Human recombinant endopeptidase PHEX has a strict S-1 ' specificity for acidic residues and cleaves peptides derived from fibroblast growth factor-23 and matrix extracellular phosphoglycoprotein

Detalhes bibliográficos
Autor(a) principal: Campos, Marcelo [UNIFESP]
Data de Publicação: 2003
Outros Autores: Couture, Constance, Hirata, Izaura Y. [UNIFESP], Juliano, Maria Aparecida [UNIFESP], Loisel, Thomas P., Crine, Philippe, Juliano, Luiz [UNIFESP], Boileau, Guy, Carmona, Adriana Karaoglanovic [UNIFESP]
Tipo de documento: Artigo
Idioma: eng
Título da fonte: Repositório Institucional da UNIFESP
Texto Completo: http://dx.doi.org/10.1042/BJ20030287
http://repositorio.unifesp.br/handle/11600/27296
Resumo: The PHEX gene (phosphate-regulating gene with homologies to endopeptidases on the X chromosome) encodes a protein (PHEX) with structural homologies to members of the M13 family of zinc metallo-endopeptidases. Mutations in the PHEX gene are responsible for X-linked hypophosphataemia in humans. However, the mechanism by which loss of PHEX function results in the disease phenotype, and the endogenous PHEX substrate(s) remain unknown. in order to study PHEX substrate specificity, combinatorial fluorescent-quenched peptide libraries containing o-aminobenzoic acid (Abz) and 2,4-dinitrophenyl (Dnp) as the donor-acceptor pair were synthesized and tested as PHEX substrates. PHEX showed a strict requirement for acidic amino acid residues (aspartate or glutamate) in S-1' subsite, with a strong preference for aspartate. Subsites S-2', S-1 and S-2 exhibited less defined specificity requirements, but the presence of leucine, proline or glycine in P-2', or valine, isoleucine or histidine in P, precluded hydrolysis of the substrate by the enzyme. the peptide Abz-GFSDYK(Dnp)-OH, which contains the most favourable residues in the P-2 to P-2' positions, was hydrolysed by PHEX at the N-terminus of aspartate with a k(cat)/K-m of 167 mM(-1) . s(-1). in addition, using quenched fluorescence peptides derived from fibroblast growth factor-23 and matrix extracellular phosphoglycoprotein sequences flanked by Abz and N-(2,4-dinitrophenyl)ethylenediamine, we showed that these physiologically relevant proteins are potential PHEX substrates. Finally, our results clearly indicate that PHEX does not have neprilysin-like substrate specificity.
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spelling Human recombinant endopeptidase PHEX has a strict S-1 ' specificity for acidic residues and cleaves peptides derived from fibroblast growth factor-23 and matrix extracellular phosphoglycoproteincombinatorial libraryinternally quenched fluorogenic substrateM13 endopeptidasePHEX substrate specificityThe PHEX gene (phosphate-regulating gene with homologies to endopeptidases on the X chromosome) encodes a protein (PHEX) with structural homologies to members of the M13 family of zinc metallo-endopeptidases. Mutations in the PHEX gene are responsible for X-linked hypophosphataemia in humans. However, the mechanism by which loss of PHEX function results in the disease phenotype, and the endogenous PHEX substrate(s) remain unknown. in order to study PHEX substrate specificity, combinatorial fluorescent-quenched peptide libraries containing o-aminobenzoic acid (Abz) and 2,4-dinitrophenyl (Dnp) as the donor-acceptor pair were synthesized and tested as PHEX substrates. PHEX showed a strict requirement for acidic amino acid residues (aspartate or glutamate) in S-1' subsite, with a strong preference for aspartate. Subsites S-2', S-1 and S-2 exhibited less defined specificity requirements, but the presence of leucine, proline or glycine in P-2', or valine, isoleucine or histidine in P, precluded hydrolysis of the substrate by the enzyme. the peptide Abz-GFSDYK(Dnp)-OH, which contains the most favourable residues in the P-2 to P-2' positions, was hydrolysed by PHEX at the N-terminus of aspartate with a k(cat)/K-m of 167 mM(-1) . s(-1). in