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Membrane interaction and antibacterial properties of two mildly cationic peptide diastereomers, bombinins H2 and H4, isolated from Bombina skin

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Abstract

Bombinins H are mildly cationic antimicrobial peptides isolated from the skin of the anuran genus Bombina, the fire-bellied toad. Some members of this peptide family coexist in skin secretions as diastereomers in which a single d-amino acid (alloisoleucine or leucine) is incorporated as a result of the post-translational modification of the respective gene-encoded l-amino acid. Here we report on the antimicrobial properties and membrane interactions of bombinins H2 and H4. The latter differs from H2 by the presence of a d-alloisoleucine at the second N-terminal position. Specifically, we have evaluated the antimicrobial activity of H2 and H4 against a large panel of reference and clinical isolates of Gram-negative and Gram-positive bacteria; performed membrane permeation assays on both intact cells and model membranes (lipid monolayers and liposomes) mimicking the composition of the plasma membrane of Gram-negative/positive bacteria; used biochemical tools, such as trypsin-encapsulated liposomes and capillary electrophoresis, to monitor the peptides’ ability to translocate through the membrane of liposomes mimicking Escherichia coli inner membrane. The results revealed interesting relationships between the presence of a single d-amino acid in the sequence of an antimicrobial peptide and its target microbial cell selectivity/membrane-perturbing activity.

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Fig. 1a, b
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Fig. 3a–d
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Fig. 5a–d

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Abbreviations

AMP:

Antimicrobial peptide

CD:

Circular dichroism

CFU:

Colony-forming units

CL:

Cardiolipin

CZE:

Capillary zone electrophoresis

FITC-D4/D20/D70:

Fluorescein isothiocyanate-dextrans of 4, 20, and 70 kDa average molecular mass, respectively

LPS:

Lipopolysaccharide

MHB:

Mueller-Hinton broth

MIC:

Minimum inhibitory concentration

PBS:

Phosphate-buffered saline

PE:

l-α-phosphatidylethanolamine

PG:

l-α-phosphatidyl-dl-glycerol

SBTI:

Soybean trypsin inhibitor

SDS:

Sodium-dodecyl sulphate

SPB:

Sodium phosphate buffer, pH 7.4 with 0.1 mM EDTA

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Acknowledgments

This work was supported by grants from Università di Roma La Sapienza, MIUR (Italian Ministry of Education, University and Research; PRIN 2008), and Istituto di Biologia e Patologia Molecolari of the Italian National Research Council (CNR).

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Authors and Affiliations

  1. Dipartimento di Scienze e Tecnologie Biomediche, Università de L’Aquila, 67010, L’Aquila, Italy

    Cristina Coccia, Argante Bozzi & Antonio Di Giulio

  2. Istituto Nazionale di Biostrutture e Biosistemi, Rome, Italy

    Argante Bozzi

  3. Dipartimento di Scienze e Tecnologie Biomediche, Università di Cagliari, 09042, Monserrato, Italy

    Andrea C. Rinaldi

  4. Istituto Pasteur-Fondazione Cenci Bolognetti, Dipartimento di Scienze Biochimiche, “A. Rossi Fanelli”, Università La Sapienza, Piazzale Aldo Moro, 5, 00185, Rome, Italy

    Vincenzo Luca, Donatella Barra & Maria Luisa Mangoni

  5. Department of Dental Basic Sciences, Section Oral Biochemistry, Academic Centre for Dentistry Amsterdam, Amsterdam, The Netherlands

    Enno C. I. Veerman

Authors
  1. Cristina Coccia
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  2. Andrea C. Rinaldi
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  3. Vincenzo Luca
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  4. Donatella Barra
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  5. Argante Bozzi
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  6. Antonio Di Giulio
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  7. Enno C. I. Veerman
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  8. Maria Luisa Mangoni
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Corresponding author

Correspondence to Maria Luisa Mangoni.

Additional information

Membrane-active peptides: 455th WE-Heraeus-Seminar and AMP 2010 Workshop.

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Coccia, C., Rinaldi, A.C., Luca, V. et al. Membrane interaction and antibacterial properties of two mildly cationic peptide diastereomers, bombinins H2 and H4, isolated from Bombina skin. Eur Biophys J 40, 577–588 (2011). https://doi.org/10.1007/s00249-011-0681-8

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  • DOI: https://doi.org/10.1007/s00249-011-0681-8

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