Substrate specificity of an amidating enzyme in porcine pituitary

A series of tripeptides was synthesised and tested as substrates for an amidating enzyme present in porcine pituitary. The rates of conversion of the tripeptides to the corresponding dipeptide amides were determined by ion exchange chromatography of the radio-iodinated peptides. The experiments showed that the amidating enzyme has a mandatory requirement for glycine in position 3 of the tripeptide substrates; peptides containing lysine, glutamic acid, leucine or alanine in the C-terminal position did not undergo reaction. In studies of the substrate requirements at position 2 of the tripeptides, facile reaction took place with neutral amino acids in this position but much slower reactions occurred with basic or acidic residues. With the neutral substrates the enzyme exhibited an optimum pH value of 6.8; with histidine in position 2 the optimum reaction occurred at a higher pH, consistent with a preference shown by the enzyme for an uncharged amino acid in the penultimate position of the peptide substrate.