33964-75-9Relevant academic research and scientific papers
Development and Application of a Peroxyl Radical Clock Approach for Measuring Both Hydrogen-Atom Transfer and Peroxyl Radical Addition Rate Constants
Do, Quynh,Lee, David D.,Dinh, Andrew N.,Seguin, Ryan P.,Zhang, Rutan,Xu, Libin
supporting information, p. 153 - 168 (2020/12/23)
The rate-determining step in free radical lipid peroxidation is the propagation of the peroxyl radical, where generally two types of reactions occur: (a) hydrogen-atom transfer (HAT) from a donor to the peroxyl radical; (b) peroxyl radical addition (PRA) to a C=C double bond. Peroxyl radical clocks have been used to determine the rate constants of HAT reactions (kH), but no radical clock is available to measure the rate constants of PRA reactions (kadd). In this work, we modified the analytical approach on the linoleate-based peroxyl radical clock to enable the simultaneous measurement of both kH and kadd. Compared to the original approach, this new approach involves the use of a strong reducing agent, LiAlH4, to completely reduce both HAT and PRA-derived products and the relative quantitation of total linoleate oxidation products with or without reduction. The new approach was then applied to measuring the kH and kadd values for several series of organic substrates, including para- and meta-substituted styrenes, substituted conjugated dienes, and cyclic alkenes. Furthermore, the kH and kadd values for a variety of biologically important lipids were determined for the first time, including conjugated fatty acids, sterols, coenzyme Q10, and lipophilic vitamins, such as vitamins D3 and A.
The CYP74B and CYP74D divinyl ether synthases possess a side hydroperoxide lyase and epoxyalcohol synthase activities that are enhanced by the site-directed mutagenesis
Gorina, Svetlana S.,Grechkin, Alexander N.,Iljina, Tatiana M.,Mukhtarova, Lucia S.,Smirnova, Elena O.,Toporkova, Yana Y.
, (2020/09/16)
The CYP74 family of cytochromes P450 includes four enzymes of fatty acid hydroperoxide metabolism: allene oxide synthase (AOS), hydroperoxide lyase (HPL), divinyl ether synthase (DES), and epoxyalcohol synthase (EAS). The present work is concerned with catalytic specificities of three recombinant DESs, namely, the 9-DES (LeDES, CYP74D1) of tomato (Solanum lycopersicum), 9-DES (NtDES, CYP74D3) of tobacco (Nicotiana tabacum), and 13-DES (LuDES, CYP74B16) of flax (Linum usitatissimum), as well as their alterations upon the site-directed mutagenesis. Both LeDES and NtDES converted 9-hydroperoxides of linoleic and α?linolenic acids to divinyl ethers colneleic and colnelenic acids (respectively) with only minorities of HPL and EAS products. In contrast, LeDES and NtDES showed low efficiency towards the linoleate 13-hydroperoxide, affording only the low yield of epoxyalcohols. LuDES exhibited mainly the DES activity towards α?linolenate 13-hydroperoxide (preferred substrate), and HPL activity towards linoleate 13-hydroperoxide, respectively. In contrast, LuDES converted 9-hydroperoxides primarily to the epoxyalcohols. The F291V and A287G mutations within the I-helix groove region (SRS-4) of LuDES resulted in the loss of DES activity and the acquirement of the epoxyalcohol synthase activity. Thus, the studied enzymes exhibited the versatility of catalysis and its qualitative alterations upon the site-directed mutagenesis.
