Subcellular Localization, Partial Purification, and Characterization of a Dynorphin Processing Endopeptidase from Bovine Pituitary

Abstract: An enzyme capable of cleaving dynorphin B‐29 to dynorphin B‐13 is present in bovine pituitary, with 40‐ to 50‐fold higher specific activity in the posterior and intermediate lobes than in the anterior lobe. Subcellular fractionation of bovine neurointermediate pituitary shows that this enzyme is present in the peptide‐containing secretory vesicles. The enzyme has been purified 2,800‐fold from whole bovine pituitaries using ion‐exchange and gel filtration chromatography. Purified dynorphin‐converting enzyme has a neutral pH optimum, and is substantially inhibited by the thiol‐protease inhibitor p‐chloromercuriphenylsulfonic acid, but not by serine or metalloprotease inhibitors. The purified enzyme processes dynorphin B‐29 at Arg¹⁴, producing both dynorphin B‐14 and dynorphin B‐13 in a 5:1 ratio. No other cleavages are observed, suggesting that the activity is free from other proteases and is specific for single Arg sequences. Purified enzyme also processes dynorphin A‐17 at the single Arg cleavage site, generating both dynorphin A‐8 and A‐9 in a 7: 1 ratio. The tissue distribution, subcellular localization, and substrate specificity of this enzyme are consistent with a physiological role in the processing of dynorphin B‐29 and dynorphin A‐17, and possibly other peptides, at single Arg residues.