CHOLECYSTOKININ AND GASTRIN ARE NOT EQUALLY SENSITIVE TO GTP-GAMMA-S AT CCKB RECEPTORS - IMPORTANCE OF THE SULFATED TYROSINE

We have shown that gastrin and cholecystokinin octapeptide (CCK-8) are differently coupled to G protein (GTP-binding protein) through type B cholecystokinin receptors in guinea-pig brain membranes and Jurkat ce lls. indeed, the gastrin-13 binding affinity is strongly reduced by st able guanyl nucleotides, whereas CCK-8 binding is only slightly affect ed. In order to determine the structural requirements regulating such coupling, we have synthesized several gastrin and cholecystokinin frag ments (sulphated or unsulphated) elongated at the N-terminus of the co mmon C-terminal tetrapeptide. We investigated their interaction with C CKB receptors in guinea pig brain membranes and Jurkat cells and their involvement in the G protein coupling. Their apparent binding affinit ies to CCKB receptors were measured by inhibition of [I-125]Bolton Hun ter-CCK-8 (3-[I-125]iodo-4-hydroxyphenyl)propionyl- binding in the pre sence or absence of GTP gamma S (guanosine 5'-O-(3-thio)triphosphate) or aluminium tetrafluoride (AlF4-). Activation of the G proteins by GT P gamma S or AlF4- led to a decrease in binding affinity for the gastr in related peptides, the common CCK-gastrin C-terminal forms, the chol ecystokinin hexapeptide and the unsulphated cholecystokinin heptapepti de. Sulphated CCK-7, CCK-8, and cionin apparent binding affinities wer e not affected. These finding indicated that the sulphated tyrosine in position 7 in CCK (as counted from the C-terminus), provides the chol ecystokinin selectivity for the CCKB receptor compared to gastrin. The results are discussed with the aim to better clarify the physiologica l relevance of gastrin and cholecystokinin toward CCKB receptors and t heir related intracellular events.