Publications

• Secretion of two novel enzymes, manganese 9S-lipoxygenase and epoxy alcohol synthase, by the rice pathogen Magnaporthe salvinii.

  Wennman A., Oliw E.H.

  The mycelium of the rice stem pathogen, Magnaporthe salvinii, secreted linoleate 9S-lipoxygenase (9S-LOX) and epoxy alcohol synthase (EAS). The EAS rapidly transformed 9S-hydroperoxy-octadeca-10E,12Z-dienoic acid (9S-HPODE) to threo 10 (11)-epoxy-9S-hydroxy-12Z-octadecenoic acid, but other hydrope ... >> More

  The mycelium of the rice stem pathogen, Magnaporthe salvinii, secreted linoleate 9S-lipoxygenase (9S-LOX) and epoxy alcohol synthase (EAS). The EAS rapidly transformed 9S-hydroperoxy-octadeca-10E,12Z-dienoic acid (9S-HPODE) to threo 10 (11)-epoxy-9S-hydroxy-12Z-octadecenoic acid, but other hydroperoxy FAs were poor substrates. 9S-LOX was expressed in Pichia pastoris. Recombinant 9S-LOX oxidized 18:2n-6 directly to 9S-HPODE, the end product, and also to two intermediates, 11S-hydroperoxy-9Z,12Z-octadecenoic acid (11S-HPODE; ∼5%) and 13R-hydroperoxy-9Z,11E-octadecadienoic acid (13R-HPODE; ∼1%). 11S- and 13R-HPODE were isomerized to 9S-HPODE, probably after oxidation to peroxyl radicals, β-fragmentation, and oxygen insertion at C-9. The 18:3n-3 was oxidized at C-9, C-11, and C-13, and to 9,16-dihydroxy-10E,12,14E-octadecatrienoic acid. 9S-LOX contained catalytic manganese (Mn:protein ∼0.2:1; Mn/Fe, 1:0.05), and its sequence could be aligned with 77% identity to 13R-LOX with catalytic manganese lipoxygenase (13R-MnLOX) of the Take-all fungus. The Leu350Met mutant of 9S-LOX shifted oxidation of 18:2n-6 from C-9 to C-13, and the Phe347Leu, Phe347Val, and Phe347Ala mutants of 13R-MnLOX from C-13 to C-9. In conclusion, M. salvinii secretes 9S-LOX with catalytic manganese along with a specific EAS. Alterations in the Sloane determinant of 9S-LOX and 13R-MnLOX with larger and smaller hydrophobic residues interconverted the regiospecific oxidation of 18:2n-6, presumably by altering the substrate position in relation to oxygen insertion. << Less

  J. Lipid Res. 54:762-775(2013) [PubMed] [EuropePMC]

  This publication is cited by 12 other entries.

• Crystal structure of manganese lipoxygenase of the rice blast fungus Magnaporthe oryzae.

  Wennman A., Oliw E.H., Karkehabadi S., Chen Y.

  Lipoxygenases (LOX) are non-heme metal enzymes, which oxidize polyunsaturated fatty acids to hydroperoxides. All LOX belong to the same gene family, and they are widely distributed. LOX of animals, plants, and prokaryotes contain iron as the catalytic metal, whereas fungi express LOX with iron or ... >> More

  Lipoxygenases (LOX) are non-heme metal enzymes, which oxidize polyunsaturated fatty acids to hydroperoxides. All LOX belong to the same gene family, and they are widely distributed. LOX of animals, plants, and prokaryotes contain iron as the catalytic metal, whereas fungi express LOX with iron or with manganese. Little is known about metal selection by LOX and the adjustment of the redox potentials of their protein-bound catalytic metals. Thirteen three-dimensional structures of animal, plant, and prokaryotic FeLOX are available, but none of MnLOX. The MnLOX of the most important plant pathogen, the rice blast fungusMagnaporthe oryzae(Mo), was expressed inPichia pastoris.Mo-MnLOX was deglycosylated, purified to homogeneity, and subjected to crystal screening and x-ray diffraction. The structure was solved by sulfur and manganese single wavelength anomalous dispersion to a resolution of 2.0 Å. The manganese coordinating sphere is similar to iron ligands of coral 8R-LOX and soybean LOX-1 but is not overlapping. The Asn-473 is positioned on a short loop (Asn-Gln-Gly-Glu-Pro) instead of an α-helix and forms hydrogen bonds with Gln-281. Comparison with FeLOX suggests that Phe-332 and Phe-525 might contribute to the unique suprafacial hydrogen abstraction and oxygenation mechanism of Mo-MnLOX by controlling oxygen access to the pentadiene radical. Modeling suggests that Arg-525 is positioned close to Arg-182 of 8R-LOX, and both residues likely tether the carboxylate group of the substrate. An oxygen channel could not be identified. We conclude that Mo-MnLOX illustrates a partly unique variation of the structural theme of FeLOX. << Less

  J. Biol. Chem. 291:8130-8139(2016) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Expression and characterization of manganese lipoxygenase of the rice blast fungus reveals prominent sequential lipoxygenation of alpha-linolenic acid.

  Wennman A., Jerneren F., Magnuson A., Oliw E.H.

  Magnaporthe oryzae causes rice blast disease and has become a model organism of fungal infections. M. oryzae can oxygenate fatty acids by 7,8-linoleate diol synthase, 10R-dioxygenase-epoxy alcohol synthase, and by a putative manganese lipoxygenase (Mo-MnLOX). The latter two are transcribed during ... >> More

  Magnaporthe oryzae causes rice blast disease and has become a model organism of fungal infections. M. oryzae can oxygenate fatty acids by 7,8-linoleate diol synthase, 10R-dioxygenase-epoxy alcohol synthase, and by a putative manganese lipoxygenase (Mo-MnLOX). The latter two are transcribed during infection. The open reading frame of Mo-MnLOX was deduced from genome and cDNA analysis. Recombinant Mo-MnLOX was expressed in Pichia pastoris and purified to homogeneity. The enzyme contained protein-bound Mn and oxidized 18:2n-6 and 18:3n-3 to 9S-, 11-, and 13R-hydroperoxy metabolites by suprafacial hydrogen abstraction and oxygenation. The 11-hydroperoxides were subject to β-fragmentation with formation of 9S- and 13R-hydroperoxy fatty acids. Oxygen consumption indicated apparent kcat values of 2.8 s(-1) (18:2n-6) and 3.9 s(-1) (18:3n-3), and UV analysis yielded apparent Km values of 8 and 12 μM, respectively, for biosynthesis of cis-trans conjugated metabolites. 9S-Hydroperoxy-10E,12Z,15Z-octadecatrienoic acid was rapidly further oxidized to a triene, 9S,16S-dihydroperoxy-10E,12Z,14E-octadecatrienoic acid. In conclusion, we have expressed, purified and characterized a new MnLOX from M. oryzae. The pathogen likely secretes Mo-MnLOX and phospholipases to generate oxylipins and to oxidize lipid membranes of rice cells and the cuticle. << Less

  Arch. Biochem. Biophys. 583:87-95(2015) [PubMed] [EuropePMC]

  This publication is cited by 8 other entries.