Specificity of the dynorphin-processing endoprotease: Comparison with prohormone convertases

The cleavage specificity of a monobasic processing dynorphin converting end oprotease is examined with a series of quench fluorescent peptide substrate s and compared with the cleavage specificity of prohormone convertases, A d ynorphin B-29-derived peptide, Abz-Arg-Arg-Gln-Phe-Lys-Val-Val-Thr-Arg-Ser- Gln-eddnp (where Abz is o-aminobenzoyl and eddnp is ethylenediamine 2,4-din itrophenyl), that contains both dibasic and monobasic cleavage sites is eff iciently cleaved by the dynorphin converting enzyme and not cleaved by two propeptide processing enzymes, furin and prohormone convertase 1. A shorter prorenin-related peptide, Dnp-Arg-Met-Ala-Arg-Leu-Thr-Leu-eddnp, that cont ains a monobasic cleavage site is cleaved by the dynorphin converting enzym e and prohormone convertase 1 and not by furin. Substitution of the P1' pos ition by Ala moderately affects cleavage by the dynorphin-processing enzyme and prohormone convertase 1. It is interesting that this substitution resu lts in efficient cleavage by furin. The site of cleavage, as determined by matrix-assisted laser desorption/ionization time of flight mass spectrometr y, is N-terminal to the Arg at the P1 position for the dynorphin converting enzyme and C-terminal to the Arg at the P1 position for furin and prohormo ne convertase 1. Peptides with additional basic residues at the P2 and at P 4 positions also serve as substrates for the dynorphin converting enzyme, T his enzyme cleaves shorter peptide substrates with significantly lower effi ciency as compared with the longer peptide substrates, suggesting that the dynorphin converting enzyme prefers longer peptides that contain monobasic processing sites as substrates, Taken together, these results suggest that the cleavage specificity of the dynorphin converting enzyme is distinct but related to the cleavage specificity of the prohormone convertases and that multiple enzymes could be involved in the processing of peptide hormones a nd neuropeptides at monobasic and dibasic sites.