Data/hora: 09/06/2024 06:46:38

Provedor de dados: 	56	
País: 	Brazil	
Título: 	Detection of the basement membrane-degrading proteolytic activity of Paracoccidioides brasiliensis after SDS-PAGE using agarose overlays containing Abz-MKALTLQ-EDDnp	
Autores: 	Puccia,R.; Juliano,M.A.; Juliano,L.; Travassos,L.R.; Carmona,A.K.	
Data: 	1999-05-01	
Ano: 	1999	
Palavras-chave: 	P. brasiliensis; Serine-thiol proteinase; SDS-PAGE; Fluorescent-quenched peptides.	
Resumo: 	We have characterized, in the Paracoccidioides brasiliensis yeast phase, an exocellular SH-dependent serine proteinase activity against Abz-MKRLTL-EDDnp and analogous fluorescent-quenched peptides, and showed that it is also active against constituents of the basement membrane in vitro. In the present study, we separated the components of P. brasiliensis culture filtrates by electrophoresis and demonstrated that the serine-thiol exocellular proteinase has a diffuse and heterogeneous migration by SDS-PAGE, localizing in a region between 69 and 43 kDa. The hydrolytic activity was demonstrable after SDS-PAGE using buffered agarose overlays of Abz-MKALTLQ-EDDnp, following incubation at 37oC, and detection of fluorescent bands with a UV transilluminator. Hydrolysis was more intense when incubation was carried out at basic pH, and was completely inhibited with 2.5 mM PMSF and partially with sodium 7-hydroxymercuribenzoate (2.5 mM p-HMB), suggesting its serine-thiol nature. A proteolytic band with similar characteristics was observed in conventional gelatin zymograms, but could not be correlated with a silver-stained component. Detection of the serine-thiol proteinase in substrate gels after SDS-PAGE provides a useful way of monitoring purification of the basement membrane degrading enzyme.	
Tipo: 	Info:eu-repo/semantics/article	
Idioma: 	Inglês	
Identificador: 	http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X1999000500019	
Editor: 	Associação Brasileira de Divulgação Científica	
Relação: 	10.1590/S0100-879X1999000500019	
Formato: 	text/html	
Fonte: 	Brazilian Journal of Medical and Biological Research  v.32 n.5 1999	
Fonte: 	info:eu-repo/semantics/openAccess