Formation of desTyr dynorphins 5-17 by a purified cytosolic aminopeptidase of rat brain.

Abstract

An aminopeptidase purified to homogeneity from cytosol of rat brain cleaved dynorphins having 5-17 residues and selected proenkephalins at the Tyr-Gly bond only to release Tyr and the desTyr fragments. The enzyme protein consisted of a single polypeptide chain of Mr 103,000 and was inhibited by puromycin, bestatin, and chelating reagents to yield Ki in the micromolar range. Hydrolysis of Leu-2-naphthylamide was inhibited by Dyn 1-5 competitively (Ki, 18 microM); the Km for Dyn 1-5, the best substrate of the series, was 63.8 microM (Kcat/Km ratio 580 mM-1 min-1). Rates of N-tyrosyl release decreased with peptide size; the presence of Arg in position 6 led to 50% loss for Dyn 1-6, and the C-terminal extensions of Dyn 1-13 or 1-17 to a 98% loss in activity as compared to the pentapeptide. Rapid degradation of small peptides is consistent with a paracrine (neurotransmitter) role as compared to the postulated precursor or exocrine roles for the dynorphins with 13 residues or more.