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
Autor(a) principal: | |
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Data de Publicação: | 2003 |
Outros Autores: | , , , , , , , |
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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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 |
_version_ |
1814268427931484160 |