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New chromogenic substrates of human neutrophil cathepsin G containing non-natural aromatic amino acid residues in position P1 selected by combinatorial chemistry methods

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Abstract

Specificity of human cathepsin G was explored using combinatorial chemistry methods. Deconvolution of a tetrapeptide library, where 5-amino-2-nitrobenzoic acid served as a chromophore attached at the C-terminus, yielded the active sequence Phe-Val-Thr-Tyr-Anb5,2-NH2. This sequence was used for a second-generation library with the general formula Ac-Phe-Val-Thr-X-Anb5,2-NH2, where position X was replaced with several amino acids: l-pyridyl- alanine (Pal), 4-nitro-l-phenylalanine (Nif), 4-amino-l- phenylalanine (Amf), 4-carboxy-l-phenylalanine (Cbf), 4-guanidine-l-phenylalanine (Gnf), 4-methyloxycarbonyl- l-phenylalanine (Mcf), 4-cyano-l-phenylalanine (Cyf), Phe, Tyr, Arg and Lys. Specificity ligand parameters, k cat and K M, with human cathepsin G were determined for all chromogenic substrates synthesized. The highest value of the specificity constant (k cat/K M) was obtained for a substrate with the Gnf residue in position P1. This peptide was 10 times more active than the second most active substrate which contained the Amf residue. The following order of potency was established: Gnf >  > Amf > Tyr = Phe > Arg= Lys > Cyf. Substrate specificity for cathepsin G is greatly enhanced when an aromatic side chain and a strong positive charge are incorporated in residue P1.

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Authors and Affiliations

  1. Bioorganic Chemistry Department, Faculty of Chemistry, University of Gdańsk, Sobieskiego 18, 80-952, Gdańsk, Poland

    Magdalena Wysocka, Anna Łȩgowska, Elżbieta Bulak, Anna Jaśkiewicz, Hanna Miecznikowska, Adam Lesner & Krzysztf Rolka

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  1. Magdalena Wysocka
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  2. Anna Łȩgowska
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  3. Elżbieta Bulak
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  4. Anna Jaśkiewicz
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  5. Hanna Miecznikowska
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  6. Adam Lesner
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  7. Krzysztf Rolka
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Correspondence to Adam Lesner.

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Wysocka, M., Łȩgowska, A., Bulak, E. et al. New chromogenic substrates of human neutrophil cathepsin G containing non-natural aromatic amino acid residues in position P1 selected by combinatorial chemistry methods. Mol Divers 11, 93–99 (2007). https://doi.org/10.1007/s11030-007-9063-7

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  • DOI: https://doi.org/10.1007/s11030-007-9063-7

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