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Type: Artigo de periódico
Title: A new method to purify hepatic CYP1A of Prochilodus scrofa, a Brazilian freshwater fish
Author: Da Silva, MEF
Silva, JA
Marangoni, S
Novello, JC
Meirelles, NC
Abstract: Cytochromes P450 constitute a superfamily of the phase I enzymes whose primary task is the detoxification of both endogenous and xenobiotic compounds. Fish, among non-mammalian species, have received great interest because they are a direct food source for humans as well as conveyors of toxic chemicals to human beings. The aim of the present study was the purification of the hepatic isoform, of CYP1A in Prochilodus scrofa (Prochilodontidae), a Brazilian fish, using only one chromatographic step. The purification of CYP1A was done by Reverse Phase HPLC on a C18 column. Purified CYP1A was characterized with respect to electrophoretic, immunochemical and biocatalyst properties. CYP1A fractions produced a single uniform band on SDS-PAGE with an apparent molecular mass of 58 kDa. Purified CYP1A of P scrofa showed strong cross-reactivity with antibodies directed against CYP1A from trout. The fraction was also encapsulated in two different reconstituted systems; one composed of neutral lipids and another of negatively charged lipids. In both of them, we could detect EROD activity but not PROD activity, which confirms that the CYP1A was purified with all its enzyme activity. There was an increase of activity when CYP1A and NADPH cytochrome P450 (CYP) reductase were encapsulated in negatively charged lipids, which confirms that the charge of lipid is essential to CYP1A activity. All these characteristics strongly suggest that this new procedure is efficient for purifying hepatic CYP1A from R scrofa, showing that the CYP1A isoform of this fish has a highly conserved protein region. (C) 2004 Elsevier Inc. All rights reserved.
Subject: CYP
NADPH cytochrome P450 reductase
Country: EUA
Editor: Elsevier Science Inc
Citation: Comparative Biochemistry And Physiology C-toxicology & Pharmacology. Elsevier Science Inc, v. 138, n. 1, n. 67, n. 74, 2004.
Rights: fechado
Identifier DOI: 10.1016/j.cca.2004.05.004
Date Issue: 2004
Appears in Collections:Unicamp - Artigos e Outros Documentos

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