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Functional similarities between heterogeneously and homogenously expressed MscL constructs

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Abstract

The mechanosensitive channel of large conductance MscL is a well-characterized mechanically gated non-selective ion channel, which often serves as a prototype mechanosensitive channel for mechanotransduction studies. However, there are some discrepancies between MscL constructs used in these studies, most notably unintended heterogeneous expression from some MscL expression constructs. In this study we investigate the possible cause of this expression pattern, and compare the original non-homogenously expressing constructs with our new homogeneously expressing one to confirm that there is little functional difference between them. In addition, a new MscL construct has been developed with an improved molar extinction coefficient at 280 nm, enabling more accurate protein quantification.

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Acknowledgments

N-terminal sequencing was facilitated by access to the Australian Proteome Research Facility, supported under the Australian Government’s National Collaborative Research Infrastructure Strategy (NCRIS), with assistance from Georgina Giannikopoulos and Bernie McInerney. Mass spectrometric analyses were carried out with the assistance of Alun Jones at the Institute for Molecular Bioscience Mass Spectrometry Facility. The authors additionally wish to thank Dr. Jonathan Beesley (QIMR Berghofer Medical Research Institute) for general advice and discussions on cloning. This work was funded in part by the National Health and Medical Research Council of Australia (APP104780).

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Authors and Affiliations

  1. Institute for Molecular Bioscience, The University of Queensland, St Lucia, QLD, 4072, Australia

    Gamma Chi, Ben Hankamer & Michael J. Landsberg

  2. Molecular Cardiology and Biophysics Division, Victor Chang Cardiac Research Institute, Darlinghurst, NSW, 2010, Australia

    Paul R. Rohde, Pietro Ridone & Boris Martinac

  3. St Vincent’s Clinical School, University of New South Wales, Darlinghurst, NSW, 2010, Australia

    Boris Martinac

  4. School of Chemistry and Molecular Biosciences, The University of Queensland, St Lucia, QLD, 4072, Australia

    Michael J. Landsberg

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  1. Gamma Chi
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  2. Paul R. Rohde
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  3. Pietro Ridone
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  4. Ben Hankamer
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  5. Boris Martinac
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  6. Michael J. Landsberg
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Correspondence to Michael J. Landsberg.

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Special Issue: Biophysics of Mechanotransduction.

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Chi, G., Rohde, P.R., Ridone, P. et al. Functional similarities between heterogeneously and homogenously expressed MscL constructs. Eur Biophys J 44, 589–598 (2015). https://doi.org/10.1007/s00249-015-1062-5

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  • DOI: https://doi.org/10.1007/s00249-015-1062-5

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