Hydrolysis by plasma kallikrein of fluorogenic peptides derived from prorenin processing site

Human plasma kallikrein (HPK) activates plasma prorenin to renin, and the p hysiological significance of this activation is still unknown. In this pape r we investigated the efficiency and the cleavage pattern of the hydrolysis by HPK of the internally quenched fluorescent peptides (qf-peptides) deriv ed from the amino acid sequence of human prorenin cleavage site. The peptid e Abz-F-S-Q-P-M-K-R-L-T-L-G-N-T-T-Q-EDDnp (Abz = ortho-aminobenzoic acid, a nd EDDnp = N-[2,4-dinitrophenyl]-ethylene diamine), that corresponds to the amino acid sequence P-7 to P-7' of human prorenin cleavage site, is hydrol yzed at the correct processing site (R-L bond) with k(cat)/K-m = 85 mM(-1) s(-1). Alanine was scanned in all positions from P-5 to P-5' in order to in vestigate the substrate specificity requirements of HPK. The qf-peptides derived from the equivalent segment of rat prorenin, that h as Lys-Lys as basic amino acid pair, and the peptide Abz-NVTSPVQ-EDDnp that contains the proposed cleavage site of rat prorenin have very low suscepti bility to hydrolysis by rat plasma kallikrein. These data are according to the previously reported absence of rat plasma prorenin activation by rat pl asma kallikrein (RPK), and with the view that prorenin activation in rat re quires alternative enzymes and/or mechanism. All the obtained peptides described in this paper were also assayed with bo vine trypsin that was taken as a reference protease because it is commonly used to activate prorenin. (C) 2000 Elsevier Science B.V. All rights reserv ed.