Site Of Lipoxygenase Research Articles

Bioassay-guided isolation, identification and molecular ligand-target insight of lipoxygenase inhibitors from leaves of Anisomeles malabarica R.Br.

Background:Anisomeles malabarica R. Br. (Lamiaceae) is extensively used in traditional medicine in major parts of India for several medicinal purposes, including their use in rheumatism.Materials and Methods:The air-dried leaves of A. malabarica were extracted with ethanol, defatted with n-hexane and then successively partitioned into chloroform and n-butanol fractions. Bioassay-guided fractionation and purification of chloroform fraction from A. malabarica lead to the isolation of lipoxygenase (LOX) inhibitors. The structures of isolated compounds were elucidated by ultraviolet, infrared, 1H nuclear magnetic resonance (NMR), 13C NMR and mass spectrometry spectroscopic techniques and assessed further by in vitro soybean lipoxygenase (sLOX) assay. In addition, the enzyme type inhibition was evaluated through molecular docking technique as a part of computational study.Results:The bioactive compounds 3, 4 dihydroxy benzoic acid (1) and 4’, 5, 7-trihydroxyflavone (2) were isolated from chloroform fraction of A. malabarica, whose bioactivity was observed to be dose-dependent compared to n-butanol fraction. Among the compounds, 3, 4 dihydroxy benzoic acid showed significant sLOX inhibitory activity with 74.04% ±2.6% followed by 4’, 5, 7-trihydroxyflavone (34.68% ±1.9%). The computational analysis of compounds showed their molecular interaction with important amino acid residues and nonheme iron atom in the catalytic site of LOX by enlightening their potential binding mode at molecular level.Conclusions:The LOX inhibitory constituents were identified from A. malabarica by means of bioassay-guided fractionation process. The results derived from in vitro and computational experiments confirm the potential of the isolated compounds and provide additional evidence for its traditional use in inflammatory disorders.

Molecular insights to explore abietane diterpenes as new LOX inhibitors

Lipoxygenase (LOX) inhibitors are well known for their anti-inflammatory effects derived from both natural products and synthetic sources. Taxus wallichiana Zucc is traditionally used in the treatment of various diseases including inflammation. In a recent study, it was used to conduce the isolation of the bioactive compounds from Taxus wallichiana Zucc. thanks to its ethnopharmacological use in diseases including inflammation. Phytochemical studies lead to the isolation of abietane diterpenoids, namely Taxusabietane C (TC) and Taxamairin F (TF). Test compounds showed significant LOX inhibitory effects. IC50 values of TC and TF were 69 ± 0.31 and 73 ± 0.14 μM, respectively. The standard compound Baicalein and Tenidap sodium showed the IC50 value being 22.1 ± 0.03 and 41.6 ± 0.02 μM. Significant LOX inhibitory activities were supported by a molecular docking simulation to explore molecular interactions of the compounds with the active (catalytic) site of LOX.

Effect of exogenous iron on aerobic catalytic mechanism of soybean lipoxygenase

Effect of exogenous iron on the enzymatic activity of soybean lipoxygenase (LOX) in aerobic catalysis is investigated in this paper. It is found that a certain level of exogenous Fe 3+, Fe 2+ or linoleic acid hydroperoxide (LA-HPOD) could speed up the oxidation process of linoleic acid catalyzed by soybean LOX. Absorbency band between 300 and 400 nm in UV spectrum of soybean LOX was strengthened once upon adding of Fe 3+ but only changed slightly with adding of Fe 2+ or other ions. Therefore, Fe 3+ is much more affinitive to the catalytic site of LOX than other ions, although the previous experimental results have indicated that the iron inside native enzyme mainly exists as a more stable Fe 2+-LOX state. The potential function of exogenous Fe 3+, Fe 2+ or LA-HPOD in aerobic catalysis mechanism of soybean LOX is proposed as electron transfer from the intrinsic electron donor to the extrinsic electron acceptor, which speeds up the conversion from Fe 2+ to Fe 3+ in situ inside the active site of LOX. Since the above exogenous matters can accelerate the initial rate and thus the overall rate of aerobic catalysis of linoleic acid catalyzed by LOX, it becomes evident from our study that the conversion from Fe 2+ to Fe 3+ in situ is the control step in the above aerobic reaction.

A nonheme iron(II) complex that models the redox cycle of lipoxygenase.

The air-stable complex [Fe(6-Me3-TPA) (O2CAr)]+ [1; 6-Me3-TPA = tris(6-methyl-2-pyridylmethyl)amine] has been synthesized as a model for the iron(II) site of lipoxygenase. This iron(II) complex reacts with 0.5 equiv ROOH to form a yellow species, which has been formulated as [FeIII(OH)(6-Me3-TPA) (O2CAr)]+ (2) by electrospray mass spectrometry. Addition of more ROOH converts 2 into a purple species, which is characterized by electrospray ionization mass spectrometry and resonance Raman spectroscopy as [FeIII(OOR)(6-Me3-TPA)(O2CAr)]+. The purple species is metastable and decomposes via Fe-O bond homolysis to regenerate the starting iron(II) complex. These metal-centered transformations parallel the changes observed for lipoxygenase in its reaction with its product hydroperoxide.

Identification of Catalytically Active Groups of Wheat Germ Lipoxygenase

Dependences of lipoxygenase activity on pH, ionization enthalpies of various chemical groups, photoinactivation of the enzyme, and effects of specific reagents (p-CMB, DPF, and PMSF) were studied to identify catalytically active groups of the enzyme. The data suggest that the catalytic site of lipoxygenase includes imidazole and hydroxyl groups of histidine and serine residues, respectively.

Electron Spin Resonance Studies on the Lipoxygenase Reaction by Spin Trapping and Spin Labelling Methods

The rate of oxygenation and that of trapping linoleic acid free radicals in the lipoxygenase [EC 1.13.11.12] reaction were measured in the presence of linoleic acid, oxygen, and nitrosobenzene at various concentrations, with a Clark oxygen electrode and ESR spectroscopy. The results were interpreted under the assumption that the free radical of linoleic acid, an intermediate of the lipoxygenase reaction, reacts competitively with oxygen or nitrosobenzene. The oxidation of the iron in the active site of lipoxygenase caused by the spin label reagent, 2-(10-carboxydecyl)-2-hexyl-4,4-dimethyl-3-oxazolidinyloxyl, was also observed by ESR- and fluorescence-spectroscopy.