http://www.cnr.it/ontology/cnr/individuo/prodotto/ID15187

Fluorescent, internally quenched, peptides for exploring the pH-dependent substrate specificity of cathepsin B. (Articolo in rivista)

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Fluorescent, internally quenched, peptides for exploring the pH-dependent substrate specificity of cathepsin B. (Articolo in rivista) (literal)

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Paolo Ruzza; Luigi Quintieri; Alessio Osler; Andrea Calderan; Barbara Biondi; Maura Floreani; Andrea Guiotto; Gianfranco Borin (2006)
Fluorescent, internally quenched, peptides for exploring the pH-dependent substrate specificity of cathepsin B.
in Journal of peptide science (Print)
(literal)

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Paolo Ruzza; Luigi Quintieri; Alessio Osler; Andrea Calderan; Barbara Biondi; Maura Floreani; Andrea Guiotto; Gianfranco Borin (literal)

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Institute of Biomolecular Chemistry of CNR, Padua Unit, 35131 Padua, Italy; Department of Pharmacology and Anesthesiology, Pharmacology Section, University of Padua, 35131 Padua, Italy (literal)

Titolo

Fluorescent, internally quenched, peptides for exploring the pH-dependent substrate specificity of cathepsin B. (literal)

Abstract

Cathepsin B is a cysteine protease that in tumor tissues is localized in both acidic lysosomes and extracellular spaces. It can catalyze the cleavage of peptide bonds by two mechanisms: endoproteolytic attack with a pH optimum around 7.4, and attack from the C-terminus with a pH optimum at 4.5-5.5. In this work, seven fluorescent, internally quenched, decapeptides have been synthesized using the prototypical cathepsin B selective substrate Z-Phe-Arg-AMC as a lead, and used to identify the structural factors determining the susceptibility of peptides to hydrolysis at acidic and neutral pH values. Each peptide differs from the others in one amino acid (residue 6) and contains a highly fluorescent Nma group linked to the ?-amino function of the N-terminal Orn residue and a Dnp group linked to the side chain of the Lys8 residue acting as a quencher. Proteolytic cleavage was monitored by measuring the increase of fluorescence at 440 nm upon excitation at 340 nm, and the cleavage sites were determined by HPLC followed by ESI-MS analysis. Peptides containing Ala or Phe at position 6 are good substrates for the enzyme at both pH 5.0 and 7.4. By contrast, those containing Glu, Asp, Lys or Val are not cleaved at all by cathepsin B at pH 7.4, and are poorly hydrolyzed at pH 5.0. These findings provide new information for the rational design of cathepsin B-activated peptide-containing anticancer drugs. (literal)

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