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Cloning of two additional catecholamine receptors from rat brain
Author(s) -
O'Dowd Brian F.,
Nguyen Tuan,
Tirpak Anne,
Jarvie Keith R.,
Israel Yedy,
Seeman Philip,
Niznik Hyman B.
Publication year - 1990
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(90)80140-e
Subject(s) - biology , microbiology and biotechnology , complementary dna , peptide sequence , molecular cloning , receptor , nucleic acid sequence , sequence analysis , conserved sequence , transmembrane domain , genetics , gene
An approach based on the polymerase chain reaction (PCR) was used to isolate additional members of the G‐linked receptor family from a rat striatal λgtII cDNA library. Priming with one degenerate probe corresponding to highly conserved consensus sequences in the third transmembrane (TM) domain of 15 G‐linked receptors and sequences in the phage vector resulted in one clone (G‐13) encoding a dopamine D2 receptor variant with a 29 amino acid insert in the third cytoplasmic loop. In addition, the amino acid sequence encoded by clone G‐36 contained conserved sequences characteristic of the G‐linked class of receptors and displayed sequence homology in TM domains with the β 2 ‐adrenergic receptor (48%). Two conserved serine residues in TM5 postulated to be part of a ligand binding site in the adrenergic receptor, suggests that G‐36 encodes a catecholaminergic receptor. Northern blot analysis confirmed the expression of G‐36 in rat brain, but not in kidney, heart and lung. Several strong hybridizing bands to G‐36 were obtained in both human and rat genomic DNA. The general PCR strategy employed here should prove to be extremely useful for the isolation of other members of the G‐linked receptor family.