addition, using quenched fluorescence peptides derived from fibroblast growth factor-23 and matrix extracellular phosphoglycoprotein sequences flanked by Abz and N-(2,4-dinitrophenyl)ethylenediamine, we showed that these physiologically relevant proteins are potential PHEX substrates. Finally, our results clearly indicate that PHEX does not have neprilysin-like substrate specificity.Universidade Federal de São Paulo, Dept Biophys, Escola Paulista Med, BR-04044020 São Paulo, BrazilUniv Montreal, Dept Biochem, Montreal, PQ H3C 3J7, CanadaBioMep Inc, Montreal, PQ H4B 2L5, CanadaUniversidade Federal de São Paulo, Dept Biophys, Escola Paulista Med, BR-04044020 São Paulo, BrazilWeb of SciencePortland PressUniversidade Federal de São Paulo (UNIFESP)Univ MontrealBioMep IncCampos, Marcelo [UNIFESP]Couture, ConstanceHirata, Izaura Y. [UNIFESP]Juliano, Maria Aparecida [UNIFESP]Loisel, Thomas P.Crine, PhilippeJuliano, Luiz [UNIFESP]Boileau, GuyCarmona, Adriana Karaoglanovic [UNIFESP]2016-01-24T12:33:55Z2016-01-24T12:33:55Z2003-07-01info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersion271-279http://dx.doi.org/10.1042/BJ20030287Biochemical Journal. London: Portland Press, v. 373, p. 271-279, 2003.10.1042/BJ200302870264-6021http://repositorio.unifesp.br/handle/11600/27296WOS:000184060200028engBiochemical Journalinfo:eu-repo/semantics/openAccessreponame:Repositório Institucional da UNIFESPinstname:Universidade Federal de São Paulo (UNIFESP)instacron:UNIFESP2016-01-24T10:33:55Zoai:repositorio.unifesp.br/:11600/27296Repositório InstitucionalPUBhttp://www.repositorio.unifesp.br/oai/requestbiblioteca.csp@unifesp.bropendoar:34652016-01-24T10:33:55Repositório Institucional da UNIFESP - Universidade Federal de São Paulo (UNIFESP)false
dc.title.none.fl_str_mv Human recombinant endopeptidase PHEX has a strict S-1 ' specificity for acidic residues and cleaves peptides derived from fibroblast growth factor-23 and matrix extracellular phosphoglycoprotein
title Human recombinant endopeptidase PHEX has a strict S-1 ' specificity for acidic residues and cleaves peptides derived from fibroblast growth factor-23 and matrix extracellular phosphoglycoprotein
spellingShingle Human recombinant endopeptidase PHEX has a strict S-1 ' specificity for acidic residues and cleaves peptides derived from fibroblast growth factor-23 and matrix extracellular phosphoglycoprotein
Campos, Marcelo [UNIFESP]
combinatorial library
internally quenched fluorogenic substrate
M13 endopeptidase
PHEX substrate specificity
title_short Human recombinant endopeptidase PHEX has a strict S-1 ' specificity for acidic residues and cleaves peptides derived from fibroblast growth factor-23 and matrix extracellular phosphoglycoprotein
title_full Human recombinant endopeptidase PHEX has a strict S-1 ' specificity for acidic residues and cleaves peptides derived from fibroblast growth factor-23 and matrix extracellular phosphoglycoprotein
title_fullStr Human recombinant endopeptidase PHEX has a strict S-1 ' specificity for acidic residues and cleaves peptides derived from fibroblast growth factor-23 and matrix extracellular phosphoglycoprotein
title_full_unstemmed Human recombinant endopeptidase PHEX has a strict S-1 ' specificity for acidic residues and cleaves peptides derived from fibroblast growth factor-23 and matrix extracellular phosphoglycoprotein
title_sort Human recombinant endopeptidase PHEX has a strict S-1 ' specificity for acidic residues and cleaves peptides derived from fibroblast growth factor-23 and matrix extracellular phosphoglycoprotein
author Campos, Marcelo [UNIFESP]
author_facet Campos, Marcelo [UNIFESP]
Couture, Constance
Hirata, Izaura Y. [UNIFESP]
Juliano, Maria Aparecida [UNIFESP]
Loisel, Thomas P.
Crine, Philippe
Juliano, Luiz [UNIFESP]
Boileau, Guy
Carmona, Adriana Karaoglanovic [UNIFESP]
author_role author
author2 Couture, Constance
Hirata, Izaura Y. [UNIFESP]
Juliano, Maria Aparecida [UNIFESP]
Loisel, Thomas P.
