Cholecystokinin-associated COOH-terminal peptides are fully sulfated in pig brain

A radioimmunoassay was developed to detect the cholecystokinin (CCK)-associated nonapeptide (CAP-9) that forms the COOH terminus of pig preproCCK. This peptide (Ser-Ala-Glu-Glu-Tyr-Glu-Tyr-Thr-Ser) is presumably produced at the time that the tyrosine-sulfated octapeptide CCK8(s) is cleaved from preproCCK. Radioimmunoassay of a dried methanol extract of pig brain revealed no detectable CAP-9 immunoreactivity, whereas acid desulfation of the dried methanol extract prior to radioimmunoassay resulted in easily measurable concentrations of CAP-9 immunoreactivity. Two peptides, CAP-9 and des-Ser⁹-CAP-9, were purified from a methanol extract of 8 kg of commercially obtained whole pig brains. Amino acid analysis showed that each peptide has both tyrosines sulfated. Thus, the likely sequence of CCK posttranslational processing events is sulfation of the three tyrosines in the COOH terminus of preproCCK followed by peptide cleavage and appearance of CCK8(s) and CAP-9(s,s).