A mutation in the second intracellular loop of the pituitary adenylate cyclase activating polypeptide type I receptor confers constitutive receptor activation.
The pituitary adenylate cyclase activating polypeptide (PACAP) type I receptor belongs to the glucagon/secretin/vasoactive intestinal polypeptide (VIP) receptor family. We mutated and deleted an amino acid residue (E261) which is located within the second intracellular loop of the rat PACAP type I receptor and which is highly conserved among the receptor family. The wild-type receptor and the mutant receptors were efficiently expressed at the surface of COS-7 cells at nearly the same level and revealed the same high affinity for the agonist PACAP-27. The cAMP contents of COS cells transfected with the E261A, E261Q, and the deletion mutant receptor were 4.6-, 5.7-, and 6.7-fold higher as compared with COS cells transfected with the wild-type receptor. Thus, all the mutant PACAP receptors were constitutively active. The data suggest that the glutamic acid in the second intracellular loop of the PACAP receptor may be a key residue to constrain the receptor in the inactive conformation with respect to its coupling to G(s) proteins.
Cao
Y J
YJ
Institut für Biochemie, Johannes Gutenberg-Universität Mainz, Becherweg 30, D-55099, Mainz, Germany.
Gimpl
G
G
Fahrenholz
F
F
D000262:Adenylyl Cyclases / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D000242:Cyclic AMP / Q000378:metabolism; D018698:Glutamic Acid / Q000235:genetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D051381:Rats; D051236:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide; D051237:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide, Type I; D011979:Receptors, Pituitary Hormone / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D017384:Sequence Deletion; D014162:Transfection
D051236:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide; D051237:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide, Type I; D011979:Receptors, Pituitary Hormone; D018698:Glutamic Acid; D000242:Cyclic AMP; D000262:Adenylyl Cyclases