The iron ligand sphere geometry of mammalian 15-lipoxygenases

Abstract

We investigated the geometry of the iron ligand sphere of the native rabbit 15-lipoxygenase (15-LOX) by X-ray absorption spectroscopy using synchrotron radiation. The soybean LOX-1 was used as a reference compound because its iron ligand sphere is well characterized. For structural information the X-ray absorption spectra were evaluated using the Excurve Program (CCLRC Daresbury Laboratory, Warrington, U.K.). From the positions of the absorption edges and from the intensities of the 1s-3d pre-edge transition peaks a six-coordinate ferrous iron was concluded for the rabbit 15-LOX. Evaluation of the extended region of the absorption spectra suggested six nitrogen and/or oxygen atoms as direct iron ligands, and the following binding distances were determined (means+/-S.D.; estimated accuracy is +/-0.001nm for bond distances, on the basis of more than 22 X-ray absorption spectra): 0.213+/-0.001nm, 0.213+/-0. 001 nm, 0.236+/-0.001 nm, 0.293+/-0.001 nm, 0.189+/-0.001 nm and 0. 242+/-0.001. Lyophilization of the LOX altered the binding distances but did not destroy the octahedral iron ligand sphere. For construction of a structural model of the iron ligand sphere the binding distances extracted from the X-ray spectra were assigned to specific amino acids (His-360, -365, -540, -544 and the C-terminal Ile-662) by molecular modelling using the crystal coordinates of the soybean LOX-1 and of a rabbit 15-LOX-inhibitor complex.

15-Lipoxygenase Catalytically Consumes Nitric Oxide and Impairs Activation of Guanylate Cyclase

“…Inhibition of both plant and mammalian 15-LOX-dependent lipid oxidation by high concentrations of nitric oxide ( ⅐ NO) was ascribed to formation of an enzyme-nitrosyl complex (25)(26)(27). Nitric oxide can form a nitrosyl complex with the active site of 15-LOX, a single six-coordinate ferrous iron liganded to nitrogen and/or oxygen atoms (28), that is detectable by electron paramagnetic resonance spectroscopy (EPR). The spectrum of the soybean 15-LOX Fe 2ϩ -⅐ NO complex contains two species, the first attributed to either high spin ferric iron, formed by transfer of an electron from Fe 2ϩ to ⅐ NO, or an S ϭ 3/2 system resulting from antiferromagnetic coupling of axial (D Ͼ E) high spin ferrous iron to ⅐ NO (29 -31).…”

Photoactivation of an Inhibitor of the 12/15‐Lipoxygenase Pathway

“…[28,29] It constitutes a regular octahedron that is slightly distorted. Four histidine residues (His361, His366, His541, and His545) and the C-terminal isoleucine residue (Ile663) have been identified as protein iron ligands and the sixth ligand position appears to be occupied by a water molecule or a hydroxy ion.…”

Structure-Function Investigation of the Interaction of 1- and 2-Substituted 3-Hydroxypyridin-4-ones with 5-Lipoxygenase and Ribonucleotide Reductase

“…The various analyses are consistent with a six-coordinate iron, bound by three or four nitrogen ligands (imidazole) and one or two oxygen ligands in an octahedral symmetry. It is evident from x-ray analysis that not all six bonds to the central iron atom are from the protein and that there are possible coordination sites that are either unoccupied or coordinated by weak ligands (12)(13)(14)(15)(16). Such sites may represent potential sites of coordination by metal chelators (8).…”

“…Of significance is a 37-amino acid sequence containing 5 histidines that are conserved in all cloned lipoxygenases (10,11). It has been suggested that this region may be involved in iron coordination (10,11,15,16). 5-LO contains 1 mol of iron/mol of enzyme (10,11).…”

“…5-LO contains 1 mol of iron/mol of enzyme (10,11). Several groups (11)(12)(13)(14)(15)(16) have investigated the geometry of the iron center in lipoxygenase using the soybean enzyme as a model due to its greater stability and availability. The various analyses are consistent with a six-coordinate iron, bound by three or four nitrogen ligands (imidazole) and one or two oxygen ligands in an octahedral symmetry.…”