A mutation in the second intracellular loop of the pituitary adenylate cyclase activating polypeptide type I receptor confers constitutive receptor activation

FEBS Letters

Fahrenholz, Falk¹  Cao, Yong-Jiang¹  Gimpl, Gerald¹ 

[1] Institut für Biochemie, Johannes Gutenberg-Universität Mainz, Becherweg 30, D-55099 Mainz, Germany

关键词: Pituitary adenylate cyclase activating polypeptide;    Peptide hormone receptor;    Site-directed mutagenesis;    Constitutive activity;    cAMP;    Signal transduction;    DEAE;    diethylaminoethyl;    DMEM;    Dulbecco's modified Eagle's medium;    EC50;    50% effective concentration;    G protein;    guanine nucleotide binding protein;    GPCR;    G protein coupled receptor;    HBSS;    Hanks’ balanced salt solution;    IC50;    median inhibitory effect;    PACAP;    pituitary adenylate cyclase activating polypeptide;    PAC1 receptor;    PACAP type I receptor;    PMSF;    phenylmethylsulfonyl fluoride;    PTH;    parathyroid hormone;    PTHrP;    PTH-related protein;    VIP;    vasoactive intestinal peptide;    VPAC receptor;    VIP-PACAP receptor;   

DOI  :  10.1016/S0014-5793(00)01269-2

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

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.

【 授权许可】

Unknown