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The Fatty Acid 8,11-Diol Synthase of Aspergillus fumigatus is Inhibited by Imidazole Derivatives and Unrelated to PpoB
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
2012 (English)In: Lipids, ISSN 0024-4201, E-ISSN 1558-9307, Vol. 47, no 7, 707-717 p.Article in journal (Refereed) Published
Abstract [en]

(8R)-Hydroperoxy-(9Z,12Z)-octadecadienoic acid (8-HPODE) is formed by aspergilli as an intermediate in biosynthesis of oxylipins with effects on sporulation. 8-HPODE is transformed by separate diol synthases to (5S,8R)-dihydroxy- and (8R,11S)-dihydroxy-(9Z,12Z)-octadecadienoic acids (5,8- and 8,11-DiHODE). The former is formed by the cytochrome P450 (P450) domain of 5,8-linoleate diol synthase (5,8-LDS or PpoA). Our aim was to characterize the 8,11-diol synthase of Aspergillus fumigatus, which is prominent in many strains. The 8,11-diol synthase was soluble and had a larger molecular size (>100 kDa) than most P450. Miconazole, ketoconazole, and 1-benzylimidazole, classical inhibitors of P450, reduced the biosynthesis of 8,11-DiHODE from 8-HPODE (apparent IC50 values similar to 0.8, similar to 5, and similar to 0.6 mu M, respectively), but did not inhibit the biosynthesis of 5,8-DiHODE. Analysis of hydroperoxides of regioisomeric C-18 and C-20 fatty acids showed that the 8,11-diol synthase was specific for certain hydroperoxides with R configuration. The suprafacial hydrogen abstraction and oxygen insertion at C-11 of 8-HPODE was associated with a small deuterium kinetic isotope effect ((H)k(cat)/(D)k(cat) similar to 1.5), consistent with P450-catalyzed oxidation. The genome of A. fumigatus contains over 70 P450 sequences. The reaction mechanism, size, and solubility of 8,11-diol synthase pointed to PpoB, a homologue of 5,8-LDS, as a possible candidate of this activity. Gene deletion of ppoB of A. fumigatus strains AF:Delta ku80 and J272 did not inhibit biosynthesis of 8,11-DiHODE and recombinant PpoB appeared to lack diol synthase activity. We conclude that 8,11-DiHODE is formed from 8-HPODE by a soluble and substrate-specific 8,11-diol synthase with catalytic characteristics of class III P450.

Place, publisher, year, edition, pages
2012. Vol. 47, no 7, 707-717 p.
Keyword [en]
Gene deletion, Sporulation, Animal heme peroxidase, LC-MS/MS, Insect cells, Protein expression
National Category
Medical and Health Sciences
URN: urn:nbn:se:uu:diva-179936DOI: 10.1007/s11745-012-3673-2ISI: 000306789400007OAI: oai:DiVA.org:uu-179936DiVA: diva2:547121
Knut and Alice Wallenberg Foundation, KAW 2004.0123
Available from: 2012-08-27 Created: 2012-08-27 Last updated: 2012-10-29Bibliographically approved

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Jerneren, FredrikOliw, Ernst H.
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