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Concentration dependent effect of GsMTx4 on mechanosensitive channels of small conductance in E. coli spheroplasts

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Abstract

The spider peptide GsMTx4, at saturating concentration of 5 μM, is an effective and specific inhibitor for stretch-activated mechanosensitive (MS) channels found in a variety of eukaryotic cells. Although the structure of the peptide has been solved, the mode of action remains to be determined. Because of its amphipathic structure, the peptide is proposed to interact with lipids at the boundaries of the MS channel proteins. In addition, GsMTx4 has antimicrobial effects, inhibiting growth of several species of bacteria in the range of 5–64 μM. Previous studies on prokaryotic MS channels, which serve as model systems to explore the principle of MS channel gating, have shown that various amphipathic compounds acting at the protein–lipid interface affect MS channel gating. We have therefore analyzed the effect of different concentrations of extracellular GsMTx4 on MS channels of small conductance, MscS and MscK, in the cytoplasmic membrane of wild-type E. coli spheroplasts using the patch-clamp technique. Our study shows that the peptide GsMTx4 exhibits a biphasic response in which peptide concentration determines inhibition or potentiation of activity in prokaryotic MS channels. At low peptide concentrations of 2 and 4 μM the gating of the prokaryotic MS channels was hampered, manifested by a decrease in pressure sensitivity. In contrast, application of peptide at concentrations of 12 and 20 μM facilitated prokaryotic MS channel opening by increasing the pressure sensitivity.

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Abbreviations

MS:

Mechanosensitive channels

WT:

Wild-type

SACs:

Stretch-activated channels

MscL:

Mechanosensitive channel of large conductance

MscS:

Small conductance

MscK:

Small conductance, K+ dependent

MscM:

Mini-conductance

POPG:

Palmitoyloleoylphosphoglycerol

POPC:

Palmitoyloleoylphosphocholine

PE:

Phosphoethanolamine

PG:

Phosphoglycerol

PC:

Pressure cycle

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Acknowledgments

We are grateful to Dr. Frederick Sachs for critical reading of the manuscript and constructive comments. Dr. Åke Wieslander and Dr. Daniel Daley are thanked for sharing important information on their lipid mutant E. coli strains. We like to thank Paul Rohde for technical assistance. The work was supported by a grant from the National Health and Medical Research Council to B.M.

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Author notes

  1. Boris Martinac

    Present address: The Victor Chang Cardiac Research Institute, Lowy Packer Building, 405 Liverpool Street, Darlinghurst, NSW, 2010, Australia

Authors and Affiliations

  1. Molecular Biophysics Laboratory, School of Biomedical Sciences, University of Queensland, St Lucia, QLD, 4072, Australia

    Annette C. Hurst & Boris Martinac

  2. Department of Physiology and Biophysics, SUNY at Buffalo, Buffalo, NY, 14214, USA

    Philip A. Gottlieb

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  1. Annette C. Hurst
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  2. Philip A. Gottlieb
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  3. Boris Martinac
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Correspondence to Boris Martinac.

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Hurst, A.C., Gottlieb, P.A. & Martinac, B. Concentration dependent effect of GsMTx4 on mechanosensitive channels of small conductance in E. coli spheroplasts. Eur Biophys J 38, 415–425 (2009). https://doi.org/10.1007/s00249-008-0386-9

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