Design of kallidin-releasing tissue kallikrein inhibitors based on the specificities of the enzyme's binding subsites

• Published in: Biochemical journal 1997-04, Vol.323 ( Pt 1) (1), p.167-171
• Main Authors: Portaro, F C, Cezari, M H, Juliano, M A, Juliano, L, Walmsley, A R, Prado, E S
• Format: Article
• Language: English
• Subjects: Benzoylarginine-2-Naphthylamide - analogs & derivatives; Benzoylarginine-2-Naphthylamide - pharmacology; Humans; Kallidin - metabolism; Kallikreins - antagonists & inhibitors; Kinetics; Peptides - chemistry; Peptides - pharmacology; Tissue Kallikreins; Vasoconstrictor Agents - antagonists & inhibitors
• ISSN: 0264-6021; 1470-8728
• DOI: 10.1042/bj3230167

The tissue kallikrein inhibitors reported in the present work were derived by selectively replacing residues in Nalpha-substituted arginine- or phenylalanine-pNA (where pNA is p-nitroanilide), and in peptide substrates for these enzymes. Phenylacetyl-Arg-pNA was found to be an efficient inhibitor of human tissue kallikrein (Ki 0.4 microM) and was neither a substrate nor an inhibitor of plasma kallikrein. The peptide inhibitors having phenylalanine as the P1 residue behaved as specific inhibitors for kallidin-releasing tissue kallikreins, while plasma kallikrein showed high affinity for inhibitors containing (p-nitro)phenylalanine at the same position. The Ki value of the most potent inhibitor developed, Abz-Phe-Arg-Arg-Pro-Arg-EDDnp [where Abz is o-aminobenzoyl and EDDnp is N-(2,4-dinitrophenyl)-ethylenediamine], was 0.08 microM for human tissue kallikrein. Progress curve analyses of the inhibition of human tissue kallikrein by benzoyl-Arg-pNA and phenylacetyl-Phe-Ser-Arg-EDDnp indicated a single-step mechanism for reversible formation of the enzyme-inhibitor complex.