Characterization of the substrate specificity of the major cysteine protease (cruzipain) from Trypanosoma cruzi using a portion-mixing combinatorial library and fluorogenic peptides
Autor(a) principal: | |
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Data de Publicação: | 1997 |
Outros Autores: | , , , , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Repositório Institucional da UNIFESP |
Texto Completo: | http://repositorio.unifesp.br/11600/42800 http://dx.doi.org/10.1042/bj3230427 |
Resumo: | The substrate specificity of the major cysteinyl proteinase of the parasitic protozoan Trypanosoma cruzi (cruzipain) was investigated, by combinatorial replacement of amino acid residues at positions P-5-P'(5), using a fluorescent quenched solid-phase library assay. Positively charged residues appear to be a general preference in the P-5-P-3 and the P'(5)-P'(3) positions, while a hydrophobic residue was always required at the P-2 position. A broad range of amino acids could be accepted at the P'(1) position. A clear difference in terms of specificity between cruzipain and human cathepsin L was observed for the accommodation of Pro at the P-2 position. The P-1 specificity was investigated by a more detailed enzyme kinetic analysis using peptidyl-MCA (where MCA is methylcoumarin amide) and Abz-peptidyl-EDDnp [where Abz is o-aminobenzoic acid and EDDnp is N-(2,4-dinitrophenyl)ethylenediamine] as substrates, and the results were compared with those obtained using human cathepsin L. Cruzipain showed a clear preference for benzyl-Cys or Arg at the P-1 position. Human cathepsin L presented similar behaviour to that of cruzipain for the hydrolysis of the epsilon-NH2-Cap-Leu-Xaa-MCA (where Cap is epsilon-aminocaproyl) and Abz-Lys-Leu-Xaa-Phe-Ser-Lys-Gln-EDDnp series, whereas the mammalian enzyme was able to tolerate large P-1 residues, such as phenylalanine, better than cruzipain in the latter series. |
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Del Nery, Elaine [UNIFESP]Juliano, Maria Aparecida [UNIFESP]Meldal, MortenSvendsen, IbScharfstein, JulioWalmsley, AdrianJuliano, Luiz [UNIFESP]Universidade Federal de São Paulo (UNIFESP)DEPT CHEMUniversidade Federal do Rio de Janeiro (UFRJ)UNIV SHEFFIELD2018-06-15T14:04:22Z2018-06-15T14:04:22Z1997-04-15Biochemical Journal. London: Portland Press, v. 323, n. 2, p. 427-433, 1997.0264-6021http://repositorio.unifesp.br/11600/42800http://dx.doi.org/10.1042/bj323042710.1042/bj3230427WOS:A1997WV76200017The substrate specificity of the major cysteinyl proteinase of the parasitic protozoan Trypanosoma cruzi (cruzipain) was investigated, by combinatorial replacement of amino acid residues at positions P-5-P'(5), using a fluorescent quenched solid-phase library assay. Positively charged residues appear to be a general preference in the P-5-P-3 and the P'(5)-P'(3) positions, while a hydrophobic residue was always required at the P-2 position. A broad range of amino acids could be accepted at the P'(1) position. A clear difference in terms of specificity between cruzipain and human cathepsin L was observed for the accommodation of Pro at the P-2 position. The P-1 specificity was investigated by a more detailed enzyme kinetic analysis using peptidyl-MCA (where MCA is methylcoumarin amide) and Abz-peptidyl-EDDnp [where Abz is o-aminobenzoic acid and EDDnp is N-(2,4-dinitrophenyl)ethylenediamine] as substrates, and the results were compared with those obtained using human cathepsin L. Cruzipain showed a clear preference for benzyl-Cys or Arg at the P-1 position. Human cathepsin L presented similar behaviour to that of cruzipain for the hydrolysis of the epsilon-NH2-Cap-Leu-Xaa-MCA (where Cap is epsilon-aminocaproyl) and Abz-Lys-Leu-Xaa-Phe-Ser-Lys-Gln-EDDnp series, whereas the mammalian enzyme was able to tolerate large P-1 residues, such as phenylalanine, better than cruzipain in the latter series.ESCOLA PAULISTA MED,DEPT BIOPHYS,BR-04044020 SAO PAULO,BRAZILDEPT CHEM,CARLSBERG LAB,DK-2500 VALBY,DENMARKCCS UNIV FED RIO DE JANEIRO,INST BIOFIS CARLOS CHAGAS FILHO,MOL IMMUNOL LAB,BR-21949 RIO JANEIRO,BRAZILUNIV SHEFFIELD,DEPT MOL BIOL & BIOTECHNOL,SHEFFIELD S10 2UH,S YORKSHIRE,ENGLANDESCOLA PAULISTA MED,DEPT BIOPHYS,BR-04044020 SAO PAULO,BRAZILWeb of Science427-433engPortland PressBiochemical JournalCharacterization of the substrate specificity of the major cysteine protease (cruzipain) from Trypanosoma cruzi using a portion-mixing combinatorial library and fluorogenic peptidesinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleinfo:eu-repo/semantics/openAccessreponame:Repositório Institucional da UNIFESPinstname:Universidade Federal de São Paulo (UNIFESP)instacron:UNIFESP11600/428002021-09-29 11:27:22.265metadata only accessoai:repositorio.unifesp.br:11600/42800Repositório InstitucionalPUBhttp://www.repositorio.unifesp.br/oai/requestopendoar:34652023-05-25T12:25:34.854967Repositório Institucional da UNIFESP - Universidade Federal de São Paulo (UNIFESP)false |
