MOLECULAR-CLONING, DEVELOPMENTAL EXPRESSION AND PHARMACOLOGICAL CHARACTERIZATION OF THE CCKB GASTRIN RECEPTOR IN THE CALF PANCREAS

We have cloned the calf predominant pancreatic cholecystokinin B (CCKB )/gastrin receptor cDNA. It encodes a 454 amino acid protein with 90% identity with the CCKB/gastrin receptor cloned in other species and ti ssues. However, the calf pancreatic CCKB/gastrin receptor contains a p entapeptide cassette within the third intracellular loop which is abse nt in the cloned human brain and stomach receptor. Quantification of t he CCKB/gastrin receptor mRNA levels by reverse transcription polymera se chain reaction demonstrated the same level of transcripts at birth, +7 and +28 days. On the other hand, binding study with pancreatic mem branes showing a dramatic increase (600-fold) in the number of CCKB/ga strin receptor sites between at birth and +28 days indicates that the development of the calf pancreatic CCKB/gastrin receptor occurs during the first 4 weeks of post-natal life. COS monkey cells (COS-7 cells) transiently transfected by the cloned cDNA exhibit binding of I-125-Bo lton-Hunter-[Thr(28),Ahx(31)]CCK-(25-33) and I-125-Bolton-Hunter-[Leu( 15)]human gastrin-(2-17) to two affinity classes of sites. K-d values of the high affinity binding components indicate a 4-fold higher affin ity of the receptor for sulfated gastrin than for CCK. Finally, the re combinant receptor is coupled to G proteins and [Ca2+](i) mobilization , and is expressed as a glycoprotein of 82 kDa.