Specific fluorogenic substrates for neprilysin (neutral endopeptidase, EC 3.4.24.11) which are highly resistant to serine- and metalloproteases

Detalhes bibliográficos
Autor(a) principal: Medeiros, M.a.s.
Data de Publicação: 1997
Outros Autores: França, M.s.f., Boileau, G., Juliano, Luiz [UNIFESP], Carvalho, K.m.
Tipo de documento: Artigo
Idioma: eng
Título da fonte: Repositório Institucional da UNIFESP
Texto Completo: http://dx.doi.org/10.1590/S0100-879X1997001000003
http://repositorio.unifesp.br/handle/11600/538
Resumo: Two intramolecularly quenched fluorogenic peptides containing o-aminobenzoyl (Abz) and ethylenediamine 2,4-dinitrophenyl (EDDnp) groups at amino- and carboxyl-terminal amino acid residues, Abz-<!-- $MVD$:face(Times) -->DArg-Arg-Leu-EDDnp (Abz-<!-- $MVD$:face(Times) -->DRRL-EDDnp) and Abz-<!-- $MVD$:face(Times) -->DArg-Arg-Phe-EDDnp (Abz-<!-- $MVD$:face(Times) -->DRRF-EDDnp), were selectively hydrolyzed by neutral endopeptidase (NEP, enkephalinase, neprilysin, EC 3.4.24.11) at the Arg-Leu and Arg-Phe bonds, respectively. The kinetic parameters for the NEP-catalyzed hydrolysis of Abz-<!-- $MVD$:face(Times) -->DRRL-EDDnp and Abz-<!-- $MVD$:face(Times) -->DRRF-EDDnp were Km = 2.8 µM, kcat = 5.3 min-1, kcat/Km = 2 min-1 µM-1 and Km = 5.0 µM, kcat = 7.0 min-1, kcat/Km = 1.4 min-1 µM-1, respectively. The high specificity of these substrates was demonstrated by their resistance to hydrolysis by metalloproteases [thermolysin (EC 3.4.24.2), angiotensin-converting enzyme (ACE; EC 3.4.24.15)], serineproteases [trypsin (EC 3.4.21.4), <!-- $MVD$:face(Symbol) -->a-chymotrypsin (EC 3.4.21.1)] and proteases present in tissue homogenates from kidney, lung, brain and testis. The blocked amino- and carboxyl-terminal amino acids protected these substrates against the action of aminopeptidases, carboxypeptidases and ACE. Furthermore, <!-- $MVD$:face(Times) -->DR amino acids ensured total protection of Abz-<!-- $MVD$:face(Times) -->DRRL-EDDnp and Abz-<!-- $MVD$:face(Times) -->DRRF-EDDnp against the action of thermolysin and trypsin. Leu-EDDnp and Phe-EDDnp were resistant to hydrolysis by <!-- $MVD$:face(Symbol) -->a-chymotrypsin. The high specifity of these substrates suggests their use for specific NEP assays in crude enzyme preparations
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spelling Specific fluorogenic substrates for neprilysin (neutral endopeptidase, EC 3.4.24.11) which are highly resistant to serine- and metalloproteasesneutral endopeptidaseenkephalinaseneprilysinfluorogenic substratesphosphoramidonTwo intramolecularly quenched fluorogenic peptides containing o-aminobenzoyl (Abz) and ethylenediamine 2,4-dinitrophenyl (EDDnp) groups at amino- and carboxyl-terminal amino acid residues, Abz-<!-- $MVD$:face(Times) -->DArg-Arg-Leu-EDDnp (Abz-<!-- $MVD$:face(Times) -->DRRL-EDDnp) and Abz-<!-- $MVD$:face(Times) -->DArg-Arg-Phe-EDDnp (Abz-<!-- $MVD$:face(Times) -->DRRF-EDDnp), were selectively hydrolyzed by neutral endopeptidase (NEP, enkephalinase, neprilysin, EC 3.4.24.11) at the Arg-Leu and Arg-Phe bonds, respectively. The kinetic parameters for the NEP-catalyzed hydrolysis of Abz-<!-- $MVD$:face(Times) -->DRRL-EDDnp and Abz-<!-- $MVD$:face(Times) -->DRRF-EDDnp were Km = 2.8 µM, kcat = 5.3 min-1, kcat/Km = 2 min-1 µM-1 and Km = 5.0 µM, kcat = 7.0 min-1, kcat/Km = 1.4 min-1 µM-1, respectively. The high specificity of these substrates was demonstrated by their resistance to hydrolysis by metalloproteases [thermolysin (EC 3.4.24.2), angiotensin-converting enzyme (ACE; EC 3.4.24.15)], serineproteases [trypsin (EC 3.4.21.4), <!