AJP - Gastrointestinal and Liver Physiology, Vol 252, Issue 2 272-G275, Copyright © 1987 by American Physiological Society
CCK-5: sequence analysis of a small cholecystokinin from canine brain and intestine
J. Shively, J. R. Reeve Jr, V. E. Eysselein, C. Ben-Avram, S. R. Vigna and J. H. Walsh
The purpose of this study is to purify and to characterize chemically
cholecystokinin (CCK)-like peptides present in brain and gut extracts that
elute from gel filtration after the octapeptide. Canine small intestinal
mucosa and brain were boiled in water and then extracted in cold
trifluoroacetic acid, and cholecystokinin-like immunoreactivity was
determined by carboxyl-terminal specific radioimmunoassay. Gel permeation
chromatography on Sephadex G-50 revealed a form of CCK apparently smaller
than CCK-8. This peptide was purified by immunoaffinity chromatography and
three successive reverse phase high-performance liquid chromatography
steps. Microsequence analysis showed that the amino terminal primary
sequence of this small CCK was Gly-Trp-Met-Asp. Immunochemical and
chromatographic analysis indicated that the carboxyl-terminal residue was
Phe-NH2 and thus the full sequence is Gly-Trp-Met-Asp-Phe-NH2. An antibody
that recognizes synthetic CCK-8, CCK-5, and CCK-4 equally did not reveal
the presence of significant amounts of CCK-4. These results indicate that
CCK-5 is the major CCK form smaller than the octapeptide present in brain
(19% of total CCK immunoreactivity) and small intestine (7% of total). This
finding, coupled with the demonstration by others that CCK-5 interacts with
high-affinity brain CCK receptors, indicates that CCK-5 may play a
physiological role in brain function.