Catalytic production of oxo-fatty acids by lipoxygenases is mediated by the radical-radical dismutation between fatty acid alkoxyl radicals and fatty acid peroxyl radicals in fatty acid assembly
Takigawa, Yuta,Koshiishi, Ichiro
, p. 258 - 264 (2020/11/26)
Oxo-octadecadienoic acids (OxoODEs) act as peroxisome proliferator-activated receptor (PPAR) agonists biologically, and are known to be produced in the lipoxygenase/linoleate system. OxoODEs seem to originate from the linoleate alkoxyl radicals that are generated from (E/Z)-hydroperoxy octadecadienoic acids ((E/Z)HpODEs) by a pseudoperoxidase reaction that is catalyzed by ferrous lipoxygenase. However, the mechanism underlying the conversion of alkoxyl radical into OxoODE remains obscure. In the present study, we confirmed that OxoODEs are produced in the lipoxygenase/linoleate system in an oxygen-dependent manner. Interestingly, we revealed a correlation between the (E/Z)-OxoODEs content and the (E/E)-HpODEs content in the system. (E/E)-HpODEs could have been derived from (E/E)-linoleate peroxyl radicals, which are generated by the reaction between a free linoleate allyl radical and an oxygen molecule. Notably, the ferrous lipoxygenase-linoleate allyl radical (LOx(Fe2+)-L·) complex, which is an intermediate in the lipoxygenase/linoleate system, tends to dissociate into LOx(Fe2+) and a linoleate allyl radical. Subsequently, LOx(Fe2+) converts (E/Z)-HpODEs to an (E/Z)-linoleate alkoxyl radical through one-electron reduction. Taken together, we propose that (E/Z)-OxoODEs and (E/E)-HpODEs are produced through radical-radical dismutation between (E/Z)-linoleate alkoxyl radical and (E/E)-linoleate peroxyl radical. Furthermore, the production of (E/Z)OxoODEs and (E/E)-HpODEs was remarkably inhibited by a hydrophobic radical scavenger, 2,2,6,6-tetra-methylpiperidine 1-oxyl (TEMPO). On the contrary, water-miscible radical scavengers, 4-hydroxyl-2,2,6,6-tetramethylpiperidine 1-oxyl (OH-TEMPO) and 3-carbamoyl-2,2,5,5-tetramethyl-3-pyrroline-N-oxyl (CmΔP) only modestly or sparingly inhibited the production of (E/Z)-OxoODEs and (E/E)-HpODEs. These facts indicate that the radical-radical dismutation between linoleate alkoxyl radical and linoleate peroxyl radical proceeds in the interior of micelles.
Oxygenation reactions catalyzed by the F557V mutant of soybean lipoxygenase-1: Evidence for two orientations of substrate binding
Hershelman, Dillon,Kahler, Kirsten M.,Price, Morgan J.,Lu, Iris,Fu,Plumeri, Patricia A.,Karaisz, Fred,Bassett, Natasha F.,Findeis, Peter M.,Clapp, Charles H.
, (2019/09/10)
Plant lipoxygenases oxygenate linoleic acid to produce 13(S)-hydroperoxy-9Z,11E-octadecadienoic acid (13(S)-HPOD) or 9-hydroperoxy-10E,12Z-octadecadienoic acid (9(S)-HPOD). The manner in which these enzymes bind substrates and the mechanisms by which they control regiospecificity are uncertain. Hornung et al. (Proc. Natl. Acad. Sci. USA 96 (1999) 4192–4197) have identified an important residue, corresponding to phe-557 in soybean lipoxygenase-1 (SBLO-1). These authors proposed that large residues in this position favored binding of linoleate with the carboxylate group near the surface of the enzyme (tail-first binding), resulting in formation of 13(S)-HPOD. They also proposed that smaller residues in this position facilitate binding of linoleate in a head-first manner with its carboxylate group interacting with a conserved arginine residue (arg-707 in SBLO-1), which leads to 9(S)-HPOD. In the present work, we have tested these proposals on SBLO-1. The F557V mutant produced 33% 9-HPOD (S:R = 87:13) from linoleic acid at pH 7.5, compared with 8% for the wild-type enzyme and 12% with the F557V,R707L double mutant. Experiments with 11(S)-deuteriolinoleic acid indicated that the 9(S)-HPOD produced by the F557V mutant involves removal of hydrogen from the pro-R position on C-11 of linoleic acid, as expected if 9(S)-HPOD results from binding in an orientation that is inverted relative to that leading to 13(S)-HPOD. The product distributions obtained by oxygenation of 10Z,13Z-nonadecadienoic acid and arachidonic acid by the F557V mutant support the hypothesis that ω6 oxygenation results from tail-first binding and ω10 oxygenation from head-first binding. The results demonstrate that the regiospecificity of SBLO-1 can be altered by a mutation that facilitates an alternative mode of substrate binding and adds to the body of evidence that 13(S)-HPOD arises from tail-first binding.
Epoxyalcohol Synthase RjEAS (CYP74A88) from the Japanese Buttercup (Ranunculus japonicus): Cloning and Characterization of Catalytic Properties
Toporkova,Fatykhova,Gorina,Mukhtarova,Grechkin
, p. 171 - 180 (2019/04/01)
Cytochromes P450 of the CYP74 family play a key role in the lipoxygenase cascade generating oxylipins (products of polyunsaturated fatty acid oxidation). The CYP74 family includes allene oxide synthases, hydroperoxide lyases, divinyl ether synthases, and epoxyalcohol synthases. In this work, we cloned the CYP74A88 gene from the Japanese buttercup (Ranunculus japonicus) and studied the properties of the encoded recombinant protein. The CYP74A88 enzyme specifically converts linoleic acid 9-and 13-hydroperoxides to oxiranyl carbinols 9,10-epoxy-11-hydroxy-12-octadecenoic acid and 11-hydroxy-12,13-epoxy-9-octadecenoic acid, respectively, which was confirmed by GC-MS analysis and kinetic studies. Therefore, the CYP74A88 enzyme is a specific epoxyalcohol synthase.
