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Measuring the Activity of Plasma Membrane and Vacuolar Transporters in Yeast

  • Melody Cools
  • Maria Rompf
  • Andreas Mayer
  • Bruno AndréEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 2049)

Abstract

The yeast proteome includes about 300 polytopic membrane proteins known or predicted to function as transporters. Such proteins ensure active or passive transport of small ions or metabolites across the plasma or internal membranes. Despite decades of research on yeast transporters, many of these remain uncharacterized in terms of substrate selectivity range, subcellular localization, and biological function. Assaying the uptake of radiolabeled compounds into whole cells or isolated organelles remains a powerful method for characterizing the function and biochemical properties of these proteins. Here we describe established protocols for measuring transporter activity in whole cells, intact vacuoles, or reconstituted vacuolar vesicles. These methods have proved particularly useful in the context of our work on yeast amino acid transporters, and can in principle be applied to assaying the uptake of other categories of compounds.

Key words

Yeast membrane transporter Yeast vacuole Uptake assay in yeast Uptake assay 

Notes

Acknowledgments

We thank Christos Gournas, Stephan Vissers, and Kathleen Broman for critical reading of the manuscript. M.C. was the recipient of a FRIA PhD fellowship. This work was supported by a PDR grant (nr. 23655065) from the FNRS (Fédération Wallonie-Bruxelles, Belgium) and by a grant from the Cystinosis Research Foundation.

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Melody Cools
    • 1
  • Maria Rompf
    • 2
  • Andreas Mayer
    • 2
  • Bruno André
    • 1
    Email author
  1. 1.Molecular Physiology of the CellUniversité Libre de Bruxelles (ULB)BioparkBelgium
  2. 2.Department of BiochemistryUniversity of Lausanne (UNIL)LausanneSwitzerland

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