Functional expression of M(1), M(3) and M(5) muscarinic acetylcholine receptors in yeast

J Neurochem. 2001 Jun;77(5):1327-37. doi: 10.1046/j.1471-4159.2001.00344.x.


The goal of this study was to functionally express the three G(q)-coupled muscarinic receptor subtypes, M(1), M(3) and M(5), in yeast (Saccharomyces cerevisiae). Transformation of yeast with expression constructs coding for the full-length receptors resulted in very low numbers of detectable muscarinic binding sites (B(max) < 5 fmol/mg). Strikingly, deletion of the central portion of the third intracellular loops of the M(1), M(3) and M(5) muscarinic receptors resulted in dramatic increases in B(max) values (53-214 fmol/mg). To monitor productive receptor/G-protein coupling, we used specifically engineered yeast strains that required agonist-stimulated receptor/G-protein coupling for cell growth. These studies showed that the shortened versions of the M(1), M(3) and M(5) receptors were unable to productively interact with the endogenous yeast G protein alpha-subunit, Gpa1p, or a Gpa1 mutant subunit that contained C-terminal mammalian Galpha(s) sequence. In contrast, all three receptors gained the ability to efficiently couple to a Gpa1/Galpha(q) hybrid subunit containing C-terminal mammalian Galpha(q) sequence, indicating that the M(1), M(3) and M(5) muscarinic receptors retained proper G-protein coupling selectivity in yeast. This is the first study to report the expression of muscarinic receptors in a coupling-competent form in yeast. The strategy described here, which involves structural modification of both receptors and co-expressed G proteins, should facilitate the functional expression of other classes of G protein-coupled receptors in yeast.

MeSH terms

  • Amino Acid Sequence
  • Blotting, Western
  • Cell Membrane / metabolism
  • GTP-Binding Proteins / metabolism
  • Gene Expression / genetics
  • Humans
  • Ligands
  • Molecular Sequence Data
  • Plasmids / genetics
  • Radioligand Assay
  • Receptor, Muscarinic M1
  • Receptor, Muscarinic M3
  • Receptor, Muscarinic M5
  • Receptors, Muscarinic / biosynthesis*
  • Receptors, Muscarinic / genetics
  • Saccharomyces cerevisiae / metabolism*


  • Ligands
  • Receptor, Muscarinic M1
  • Receptor, Muscarinic M3
  • Receptor, Muscarinic M5
  • Receptors, Muscarinic
  • GTP-Binding Proteins