Crine, Philippe
Juliano, Luiz [UNIFESP]
Boileau, Guy
Carmona, Adriana Karaoglanovic [UNIFESP]
author2_role author
author
author
author
author
author
author
author
dc.contributor.none.fl_str_mv Universidade Federal de São Paulo (UNIFESP)
Univ Montreal
BioMep Inc
dc.contributor.author.fl_str_mv Campos, Marcelo [UNIFESP]
Couture, Constance
Hirata, Izaura Y. [UNIFESP]
Juliano, Maria Aparecida [UNIFESP]
Loisel, Thomas P.
Crine, Philippe
Juliano, Luiz [UNIFESP]
Boileau, Guy
Carmona, Adriana Karaoglanovic [UNIFESP]
dc.subject.por.fl_str_mv combinatorial library
internally quenched fluorogenic substrate
M13 endopeptidase
PHEX substrate specificity
topic combinatorial library
internally quenched fluorogenic substrate
M13 endopeptidase
PHEX substrate specificity
description The PHEX gene (phosphate-regulating gene with homologies to endopeptidases on the X chromosome) encodes a protein (PHEX) with structural homologies to members of the M13 family of zinc metallo-endopeptidases. Mutations in the PHEX gene are responsible for X-linked hypophosphataemia in humans. However, the mechanism by which loss of PHEX function results in the disease phenotype, and the endogenous PHEX substrate(s) remain unknown. in order to study PHEX substrate specificity, combinatorial fluorescent-quenched peptide libraries containing o-aminobenzoic acid (Abz) and 2,4-dinitrophenyl (Dnp) as the donor-acceptor pair were synthesized and tested as PHEX substrates. PHEX showed a strict requirement for acidic amino acid residues (aspartate or glutamate) in S-1' subsite, with a strong preference for aspartate. Subsites S-2', S-1 and S-2 exhibited less defined specificity requirements, but the presence of leucine, proline or glycine in P-2', or valine, isoleucine or histidine in P, precluded hydrolysis of the substrate by the enzyme. the peptide Abz-GFSDYK(Dnp)-OH, which contains the most favourable residues in the P-2 to P-2' positions, was hydrolysed by PHEX at the N-terminus of aspartate with a k(cat)/K-m of 167 mM(-1) . s(-1). in addition, using quenched fluorescence peptides derived from fibroblast growth factor-23 and matrix extracellular phosphoglycoprotein sequences flanked by Abz and N-(2,4-dinitrophenyl)ethylenediamine, we showed that these physiologically relevant proteins are potential PHEX substrates. Finally, our results clearly indicate that PHEX does not have neprilysin-like substrate specificity.
publishDate 2003
dc.date.none.fl_str_mv 2003-07-01
2016-01-24T12:33:55Z
2016-01-24T12:33:55Z
dc.type.driver.fl_str_mv info:eu-repo/semantics/article
dc.type.status.fl_str_mv info:eu-repo/semantics/publishedVersion
format article
status_str publishedVersion
dc.identifier.uri.fl_str_mv http://dx.doi.org/10.1042/BJ20030287
Biochemical Journal. London: Portland Press, v. 373, p. 271-279, 2003.
10.1042/BJ20030287
0264-6021
http://repositorio.unifesp.br/handle/11600/27296
WOS:000184060200028
url http://dx.doi.org/10.1042/BJ20030287
http://repositorio.unifesp.br/handle/11600/27296
identifier_str_mv Biochemical Journal. London: Portland Press, v. 373, p. 271-279, 2003.
10.1042/BJ20030287
0264-6021
WOS:000184060200028
dc.language.iso.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv Biochemical Journal
dc.rights.driver.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv 271-279
dc.publisher.none.fl_str_mv Portland Press
publisher.none.fl_str_mv Portland Press
dc.source.none.fl_str_mv reponame:Repositório Institucional da UNIFESP
instname:Universidade Federal de São Paulo (UNIFESP)
instacron:UNIFESP
instname_str Universidade Federal de São Paulo (UNIFESP)
instacron_str UNIFESP
institution UNIFESP
reponame_str Repositório Institucional da UNIFESP
collection Repositório Institucional da UNIFESP
repository.name.fl_str_mv Repositório Institucional da UNIFESP - Universidade Federal de São Paulo (UNIFESP)
repository.mail.fl_str_mv biblioteca.csp@unifesp.br
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