Mapping of the substrate-binding site of the human granulocyte elastase by the aid of tripeptidyl-p-nitroanilide substrates |
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Authors: | Katalin Marossy Gabriella Cs. Szabó Marianne Pozsgay Pál Elődi |
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Affiliation: | Department of Biochemistry, University Medical School, Debrecen, Hungary |
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Abstract: | The kinetic properties of the human granulocyte elastase /EC 3.4.21.11/ were investigated with 24 tripeptidyl-pNA substrates. By the regression analysis of the kinetic data obtained with 15 substrates a relatively hydrophobic compound, Boc-D-Phe-Ala-Nle-pNA, was predicted as the optimal substrate sequence. The compound was synthesized, assayed and the predicted Km = 4.2 uM was confirmed experimentally. The substrate-binding site of granulocyte elastase appeared to be hydrophobic and very much similar to that of the pancreatic enzyme at the S2–S4 subsites, but the S1 subsite, which determines the primary specificity, could accomodate bulkier residues and it was less selective than that in the pancreatic enzyme. |
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Keywords: | pNA p-nitroanilide Nle norleucine Suc succinyl- Bz benzoyl- Boc tert-butoxycarbonyl- |
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