and lipid moieties in the activity. MALDI-TOF/MS analysis of purified enterocin LD3 resolved m/z 4114.6, and N-terminal amino acid sequence was found to be H2NQGGQANQ–COOH suggesting a new bacteriocin. Dissipation of membrane potential, loss of internal ATP and bactericidal effect were recorded when indicator strain Micrococcus luteus was treated with enterocin LD3. It inhibited Gram-positive and Gram-negative bacteria including human pathogens such as Staphylococcus aureus, Pseudomonas fluorescens, Pseudomonas aeruginosa, Salmonella typhi, Shigella flexneri, Listeria monocytogenes, Escherichia coli O157:H7, E. coli (urogenic, a clinical isolate) and Vibrio sp. These properties of purified enterocin LD3 suggest its applications as a food biopreservative and as an alternative to clinical antibiotics.

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Acknowledgments

The present work and AG were financially supported by University Grant Commission, New Delhi, under major research project (No. 36-38/2008 SR). SKT was supported by Indo-US Science and Technology Forum, New Delhi, and was truly acknowledged.

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Authors and Affiliations

  1. Department of Genetics, Maharshi Dayanand University, Rohtak, Haryana, 124001, India

    Aabha Gupta & Santosh Kumar Tiwari

  2. Department of Bioscience and Biotechnology, Banasthali University, Tonk, Rajasthan, 304022, India

    Aabha Gupta & Santosh Kumar Tiwari

  3. Health Promoting Naturals Laboratory, School of Environmental and Biological Sciences, Rutgers State University, New Brunswick, NJ, 08901, USA

    Santosh Kumar Tiwari, Victoria Netrebov & Michael L. Chikindas

  4. Center for Digestive Health, New Jersey Institute for Food, Nutrition and Health, New Brunswick, NJ, 08901, USA

    Michael L. Chikindas

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  1. Aabha Gupta
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Correspondence to Santosh Kumar Tiwari.

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Gupta, A., Tiwari, S.K., Netrebov, V. et al. Biochemical Properties and Mechanism of Action of Enterocin LD3 Purified from Enterococcus hirae LD3. Probiotics & Antimicro. Prot. 8, 161–169 (2016). https://doi.org/10.1007/s12602-016-9217-y

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