The mechanism of α-oxidation in leaves

Hitchcock, C. H. S.; James, A. T.

doi:10.1016/0005-2760(66)90112-3

Cited by 76 publications

(26 citation statements)

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“…The addition of palmitic acid to the crude homogenate prepared from Ulva pertusa led to an increased yield of the corresponding (R)-2-hydroperoxy acid (5), which is further decarboxylated to form n-pentadecanal (Scheme 1). The fatty acid hydroperoxides as the reaction intermediates in plant ␣-oxidation system were also proposed in higher plants, such as peas (Pisum sativum) leaf (6), germinating peanuts (Arachis hypogaea) (7), cucumbers (Cucumis sativus) (8), and potatoes (Solanum tuberosum) (9) as well as in marine green algae (U. pertusa). The enzyme system requires neither hydrogen peroxide nor 2-oxoglutarate, but it needs molecular oxygen to facilitate its catalytic reaction, which suggested that the enzyme involved in plant ␣-oxidation system is distinct from those in mammals or bacteria.…”

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Catalytic Properties of Rice α-Oxygenase

Koeduka

Matsui

Akakabe

et al. 2002

Journal of Biological Chemistry

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Long-chain fatty acids can be metabolized to C n؊1 aldehydes by ␣-oxidation in plants. The reaction mechanism of the enzyme has not been elucidated. In this study, a complete nucleotide sequence of fatty acid ␣-oxygenase gene in rice plants (Oryza sativa) was isolated. The deduced amino acid sequence showed some similarity with those of mammalian prostaglandin H synthases (PGHSs). The gene was expressed in Escherichia coli and purified to apparently homogenous state. It showed the highest activity with linoleic acid and predominantly formed 2-hydroperoxide of the fatty acid (C n ), which is then spontaneously decarboxylated to form corresponding C n؊1 aldehyde. With linoleic or linoleic acids as a substrate, rice ␣-oxygenase formed no product having a max at approximately 234 nm, which indicated that the enzyme could not oxygenize the pentadiene system in the substrate. The spectroscopic feature of the purified enzyme in its ferrous state is similar to that of mammalian PGHS, whereas that of dithionite-reduced state showed significant difference. Site-directed mutagenesis revealed that His-158, Tyr-380, and Ser-558 were essential for the ␣-oxygenase activity. These residues are conserved in PGHS and known as a heme ligand, a source of a radical species to initiate oxygenation reaction and a residue involved in substrate binding, respectively. This finding suggested that the initial step of the oxygenation reaction in ␣-oxygenase has a high similarity with that of PGHS. The rice ␣-oxygenase activity was inhibited by imidazole but hardly inhibited by nonsteroidal anti-inflammatory drugs, such as aspirin, ibuprofen, and flurbiprofen, which are known as typical PGHS inhibitors. In addition, peroxidase activity could not be detected with ␣-oxygenase when palmitic acid 2-hydroperoxide was used as a substrate. From these findings, the catalytic resemblance between ␣-oxygenase and PGHS seems to be evident, although there still are differences in their substrate recognitions and peroxidation activities.

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Catalytic Properties of Rice α-Oxygenase

Koeduka

Matsui

Akakabe

et al. 2002

Journal of Biological Chemistry

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“…␣-Oxidation of fatty acids is widely observed in bacteria (1), yeasts (2), plants (3,4), and mammals (5,6). This reaction pathway results in catalysis of fatty acid to produce the corresponding fatty acid with one less carbon atom, where the fatty acid is first hydroxylated at the 2-position and sequentially decarboxylated to release carbon dioxide.…”

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Molecular Cloning and Expression of Fatty Acid α-Hydroxylase from Sphingomonas paucimobilis

Matsunaga¹,

Yokotani²,

Gotoh³

et al. 1997

Journal of Biological Chemistry

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Fatty acid ␣-hydroxylase (FAAH) catalyzes the initial reaction in ␣-oxidation of fatty acid to produce 2-hydroxy fatty acid. FAAH activity has been detected in a wide range of organisms from prokaryotes to eukaryotes. Here, we describe cloning of the FAAH gene from Sphingomonas paucimobilis, a sphingolipid-and 2-hydroxymyristic acid-rich bacterium. The isolated gene encoded 415 amino acids. A homology search revealed that amino acid sequences highly conserved in cytochrome P450 (P450) were present in FAAH. Although the heme-binding cysteine was recognizable at position 361, the consensus in the heme-binding region was modified by an insertion. Overall, FAAH has no significant identity to the known P450s. CO difference spectrum of recombinant FAAH showed the characteristic one of P450, except this peak was at 445 nm. These results suggest bacterial FAAH is a novel member of the P450 superfamily.