-- $MVD$:face(Symbol) -->a-chymotrypsin (EC 3.4.21.1)] and proteases present in tissue homogenates from kidney, lung, brain and testis. The blocked amino- and carboxyl-terminal amino acids protected these substrates against the action of aminopeptidases, carboxypeptidases and ACE. Furthermore, <!-- $MVD$:face(Times) -->DR amino acids ensured total protection of Abz-<!-- $MVD$:face(Times) -->DRRL-EDDnp and Abz-<!-- $MVD$:face(Times) -->DRRF-EDDnp against the action of thermolysin and trypsin. Leu-EDDnp and Phe-EDDnp were resistant to hydrolysis by <!-- $MVD$:face(Symbol) -->a-chymotrypsin. The high specifity of these substrates suggests their use for specific NEP assays in crude enzyme preparationsUniversidade Federal do CearáUniversité de MontréalUniversidade Federal de São Paulo (UNIFESP)UNIFESP, EPM, São Paulo, BrazilSciELOAssociação Brasileira de Divulgação CientíficaUniversidade Federal do CearáUniversité de MontréalUniversidade Federal de São Paulo (UNIFESP)Medeiros, M.a.s.França, M.s.f.Boileau, G.Juliano, Luiz [UNIFESP]Carvalho, K.m.2015-06-14T13:24:39Z2015-06-14T13:24:39Z1997-10-01info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersion1157-1162application/pdfhttp://dx.doi.org/10.1590/S0100-879X1997001000003Brazilian Journal of Medical and Biological Research. Associação Brasileira de Divulgação Científica, v. 30, n. 10, p. 1157-1162, 1997.10.1590/S0100-879X1997001000003S0100-879X1997001000003.pdf0100-879XS0100-879X1997001000003http://repositorio.unifesp.br/handle/11600/538WOS:A1997YA76600003engBrazilian Journal of Medical and Biological Researchinfo:eu-repo/semantics/openAccessreponame:Repositório Institucional da UNIFESPinstname:Universidade Federal de São Paulo (UNIFESP)instacron:UNIFESP2024-07-29T16:54:22Zoai:repositorio.unifesp.br/:11600/538Repositório InstitucionalPUBhttp://www.repositorio.unifesp.br/oai/requestbiblioteca.csp@unifesp.bropendoar:34652024-07-29T16:54:22Repositório Institucional da UNIFESP - Universidade Federal de São Paulo (UNIFESP)false
dc.title.none.fl_str_mv Specific fluorogenic substrates for neprilysin (neutral endopeptidase, EC 3.4.24.11) which are highly resistant to serine- and metalloproteases
title Specific fluorogenic substrates for neprilysin (neutral endopeptidase, EC 3.4.24.11) which are highly resistant to serine- and metalloproteases
spellingShingle Specific fluorogenic substrates for neprilysin (neutral endopeptidase, EC 3.4.24.11) which are highly resistant to serine- and metalloproteases
Medeiros, M.a.s.
neutral endopeptidase
enkephalinase
neprilysin
fluorogenic substrates
phosphoramidon
title_short Specific fluorogenic substrates for neprilysin (neutral endopeptidase, EC 3.4.24.11) which are highly resistant to serine- and metalloproteases
title_full Specific fluorogenic substrates for neprilysin (neutral endopeptidase, EC 3.4.24.11) which are highly resistant to serine- and metalloproteases
title_fullStr Specific fluorogenic substrates for neprilysin (neutral endopeptidase, EC 3.4.24.11) which are highly resistant to serine- and metalloproteases
title_full_unstemmed Specific fluorogenic substrates for neprilysin (neutral endopeptidase, EC 3.4.24.11) which are highly resistant to serine- and metalloproteases
title_sort Specific fluorogenic substrates for neprilysin (neutral endopeptidase, EC 3.4.24.11) which are highly resistant to serine- and metalloproteases
author Medeiros, M.a.s.
author_facet Medeiros, M.a.s.
França, M.s.f.
Boileau, G.
Juliano, Luiz [UNIFESP]
Carvalho, K.m.
author_role author
author2 França, M.s.f.
