| Texto completo | |
| Autor(es): Mostrar menos - |
Oliveira, Lilian C. G.
[1]
;
Silva, Vinicius O.
[1]
;
Okamoto, Debora N.
[1]
;
Kondo, Marcia Y.
[1]
;
Santos, Saara M. B.
[1]
;
Hirata, Isaura Y.
[1]
;
Vallim, Marcelo A.
[1]
;
Pascon, Renata C.
[1]
;
Gouvea, Iuri E.
[1]
;
Juliano, Maria A.
[1]
;
Juliano, Luiz
[1]
Número total de Autores: 11
|
| Afiliação do(s) autor(es): | [1] Univ Fed Sao Paulo, Escola Paulista Med, Dept Biophys, BR-04044020 Sao Paulo - Brazil
Número total de Afiliações: 1
|
| Tipo de documento: | Artigo Científico |
| Fonte: | Analytical Biochemistry; v. 421, n. 1, p. 299-307, FEB 1 2012. |
| Citações Web of Science: | 18 |
| Assunto(s): | Peptídeo hidrolases Enzimas proteolíticas Catepsina L |
| Resumo | |
Identification of synthetic peptide substrates for novel peptidases is an essential step for their study. With this purpose we synthesized fluorescence resonance energy transfer (FRET) peptide libraries Abz (or MCA)-GXXXXXQ-EDDnp and Abs (or MCA)-GXXZXXQ-EDDnp, where X consists of an equimolar mixture of all amino acids, the Z position is fixed with one of the proteinogenic amino acids (cysteine was excluded), Abz (ortho-aminobenzoic acid) or MCA ({[}7-amino-4-methyl]coumarin) is the fluorescence donor and Q-EDDnp (glutamine-{[}N-(2,4-dinitrophenyl)-ethylenediamine]) is the fluorescence acceptor. The peptide libraries MCA-GXXX down arrow XXQ-EDDnp and MCA-GXXZ down arrow XXQ-EDDnp were cleaved as indicated (1) by trypsin, chymotrypsin, cathepsin L, pepsin A, and Eqolisin as confirmed by Edman degradation of the products derived from the digestion of these libraries. The best hydrolyzed Abz-GXXZXXQ-EDDnp sublibraries by these proteases, including Dengue 2 virus NS2B-NS3 protease, contained amino acids at the Z position that are reported to be well accepted by their S-1 subsite. The pH profiles of the hydrolytic activities of these canonical proteases on the libraries were similar to those reported for typical substrates. The FRET peptide libraries provide an efficient and simple approach for detecting nanomolar concentrations of endopeptidases and are useful for initial specificity characterization as performed for two proteases secreted by a Bacillus subtilis. (C) 2011 Elsevier Inc. All rights reserved. (AU) | |
| Processo FAPESP: | 09/52030-5 - Estabelecimento de um laboratório de microbiologia aplicada no Parque Zoológico de São Paulo: identificação e isolamento de microorganismos que produzam enzimas e seus inibidores de aplicação nas áreas médica, veterinária e industrial |
| Beneficiário: | Luiz Juliano Neto |
| Modalidade de apoio: | Auxílio à Pesquisa - Regular |