Production of adrenergic receptors in yeast

Dorothea Sizmann, Arja Kuusinen, Sirkka Keränen, J. Lomasney, M. Caron

Research output: Contribution to journalArticleScientificpeer-review

24 Citations (Scopus)

Abstract

Using a recombinant yeast strain expressing human beta 2 adrenergic receptors under a galactose-inducible promoter, we established conditions for receptor production in 1-15 liter fermenter culture. Crucial factors contributing to consistent high-level expression included the use of selective glucose-free medium, the maintenance of the pH of the culture at 7.2-7.5 and the presence of an antagonist. The expression strategy and production conditions used with the beta 2 adrenergic receptor were then employed to express the human alpha 2-C2 adrenergic receptor in Saccharomyces cerevisiae. Galactose-induced yeast cells displayed specific, high-affinity [3H]rauwolscine binding and contained a 50-kDa species recognized by an alpha 2-C2 receptor specific antiserum. In fermenter culture, up to 10(5) high-affinity [3H]rauwolscine binding sites per cell (corresponding to 30-60 pmol/mg of protein) were obtained. The high expression level combined with relative ease and low cost of scaling-up make yeast a promising alternative to mammalian cells for the production of adrenergic and other G-protein-coupled receptors for structural studies.
Original languageEnglish
Pages (from-to)197-203
Number of pages7
JournalReceptors and Channels
Volume4
Issue number3
Publication statusPublished - 1996
MoE publication typeA1 Journal article-refereed

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Adrenergic beta-2 Receptors
Adrenergic Receptors
Yohimbine
Yeasts
Galactose
Adrenergic alpha-2 Receptors
G-Protein-Coupled Receptors
Adrenergic Agents
Saccharomyces cerevisiae
Immune Sera
Binding Sites
Maintenance
Costs and Cost Analysis
Glucose
Proteins

Cite this

Sizmann, D., Kuusinen, A., Keränen, S., Lomasney, J., & Caron, M. (1996). Production of adrenergic receptors in yeast. Receptors and Channels, 4(3), 197-203.
Sizmann, Dorothea ; Kuusinen, Arja ; Keränen, Sirkka ; Lomasney, J. ; Caron, M. / Production of adrenergic receptors in yeast. In: Receptors and Channels. 1996 ; Vol. 4, No. 3. pp. 197-203.
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Sizmann, D, Kuusinen, A, Keränen, S, Lomasney, J & Caron, M 1996, 'Production of adrenergic receptors in yeast', Receptors and Channels, vol. 4, no. 3, pp. 197-203.

Production of adrenergic receptors in yeast. / Sizmann, Dorothea; Kuusinen, Arja; Keränen, Sirkka; Lomasney, J.; Caron, M.

In: Receptors and Channels, Vol. 4, No. 3, 1996, p. 197-203.

Research output: Contribution to journalArticleScientificpeer-review

TY - JOUR

T1 - Production of adrenergic receptors in yeast

AU - Sizmann, Dorothea

AU - Kuusinen, Arja

AU - Keränen, Sirkka

AU - Lomasney, J.

AU - Caron, M.

N1 - Project code: BEL2031

PY - 1996

Y1 - 1996

N2 - Using a recombinant yeast strain expressing human beta 2 adrenergic receptors under a galactose-inducible promoter, we established conditions for receptor production in 1-15 liter fermenter culture. Crucial factors contributing to consistent high-level expression included the use of selective glucose-free medium, the maintenance of the pH of the culture at 7.2-7.5 and the presence of an antagonist. The expression strategy and production conditions used with the beta 2 adrenergic receptor were then employed to express the human alpha 2-C2 adrenergic receptor in Saccharomyces cerevisiae. Galactose-induced yeast cells displayed specific, high-affinity [3H]rauwolscine binding and contained a 50-kDa species recognized by an alpha 2-C2 receptor specific antiserum. In fermenter culture, up to 10(5) high-affinity [3H]rauwolscine binding sites per cell (corresponding to 30-60 pmol/mg of protein) were obtained. The high expression level combined with relative ease and low cost of scaling-up make yeast a promising alternative to mammalian cells for the production of adrenergic and other G-protein-coupled receptors for structural studies.

AB - Using a recombinant yeast strain expressing human beta 2 adrenergic receptors under a galactose-inducible promoter, we established conditions for receptor production in 1-15 liter fermenter culture. Crucial factors contributing to consistent high-level expression included the use of selective glucose-free medium, the maintenance of the pH of the culture at 7.2-7.5 and the presence of an antagonist. The expression strategy and production conditions used with the beta 2 adrenergic receptor were then employed to express the human alpha 2-C2 adrenergic receptor in Saccharomyces cerevisiae. Galactose-induced yeast cells displayed specific, high-affinity [3H]rauwolscine binding and contained a 50-kDa species recognized by an alpha 2-C2 receptor specific antiserum. In fermenter culture, up to 10(5) high-affinity [3H]rauwolscine binding sites per cell (corresponding to 30-60 pmol/mg of protein) were obtained. The high expression level combined with relative ease and low cost of scaling-up make yeast a promising alternative to mammalian cells for the production of adrenergic and other G-protein-coupled receptors for structural studies.

M3 - Article

VL - 4

SP - 197

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JO - Receptors and Channels

JF - Receptors and Channels

SN - 1060-6823

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Sizmann D, Kuusinen A, Keränen S, Lomasney J, Caron M. Production of adrenergic receptors in yeast. Receptors and Channels. 1996;4(3):197-203.