There is a potential phosphorylation site in the C-terminal region of the precursor for the
acid-stimulating
hormone gastrin, which is immediately adjacent to an important cleavage point. In the present study we have sought to identify, separate, quantify and characterize phosphorylated and unphosphorylated forms of human
progastrin and its fragments. Identification was made by two radioimmunoassays: (a) a novel assay employing an antibody raised to intact human
progastrin; and (b) an assay using antibody reacting with the C-terminal tryptic fragment of human
progastrin, as well as
progastrin itself. Two forms of human
progastrin isolated from a
gastrinoma were separated by ion-exchange h.p.l.c., and had similar elution positions on reverse-phase h.p.l.c. and on gel filtration. The more acidic
peptide contained close to equimolar amounts of
phosphate. On trypsinization,
peptides were released that co-eluted on ion-exchange h.p.l.c. with, and had the immunochemical properties of, naturally occurring C-terminal fragments of
progastrin. One of the latter was isolated and shown by Edman degradation after derivatization with
ethanethiol to have the sequence Ser (P)-
Ala-Glu-Asp-Glu-Asn. Similar
peptides occur in
antral mucosa resected from
ulcer patients. The unphosphorylated forms of
progastrin predominated, whereas the phosphorylated forms of the C-terminal fragments were predominant. This distribution could be explained by preferential cleavage of phosphorylated
progastrin. We conclude that in human
progastrin, Ser-96 can occur in the phosphorylated form; this residue immediately follows a pair of basic residues (
Arg-Arg) that are cleaved during synthesis of the biologically active product.