dc.title.en.fl_str_mv |
Characterization of the substrate specificity of the major cysteine protease (cruzipain) from Trypanosoma cruzi using a portion-mixing combinatorial library and fluorogenic peptides |
title |
Characterization of the substrate specificity of the major cysteine protease (cruzipain) from Trypanosoma cruzi using a portion-mixing combinatorial library and fluorogenic peptides |
spellingShingle |
Characterization of the substrate specificity of the major cysteine protease (cruzipain) from Trypanosoma cruzi using a portion-mixing combinatorial library and fluorogenic peptides Del Nery, Elaine [UNIFESP] |
title_short |
Characterization of the substrate specificity of the major cysteine protease (cruzipain) from Trypanosoma cruzi using a portion-mixing combinatorial library and fluorogenic peptides |
title_full |
Characterization of the substrate specificity of the major cysteine protease (cruzipain) from Trypanosoma cruzi using a portion-mixing combinatorial library and fluorogenic peptides |
title_fullStr |
Characterization of the substrate specificity of the major cysteine protease (cruzipain) from Trypanosoma cruzi using a portion-mixing combinatorial library and fluorogenic peptides |
title_full_unstemmed |
Characterization of the substrate specificity of the major cysteine protease (cruzipain) from Trypanosoma cruzi using a portion-mixing combinatorial library and fluorogenic peptides |
title_sort |
Characterization of the substrate specificity of the major cysteine protease (cruzipain) from Trypanosoma cruzi using a portion-mixing combinatorial library and fluorogenic peptides |
author |
Del Nery, Elaine [UNIFESP] |
author_facet |
Del Nery, Elaine [UNIFESP] Juliano, Maria Aparecida [UNIFESP] Meldal, Morten Svendsen, Ib Scharfstein, Julio Walmsley, Adrian Juliano, Luiz [UNIFESP] |
author_role |
author |
author2 |
Juliano, Maria Aparecida [UNIFESP] Meldal, Morten Svendsen, Ib Scharfstein, Julio Walmsley, Adrian Juliano, Luiz [UNIFESP] |
author2_role |
author author author author author author |
dc.contributor.institution.none.fl_str_mv |
Universidade Federal de São Paulo (UNIFESP) DEPT CHEM Universidade Federal do Rio de Janeiro (UFRJ) UNIV SHEFFIELD |
dc.contributor.author.fl_str_mv |
Del Nery, Elaine [UNIFESP] Juliano, Maria Aparecida [UNIFESP] Meldal, Morten Svendsen, Ib Scharfstein, Julio Walmsley, Adrian Juliano, Luiz [UNIFESP] |
description |
The substrate specificity of the major cysteinyl proteinase of the parasitic protozoan Trypanosoma cruzi (cruzipain) was investigated, by combinatorial replacement of amino acid residues at positions P-5-P'(5), using a fluorescent quenched solid-phase library assay. Positively charged residues appear to be a general preference in the P-5-P-3 and the P'(5)-P'(3) positions, while a hydrophobic residue was always required at the P-2 position. A broad range of amino acids could be accepted at the P'(1) position. A clear difference in terms of specificity between cruzipain and human cathepsin L was observed for the accommodation of Pro at the P-2 position. The P-1 specificity was investigated by a more detailed enzyme kinetic analysis using peptidyl-MCA (where MCA is methylcoumarin amide) and Abz-peptidyl-EDDnp [where Abz is o-aminobenzoic acid and EDDnp is N-(2,4-dinitrophenyl)ethylenediamine] as substrates, and the results were compared with those obtained using human cathepsin L. Cruzipain showed a clear preference for benzyl-Cys or Arg at the P-1 position. Human cathepsin L presented similar behaviour to that of cruzipain for the hydrolysis of the epsilon-NH2-Cap-Leu-Xaa-MCA (where Cap is epsilon-aminocaproyl) and Abz-Lys-Leu-Xaa-Phe-Ser-Lys-Gln-EDDnp series, whereas the mammalian enzyme was able to tolerate large P-1 residues, such as phenylalanine, better than cruzipain in the latter series. |
publishDate |
1997 |
dc.date.issued.fl_str_mv |
1997-04-15 |
dc.date.accessioned.fl_str_mv |
2018-06-15T14:04:22Z |
dc.date.available.fl_str_mv |
2018-06-15T14:04:22Z |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/article |
format |
article |
status_str |
publishedVersion |
dc.identifier.citation.fl_str_mv |
Biochemical Journal. London: Portland Press, v. 323, n. 2, p. 427-433, 1997. |
dc.identifier.uri.fl_str_mv |
http://repositorio.unifesp.br/11600/42800 http://dx.doi.org/10.1042/bj3230427 |
dc.identifier.issn.none.fl_str_mv |
0264-6021 |
dc.identifier.doi.none.fl_str_mv |
10.1042/bj3230427 |
dc.identifier.wos.none.fl_str_mv |
WOS:A1997WV76200017 |
identifier_str_mv |
Biochemical Journal. London: Portland Press, v. 323, n. 2, p. 427-433, 1997. 0264-6021 10.1042/bj3230427 WOS:A1997WV76200017 |
url |
http://repositorio.unifesp.br/11600/42800 http://dx.doi.org/10.1042/bj3230427 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.ispartof.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 |
427-433 |
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 |
|
_version_ |
1783460289399750656 |