Defining the substrate specificity of mouse cathepsin P

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dc.contributor.authorPuzer, L.
dc.contributor.authorBarros, NMT
dc.contributor.authorOliveira, V
dc.contributor.authorJuliano, M. A.
dc.contributor.authorLu, G. Z.
dc.contributor.authorHassanein, M.
dc.contributor.authorJuliano, L.
dc.contributor.authorMason, R. W.
dc.contributor.authorCarmona, A. K.
dc.contributor.institutionUniversidade Federal de São Paulo (UNIFESP)
dc.contributor.institutionUniv Cidade São Paulo
dc.contributor.institutionAlfred I DuPont Hosp Children
dc.contributor.institutionUniv Delaware
dc.date.accessioned2016-01-24T12:37:42Z
dc.date.available2016-01-24T12:37:42Z
dc.date.issued2005-03-01
dc.description.abstractCathepsin P is a recently discovered placental cysteine protease that is structurally related to the more ubiquitously expressed, broad-specificity enzyme, cathepsin L. We studied the substrate specificity requirements of recombinant mouse cathepsin P using fluorescence resonance energy transfer (FRET) peptides derived from the lead sequence Abz-KLRSSKQ-EDDnp (Abz, ortho-aminobenzoic acid and EDDnp, N-[2,4-dinitroplieiiyl]ethyleiiediamine). Systematic modifications were introduced resulting in five series of peptides to map the S-3 to S-2' subsites of the enzyme. the results indicate that the subsites S-1, S-2, S-1' and S-2', present a clear preference for hydrophobic residues. the specificity requirements of the S, subsite were found to be more restricted, preferring hydrophobic aliphatic amino acids. the S-3 subsite of the enzyme presents a broad specificity, accepting negatively charged (Glu), positively charged (Lys, Arg), and hydrophobic aliphatic or aromatic residues (Val, Phe). for several substrates, the activity of cathepsin P was markedly regulated by kosmotropic salts, particularly Na2SO4. No significant effect on secondary or tertiary structure could be detected by either circular dichroism or size exclusion chromatography, indicating that the salts most probably disrupt unfavorable ionic interactions between the substrate and enzyme active site. A substrate based upon the preferred P-3 to P-2' defined by the screening study, ortho-aminobenzoic-Glu-Ile-Phe-Val-Phe-Lys-Gln-N-(2,4-dinitrophenyl)ethylenediamine (cleaved at the Phe-Val bond) was efficiently hydrolyzed in the absence of high salt. the k(cat)/K-m for this substrate was almost two orders of magnitude higher than that of the original parent compound. These results show that cathepsin P, in contrast to other mammalian cathepsins, has a restricted catalytic specificity. (C) 2004 Elsevier Inc. All rights reserved.en
dc.description.affiliationUNIFESP, Escola Paulista Med, Dept Biophys, BR-04044020 São Paulo, Brazil
dc.description.affiliationUniv Cidade São Paulo, Neurosci Lab, BR-03071000 São Paulo, Brazil
dc.description.affiliationAlfred I DuPont Hosp Children, Dept Biomed Res, Wilmington, de 19803 USA
dc.description.affiliationUniv Delaware, Dept Biol Sci, Newark, de 19716 USA
dc.description.affiliationUnifespUNIFESP, Escola Paulista Med, Dept Biophys, BR-04044020 São Paulo, Brazil
dc.description.sourceWeb of Science
dc.format.extent190-196
dc.identifierhttp://dx.doi.org/10.1016/j.abb.2004.12.007
dc.identifier.citationArchives of Biochemistry and Biophysics. New York: Elsevier B.V., v. 435, n. 1, p. 190-196, 2005.
dc.identifier.doi10.1016/j.abb.2004.12.007
dc.identifier.issn0003-9861
dc.identifier.urihttp://repositorio.unifesp.br/handle/11600/28174
dc.identifier.wosWOS:000226782400021
dc.language.isoeng
dc.publisherElsevier B.V.
dc.relation.ispartofArchives of Biochemistry and Biophysics
dc.rightsinfo:eu-repo/semantics/restrictedAccess
dc.rights.licensehttp://www.elsevier.com/about/open-access/open-access-policies/article-posting-policy
dc.subjectcysteine proteasesen
dc.subjectplacentally expressed cathepsinsen
dc.subjectcathepsin Pen
dc.subjectflurogenic substratesen
dc.subjectsubstrate specificityen
dc.titleDefining the substrate specificity of mouse cathepsin Pen
dc.typeinfo:eu-repo/semantics/article
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