Constitutive expression of the human D2S-dopamine receptor in the unicellular yeast Saccharomyces cerevisiae.
The cDNA for the human D2S-dopamine receptor has been functionally expressed in the unicellular yeast Saccharomyces cerevisiae. Two expression plasmids pRS421D2 (original D2S-gene coding region) and pRS421D2S (the first 24 aa of the yeast STE2-gene are fused to the N-terminus of the D2S-gene) were constructed and transformed into the protease deficient S. cerevisiae strain cI3-ABYS-86. Northern blot analysis of total RNA from transformed yeast clones revealed that for both constructs the D2S-gene was constitutively transcribed from the plasmids PMA1 promoter. Membranes prepared from recombinant S. cerevisiae exhibited saturable binding with the antagonist [3H]methylspiperone. Competition studies revealed pharmacological properties for these sites which were comparable to those reported for the D2-receptor heterologously expressed in mammalian cells. The expression of the receptor was monitored by Western blot analysis using an antiserum raised against a peptide from the third intracellular domain of the receptor protein and by ligand binding assay.
Sander
P
P
Max-Planck-Institut für Biophysik, Abt. Molekulare Membranbiologie, Frankfurt/M, Germany.
Grünewald
S
S
Maul
G
G
Reiländer
H
H
Michel
H
H
D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism; D015870:Gene Expression; D008969:Molecular Sequence Data; D017448:Receptors, Dopamine D2 / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D013134:Spiperone / Q000031:analogs & derivatives / Q000378:metabolism
D017448:Receptors, Dopamine D2; D013134:Spiperone; C039167:3-N-methylspiperone