Preparation of a verifiable peptide-protein immunogen: direction-controlled conjugation of a synthetic fragment of the monitor peptide with myoglobin and application for sequence analysis.

K Iwai, S Fukuoka, T Fushiki, K Kido, Y Sengoku, T Semba

Index: Anal. Biochem. 171(2) , 277-82, (1988)

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Abstract

A useful method for preparing a synthetic peptide-carrying protein for specific antibody production was established. The monitor peptide is a trypsin-sensitive cholecystokinin-releasing peptide purified from rat pancreatic juice on the basis of its stimulatory activity toward pancreatic enzyme secretion. The NH2-terminus fragment of the monitor peptide (residues 1-14) was synthesized by a solid phase method. Cysteine at the COOH terminus of the fragment was conjugated with amino groups of myoglobin using a hetero-bifunctional reagent. Sequence analysis of the fragment-myoglobin conjugate indicated that the peptide/myoglobin conjugation ratio was about 1/1 (mol/mol). Antiserum against the conjugate from a rabbit effectively abolished the stimulatory activity of the monitor peptide in the rat small intestine.