Biosynthesis of Jasmonates from Linoleic Acid by the Fungus Fusarium oxysporum. Evidence for a Novel Allene Oxide Cyclase
Oliw, Ernst H.,Hamberg, Mats
, p. 543 - 556 (2019/08/12)
Fusarium oxysporum f. sp. tulipae (FOT) secretes (+)-7-iso-jasmonoyl-(S)-isoleucine ((+)-JA-Ile) to the growth medium together with about 10 times less 9,10-dihydro-(+)-7-iso-JA-Ile. Plants and fungi form (+)-JA-Ile from 18:3n-3 via 12-oxophytodienoic acid (12-OPDA), which is formed sequentially by 13S-lipoxygenase, allene oxide synthase (AOS), and allene oxide cyclase (AOC). Plant AOC does not accept linoleic acid (18:2n-6)-derived allene oxides and dihydrojasmonates are not commonly found in plants. This raises the question whether 18:2n-6 serves as the precursor of 9,10-dihydro-JA-Ile in Fusarium, or whether the latter arises by a putative reductase activity operating on the n-3 double bond of (+)-JA-Ile or one of its precursors. Incubation of pentadeuterated (d5) 18:3n-3 with mycelia led to the formation of d5-(+)-JA-Ile whereas d5-9,10-dihydro-JA-Ile was not detectable. In contrast, d5-9,10-dihydro-(+)-JA-Ile was produced following incubation of [17,17,18,18,18-2H5]linoleic acid (d5-18:2n-6). Furthermore, 9(S),13(S)-12-oxophytoenoic acid, the 15,16-dihydro analog of 12-OPDA, was formed upon incubation of unlabeled or d5-18:2n-6. Appearance of the α-ketol, 12-oxo-13-hydroxy-9-octadecenoic acid following incubation of unlabeled or [13C18]-labeled 13(S)-hydroperoxy-9(Z),11(E)-octadecadienoic acid confirmed the involvement of AOS and the biosynthesis of the allene oxide 12,13(S)-epoxy-9,11-octadecadienoic acid. The lack of conversion of this allene oxide by AOC in higher plants necessitates the conclusion that the fungal AOC is distinct from the corresponding plant enzyme.
Allene Oxide Synthase Pathway in Cereal Roots: Detection of Novel Oxylipin Graminoxins
Grechkin, Alexander N.,Ogorodnikova, Anna V.,Egorova, Alevtina M.,Mukhitova, Fakhima K.,Ilyina, Tatiana M.,Khairutdinov, Bulat I.
, p. 336 - 343 (2018/06/04)
Young roots of wheat, barley, and sorghum, as well as methyl jasmonate pretreated rice seedlings, undergo an unprecedented allene oxide synthase pathway targeted to previously unknown oxylipins 1–3. These Favorskii-type products, (4Z)-2-pentyl-4-tridecene-1,13-dioic acid (1), (2′Z)-2-(2′-octenyl)-decane-1,10-dioic acid (2), and (2′Z,5′Z)-2-(2′,5′-octadienyl)-decane-1,10-dioic acid (3), have a carboxy function at the side chain, as revealed by their MS and NMR spectral data. Compounds 1–3 were the major oxylipins detected, along with the related α-ketols. Products 1–3 were biosynthesized from (9Z,11E,13S)-13-hydroperoxy-9,11-octadecadienoic acid, (9S,10E,12Z)-9-hydroperoxy-10,12-octadecadienoic acid (9-HPOD), and (9S,10E,12Z,15Z)-9-hydroperoxy-10,12,15-octadecatrienoic acid, respectively, via the corresponding allene oxides and cyclopropanones. The data indicate that conversion of the allene oxide into the cyclopropanone is controlled by soluble cyclase. The short-lived cyclopropanones are hydrolyzed to products 1–3. The collective name “graminoxins” has been ascribed to oxylipins 1–3.
