title: Effects of heme precursors on CYP1A2 and POR expression in the baculovirus/Spodoptera frugiperda system.
abstract: OBJECTIVE
  CYP1A2 and NADPH-CYP450 oxidoreductase (POR) were expressed in the baculovirus/Spodoptera frugiperda (sf9) system. The aim of this study was to investigate the effects of heme precursors on the expression of CYP1A2 and POR.
  
  
  METHODS
  The heme precursors [δ-Aminolaevulinic Acid (5-ALA), Fe(3+) and hemin] were introduced into the system to evaluate their effects on the expression of CYP1A2, POR and their co-expression. All the proteins were identified using immunoblotting, CO-difference spectroscopy, or cytochrome c assay.
  
  
  RESULTS
  In the present study, functional CYP1A2 and POR were successfully expressed in the baculovirus/sf9 system, and both of them showed high activities. Co-addition of 5-ALA and Fe(3+) significantly improved expression of CYP1A2 by about 50% compared with the addition of 5-ALA, Fe(3+) or hemin alone. Either co-addition of 5-ALA and Fe(3+) or addition of 5-ALA or Fe(3+) alone improved the POR expression level 2 fold and its activity 7-10 fold compared with control (no addition). However, unlike CYP1A2, there was no difference between the co-addition and addition of these heme precursors alone. Different ratios of BvCYP1A2 to BvPOR also affected the co-expression of CYP1A2 and POR, with a 3:1 ratio of BvCYP1A2 / BvPOR significantly increasing their co-expression. Surprisingly, the addition of 0.1 mM 5-ALA or Fe(3+) alone, but not their co-addition, could significantly improve the CYP1A2 and POR co-expression (P < 0.05).
  
  
  CONCLUSIONS
  5-ALA and Fe(3+) increased the expression of CYP1A2 and POR in a baculovirus/sf9 system, but the pattern of their expression was different between their expression alone and co-expression.
authors.affiliation: Key Lab of Modern Toxicology of Ministry of Education, School of Public Health, Nanjing Medical University, Nanjing 210029, Jiangsu Province, China.
authors.forename: Huiyuan
authors.initials: H
authors.lastname: Lu
authors.forename: Jun
authors.initials: J
authors.lastname: Ma
authors.forename: Nian
authors.initials: N
authors.lastname: Liu
authors.forename: Shoulin
authors.initials: S
authors.lastname: Wang
keywords: CYP1A2; POR; baculovirus/Spodoptera frugiperda system; co-expression; heme precursors
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