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Influence of electron transport proteins on the reactions catalyzed by Fusarium fujikuroi gibberellin monooxygenases

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2008-02
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Troncoso, Claudia
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Influence of electron transport proteins on the reactions catalyzed by Fusarium fujikuroi gibberellin monooxygenases
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  • Troncoso, Claudia;
  • Cárcamo, José J.;
  • Hedden, Peter;
  • Tudzynski, Bettina;
  • Rojas, M. Cecilia;
Abstract
The multifunctional cytochrome P450 monooxygenases P450-1 and P450-2 from Fusarium fujikuroi catalyze the formation of GA14 and GA4, respectively, in the gibberellin (GA)-biosynthetic pathway. However, the activity of these enzymes is qualitatively and quantitatively different in mutants lacking the NADPH:cytochrome P450 oxidoreductase (CPR) compared to CPR-containing strains. 3b- Hydroxylation, a major P450-1 activity in wild-type strains, was strongly decreased in the mutants relative to oxidation at C-6 and C-7, while synthesis of C19-GAs as a result of oxidative cleavage of C-20 by P450-2 was almost absent whereas the C-20 alcohol, aldehyde and carboxylic acid derivatives accumulated. Interaction of the monooxygenases with alternative electron transport proteins could account for these different product distributions. In the absence of CPR, P450-1 activities were NADH-dependent, and stimulated by cytochrome b5 or by added FAD. These properties as well as the decreased efficiency of P450-1 and P450-2 in the mutants are consistent with the participation of cytochrome b5:NADH cytochrome b5 reductase as redox partner of the gibberellin monooxygenases in the absence of CPR. We provide evidence, from either incubations of GA12 (C-20 methyl) with cultures of the mutant suspended in [18O]H2O or maintained under an atmosphere of [18O]O2:N2 (20:80), that GA15 (C-20 alcohol) and GA24 (C-20 aldehyde) are formed directly from dioxygen and not from hydrolysis of covalently enzyme-bound intermediates. Thus these partially oxidized GAs correspond to intermediates of the sequential oxidation of C-20 catalyzed by P450-2.
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This work was supported by Fondo Nacional de Ciencia y Tecnologı´a (Grant 1020140) and by the CONICYT/ DAAD Cooperation Program.
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URI: https://repositorio.uchile.cl/handle/2250/118972
ISSN: 0031-9422
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PHYTOCHEMISTRY, Volume: 69,Issue: 3, Pages: 672-683, 2008
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