Boileau, G.
Juliano, Luiz [UNIFESP]
Carvalho, K.m.
author2_role author
author
author
author
dc.contributor.none.fl_str_mv Universidade Federal do Ceará
Université de Montréal
Universidade Federal de São Paulo (UNIFESP)
dc.contributor.author.fl_str_mv Medeiros, M.a.s.
França, M.s.f.
Boileau, G.
Juliano, Luiz [UNIFESP]
Carvalho, K.m.
dc.subject.por.fl_str_mv neutral endopeptidase
enkephalinase
neprilysin
fluorogenic substrates
phosphoramidon
topic neutral endopeptidase
enkephalinase
neprilysin
fluorogenic substrates
phosphoramidon
description Two intramolecularly quenched fluorogenic peptides containing o-aminobenzoyl (Abz) and ethylenediamine 2,4-dinitrophenyl (EDDnp) groups at amino- and carboxyl-terminal amino acid residues, Abz-<!-- $MVD$:face(Times) -->DArg-Arg-Leu-EDDnp (Abz-<!-- $MVD$:face(Times) -->DRRL-EDDnp) and Abz-<!-- $MVD$:face(Times) -->DArg-Arg-Phe-EDDnp (Abz-<!-- $MVD$:face(Times) -->DRRF-EDDnp), were selectively hydrolyzed by neutral endopeptidase (NEP, enkephalinase, neprilysin, EC 3.4.24.11) at the Arg-Leu and Arg-Phe bonds, respectively. The kinetic parameters for the NEP-catalyzed hydrolysis of Abz-<!-- $MVD$:face(Times) -->DRRL-EDDnp and Abz-<!-- $MVD$:face(Times) -->DRRF-EDDnp were Km = 2.8 µM, kcat = 5.3 min-1, kcat/Km = 2 min-1 µM-1 and Km = 5.0 µM, kcat = 7.0 min-1, kcat/Km = 1.4 min-1 µM-1, respectively. The high specificity of these substrates was demonstrated by their resistance to hydrolysis by metalloproteases [thermolysin (EC 3.4.24.2), angiotensin-converting enzyme (ACE; EC 3.4.24.15)], serineproteases [trypsin (EC 3.4.21.4), <!-- $MVD$:face(Symbol) -->a-chymotrypsin (EC 3.4.21.1)] and proteases present in tissue homogenates from kidney, lung, brain and testis. The blocked amino- and carboxyl-terminal amino acids protected these substrates against the action of aminopeptidases, carboxypeptidases and ACE. Furthermore, <!-- $MVD$:face(Times) -->DR amino acids ensured total protection of Abz-<!-- $MVD$:face(Times) -->DRRL-EDDnp and Abz-<!-- $MVD$:face(Times) -->DRRF-EDDnp against the action of thermolysin and trypsin. Leu-EDDnp and Phe-EDDnp were resistant to hydrolysis by <!-- $MVD$:face(Symbol) -->a-chymotrypsin. The high specifity of these substrates suggests their use for specific NEP assays in crude enzyme preparations
publishDate 1997
dc.date.none.fl_str_mv 1997-10-01
2015-06-14T13:24:39Z
2015-06-14T13:24:39Z
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/10.1590/S0100-879X1997001000003
Brazilian Journal of Medical and Biological Research. Associação Brasileira de Divulgação Científica, v. 30, n. 10, p. 1157-1162, 1997.
10.1590/S0100-879X1997001000003
S0100-879X1997001000003.pdf
0100-879X
S0100-879X1997001000003
http://repositorio.unifesp.br/handle/11600/538
WOS:A1997YA76600003
url http://dx.doi.org/10.1590/S0100-879X1997001000003
http://repositorio.unifesp.br/handle/11600/538
identifier_str_mv Brazilian Journal of Medical and Biological Research. Associação Brasileira de Divulgação Científica, v. 30, n. 10, p. 1157-1162, 1997.
10.1590/S0100-879X1997001000003
S0100-879X1997001000003.pdf
0100-879X
S0100-879X1997001000003
WOS:A1997YA76600003
dc.language.iso.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv Brazilian Journal of Medical and Biological Research
dc.rights.driver.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv 1157-1162
application/pdf
dc.publisher.none.fl_str_mv Associação Brasileira de Divulgação Científica
publisher.none.fl_str_mv Associação Brasileira de Divulgação Científica
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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