Protocol

Cell-Based Assays for High-Throughput Screening

Volume 486 of the series Methods in Molecular Biology pp 125-137

Date:

Screening for Chemical Inhibitors of Heterologous Proteins Expressed in Yeast Using a Simple Growth-Restoration Assay

  • Aruna D. BalgiAffiliated withDepartment of Biochemistry and Molecular Biology, University of British Columbia
  • , Michel RobergeAffiliated withDepartment of Biochemistry and Molecular Biology, University of British Columbia Email author 

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Summary

Overexpression of heterologous proteins in the yeast Saccharomyces cerevisiae often inhibits its growth, while inhibitors of the overexpressed proteins can restore growth. These simple observations form the basis of a technically easy, inexpensive, scalable, and widely applicable assay to identify inhibitors of such proteins. An expression plasmid for the inducible expression of a gene of interest is introduced into a yeast strain rendered more sensitive to chemicals by deletion of efflux pumps. Protein expression is induced, cells are exposed to test chemicals, and growth is measured by A 600 reading. The chemicals that relieve growth inhibition are subjected to secondary assays to establish their selectivity toward the protein of interest. This assay has been used successfully to identify inhibitors of proteins of viral, microbial, and mammalian origin.

Key words

Chemical inhibitor Drug screening Growth restoration Saccharomyces cerevisiae