N-linoleoylamino acids as chiral probes of substrate binding by soybean lipoxygenase-1
Clapp, Charles H.,Pachuski, Justin,Bassett, Natasha F.,Bishop, Kathleen A.,Carter, Gillian,Young, Megan,Young, Thomas,Fu, Yuhan
, p. 170 - 177 (2018/03/24)
Lipoxygenases catalyze the oxygenation of polyunsaturated fatty acids and their derivatives to produce conjugated diene hydroperoxides. Soybean lipoxygenase-1 (SBLO-1) has been the subject of intensive structural and mechanistic study, but the manner in which this enzyme binds substrates is uncertain. Previous studies suggest that the fatty acyl group of the substrate binds in an internal cavity near the catalytic iron with the polar end at the surface of the protein or perhaps external to the protein. To test this model, we have investigated two pairs of enantiomeric N-linoleoylamino acids as substrates for SBLO-1. If the amino acid moiety binds external to the protein, the kinetics and product distribution should show little or no sensitivity to the stereochemical configuration of the amino acid moiety. Consistent with this expectation, N-linoleoyl-L-valine (LLV) and N-linoleoyl-D-valine (LDV) are both good substrates with kcat/Km values that are equal within error and about 40% higher than kcat/Km for linoleic acid. Experiments with N-linoleoyl-L-tryptophan (LLT) and N-linoleoyl-D-tryptophan (LDT) were complicated by the low critical micelle concentrations (CMC = 6–8 μM) of these substances. Below the CMC, LDT is a better substrate by a factor of 2.7. The rates of oxygenation of LDT and LLT continue to rise above the CMC, with modest stereoselectivity in favor of the D enantiomer. With all of the substrates tested, the major product is the 13(S)-hydroperoxide, and the distribution of minor products is not appreciably affected by the configuration of the amino acid moiety. The absence of stereoselectivity with LLV and LDV, the modest magnitude of the stereoselectivity with LLT and LDT, and the ability micellar forms of LLT and LDT to increase the concentration of available substrate are all consistent with the hypothesis that the amino acid moiety binds largely external to SBLO-1 and interacts with it only weakly.
Ascorbic acid 6-palmitate: A potent inhibitor of human and soybean lipoxygenase-dependent lipid peroxidation
Mohamed, Riyaz,Tarannum, Shaista,Yariswamy, Manjunath,Vivek, Hamse K.,Siddesha, Jalahalli M.,Angaswamy, Nataraju,Vishwanath, Bannikuppe S.
, p. 769 - 778 (2014/06/09)
Objectives Lipoxygenases (LOX) are the key enzymes involved in the biosynthesis of leukotrienes and reactive oxygen species, which are implicated in pathophysiology of inflammatory disorders. This study was conducted to evaluate the inhibitory effect of water-soluble antioxidant ascorbic acid and its lipophilic derivative, ascorbic acid 6-palmitate (Vcpal) on polymorphonuclear lymphocyte 5-LOX and soybean 15-LOX (sLOX) in vitro. Methods LOX activity was determined by measuring the end products, 5-hydroperoxy eicosatetraenoic acid (5-HETE) and lipid hydroperoxides, by spectrophotometric and high performance liquid chromatography methods. The substrate-dependent enzyme kinetics and docking studies were carried out to understand the nature of inhibition. Key findings Vcpal potently inhibited 5-LOX when compared with its inhibitory effect on sLOX (IC50; 2.5 and 10.3μm respectively, P= 0.003). Further, Vcpal inhibited 5-LOX more strongly than the known synthetic drugs: phenidone and nordihydroguaiaretic acid (P= 0.0007). Enzyme kinetic studies demonstrated Vcpal as a non-competitive reversible inhibitor of 5-LOX. In-silico molecular docking revealed high MolDock and Rerank score for Vcpal than ascorbic acid, complementing in-vitro results. Conclusion Both in-vitro and docking studies demonstrated Vcpal but not ascorbic acid as a non-competitive inhibitor of 5-LOX- and sLOX-induced lipid peroxidation, suggesting a key role for lipophilic nature in bringing about inhibition.
The hydroperoxide moiety of aliphatic lipid hydroperoxides is not affected by hypochlorous acid
Zschaler, Josefin,Arnhold, Juergen
, p. 42 - 51 (2015/01/09)
The oxidation of polyunsaturated fatty acids to the corresponding hydroperoxide by plant and animal lipoxygenases is an important step for the generation of bioactive lipid mediators. Thereby fatty acid hydroperoxide represent a common intermediate, also in human innate immune cells, like neutrophil granulocytes. In these cells a further key component is the heme protein myeloperoxidase producing HOCl as a reactive oxidant. On the basis of different investigation a reaction of the fatty acid hydroperoxide and hypochlorous acid (HOCl) could be assumed. Here, chromatographic and spectrometric analysis revealed that the hydroperoxide moiety of 15S-hydroperoxy-5Z,8Z,11Z,13E-eicosatetraenoic acid (15-HpETE) and 13S-hydroperoxy-9Z,11E-octadecadienoic acid (13-HpODE) is not affected by HOCl. No reduction of the hydroperoxide group due to a reaction with HOCl could be measured. It could be demonstrated that the double bonds of the fatty acid hydroperoxides are the major target of HOCl, present either as reagent or formed by the myeloperoxidase-hydrogen peroxide-chloride system.
