The 78-kDa
gastrin-
binding protein (GBP) is a likely target for the antiproliferative effects of
gastrin/
cholecystokinin receptor antagonists on
colorectal carcinoma cell lines. Both the N- and C-terminal halves of the GBP bind
gastrin, but the affinity of the N-terminal half for
gastrin is 7.2-fold higher than the affinity of the C-terminal half. In order to define the
gastrin-binding sites of the GBP in greater detail, we have constructed a truncation mutant lacking residues 221-318 of the N-terminal domain and a series of point mutants in which the
lysine residues in the first 220 residues of the N-terminal domain were mutated to
arginine residues. The effect of these mutations on both the extent of covalent cross-linking of iodinated gastrin2,17 and on the affinity for gastrin17 was investigated. Deletion of residues 221-318 of the GBP decreased the affinity 5.5-fold and reduced, but did not abolish, the extent of covalent cross-linking. Mutation of the 17 lysines in residues 1-220 of the GBP decreased the affinity for
gastrin between 1.7- and 3.5-fold and in some cases reduced, but did not abolish, the extent of covalent cross-linking. We conclude that one or more
lysine residues are involved in binding of
gastrin to the GBP, but that no single
lysine residue is the preferred target for covalent cross-linking of iodinated gastrin2,17 to the GBP.