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“…When D-[1 J4C, 2-3Hlpalmitic acid is similarly oxidised, D-2-hydroxy [ l-14C]palmitate is formed with loss of tritium (2401, retention). The mechanism of cr-hydroxylation is therefore replacement of x-hydrogen by hydroxyl with overall retention of configuration, Long-chain fatty acids are catabolised in leaves by successive a-oxidation [ 11 involving the formation of the corresponding 2-hydroxyacids [2]. During the oxidative attack on the a-position of [l4C]pa1mit8ic acid, L-2-hydroxypalmitic acid is apparently formed preferentially, but is immediately further oxidised [3] ; the hydroxyacid which accumulates has the D-configuration [4] (Fig.…”

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“…During the oxidative attack on the a-position of [l4C]pa1mit8ic acid, L-2-hydroxypalmitic acid is apparently formed preferentially, but is immediately further oxidised [3] ; the hydroxyacid which accumulates has the D-configuration [4] (Fig. 1) ration of acetone-dried powder has been described previously [1,2]. This was stirred in about forty times its weight of cold 0.2M phosphate buffer (pH 7.0) for 1 h and briefly centrifuged to remove most of the insoluble material.…”

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The Stereochemistry of α‐Oxidation of Fatty Acids in Plants

Hitchcock

1968

European Journal of Biochemistry

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ration of acetone-dried powder has been described previously [1,2]. This was stirred in about forty times its weight of cold 0.2M phosphate buffer (pH 7.0) for 1 h and briefly centrifuged to remove most of the insoluble material. Homogenates were made by coarsely chopping the leaves into about five times their weight of cold 0.2 M phosphate buffer (pH7.0) and homogenising with an Ultra Turrax tissue grinder (Hudes Merchandising, Leeke St., London) for about 45 sec a t a temperature below 10". Homogenates and acetone-dried powder solutions were used immediately after preparation.The Radiochemical Centre (Amersham) provided [1-14C]palmitic acid (specific activity, 44 mC/mmole). Preparation of ~-[ 2 -~H ] 1 6 :0 and ~-[ 2 -~H ] 1 6 : 0The preparation of D-and L-2-hydroxypalmitic acid, essentially by the method of Horn and Pretorius [5], has been described previously [3]. Each enantiomer was converted into its tosyl derivative with retention of configuration by treatment with p-toluenesulphonyl chloride in pyridine. Each tosylate was refluxed in anhydrous tetrahydrofuran for 20 h with lithium boron [3H]hydride, when hydrogenolysis with inversion of configuration occurs [6,7]. The hexadecanol formed was oxidised t o the acid with chromium trioxide in acetic acid; D-16h:O thus gave ~-[2-~H]16 : 0, which was purified by thin-layer and gas-liquid chromatography. Similarly L-16 h : 0 yielded ~-[ 2 -~H ] 1 6 : 0 (Fig.2). METHODSSubstrates were prepared by mixing [1-l4C]16: 0 and the [2-3H]16:0 in benzene solution, aliquots of which were assayed for 14C and 3H simultaneously in a Packard Tri Carb spectrometer. The 3H :14C ratio

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The mechanism of α-oxidation in leaves

Cited by 76 publications

References 49 publications

Catalytic Properties of Rice α-Oxygenase

Catalytic Properties of Rice α-Oxygenase

Molecular Cloning and Expression of Fatty Acid α-Hydroxylase from Sphingomonas paucimobilis

The Stereochemistry of α‐Oxidation of Fatty Acids in Plants

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