Kunitz-type Bauhinia bauhinioides inhibitors devoid of disulfide bridges: isolation of the cDNAs, heterologous expression and structural studies

Kunitz-type Bauhinia bauhinioides inhibitors devoid of disulfide bridges: isolation of the cDNAs, heterologous expression and structural studies

Autor Araujo, APU Google Scholar
Hansen, D. Google Scholar
Vieira, D. F. Google Scholar
Oliveira, C. de Google Scholar
Santana, L. A. Google Scholar
Beltramini, L. M. Google Scholar
Sampaio, CAM Google Scholar
Sampaio, M. U. Google Scholar
Oliva, MLV Google Scholar
Instituição Universidade Federal de São Paulo (UNIFESP)
Universidade de São Paulo (USP)
Resumo Bauhinia bauhinoides cruzipain inhibitor (BbCl) and Bauhinia bauhinioides kallikrein inhibitor (BbKl) are cysteine and serine proteinase inhibitors structurally homologous to plant Kunitz-type inhibitors, but are devoid of disulfide bridges. Based on cDNA sequences, we found that BbKI and BbCI are initially synthesized as a prepropepticle comprising an N-terminal signal peptide (19 residues), the mature protein (164 residues) and a C-terminal targeting peptide (10 residues). Partial cDNAs encoding the mature enzymes plus N-terminal His-tags and thrombin cleavage sites were expressed in E coli and the soluble proteins were purified by one-step nickel affinity chromatography. After thrombin cleavage, both proteins exhibited potent inhibitory activities toward their cognate proteinases like the wild-type proteins. BbCl inhibits human neutrophil elastase (K-i(app) 5.3 nM), porcine pancreatic elastase (K-i(app) 40 nM), cathepsin G (K-i(app) 160 nM) and the cysteine proteinases cruzipain (K-i(app) 1.2 nM), cruzain (K-i(app) 0.3 nM) and cathepsin L (K-i(app) 2.2 nM), while BbKl strongly inhibits plasma kallikrein (K-i(app) 2.4 nM) and plasmin (K-i(app) 33 nM). Circular dichroism spectra of BbCl and BbKl were in agreement with the P-trefoil fold described for Kunitz inhibitors. the inhibitory potency of both BbCl- and BbKl-type inhibitors suggests that other, non-covalent interactions may compensate for the lack of disulficle bridges.
Palavra-chave cathepsins
cruzipain
elastase
gene
kallikreins
proteinase inhibitors
Idioma Inglês
Data de publicação 2005-06-01
Publicado em Biological Chemistry. Berlin: Walter de Gruyter & Co, v. 386, n. 6, p. 561-568, 2005.
ISSN 1431-6730 (Sherpa/Romeo, fator de impacto)
Publicador Walter de Gruyter & Co
Extensão 561-568
Fonte http://dx.doi.org/10.1515/BC.2005.066
Direito de acesso Acesso restrito
Tipo Artigo
Web of Science WOS:000230256800007
Endereço permanente http://repositorio.unifesp.br/handle/11600/28340

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