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

Abstract

Analysis of purified soybean and rabbit reticulocyte 15-lipoxygenase (15-LOX) and PA317 cells transfected with human 15-LOX revealed a rapid rate of linoleate-dependent nitric oxide (⋅NO) uptake that coincided with reversible inhibition of product ((13S)-hydroperoxyoctadecadienoic acid, or (13S)-HPODE) formation. No reaction of ⋅NO (up to 2 μm) with either native (Ered) or ferric LOXs (0.2 μm) metal centers to form nitrosyl complexes occurred at these ⋅NO concentrations. During HPODE-dependent activation of 15-LOX, there was consumption of 2 mol of ⋅NO/mol of 15-LOX. Stopped flow fluorescence spectroscopy showed that ⋅NO (2.2 μm) did not alter the rate or extent of (13S)-HPODE-induced tryptophan fluorescence quenching associated with 15-LOX activation. Additionally,⋅NO does not inhibit the anaerobic peroxidase activity of 15-LOX, inferring that the inhibitory actions of ⋅NO are due to reaction with the enzyme-bound lipid peroxyl radical, rather than impairment of (13S)-HPODE-dependent enzyme activation. From this, a mechanism of 15-LOX inhibition by ⋅NO is proposed whereby reaction of ⋅NO with EredLOO⋅ generates Ered and LOONO, which hydrolyzes to (13S)-HPODE and nitrite (NO 2 −). Reactivation of Ered, considerably slower than dioxygenase activity, is then required to complete the catalytic cycle and leads to a net inhibition of rates of (13S)-HPODE formation. This reaction of ⋅NO with 15-LOX inhibited ⋅NO-dependent activation of soluble guanylate cyclase and consequent cGMP production. Since accelerated ⋅NO production, enhanced 15-LOX gene expression, and 15-LOX product formation occurs in diverse inflammatory conditions, these observations indicate that reactions of ⋅NO with lipoxygenase peroxyl radical intermediates will result in modulation of both ⋅NO bioavailability and rates of production of lipid signaling mediators. Analysis of purified soybean and rabbit reticulocyte 15-lipoxygenase (15-LOX) and PA317 cells transfected with human 15-LOX revealed a rapid rate of linoleate-dependent nitric oxide (⋅NO) uptake that coincided with reversible inhibition of product ((13S)-hydroperoxyoctadecadienoic acid, or (13S)-HPODE) formation. No reaction of ⋅NO (up to 2 μm) with either native (Ered) or ferric LOXs (0.2 μm) metal centers to form nitrosyl complexes occurred at these ⋅NO concentrations. During HPODE-dependent activation of 15-LOX, there was consumption of 2 mol of ⋅NO/mol of 15-LOX. Stopped flow fluorescence spectroscopy showed that ⋅NO (2.2 μm) did not alter the rate or extent of (13S)-HPODE-induced tryptophan fluorescence quenching associated with 15-LOX activation. Additionally,⋅NO does not inhibit the anaerobic peroxidase activity of 15-LOX, inferring that the inhibitory actions of ⋅NO are due to reaction with the enzyme-bound lipid peroxyl radical, rather than impairment of (13S)-HPODE-dependent enzyme activation. From this, a mechanism of 15-LOX inhibition by ⋅NO is proposed whereby reaction of ⋅NO with EredLOO⋅ generates Ered and LOONO, which hydrolyzes to (13S)-HPODE and nitrite (NO 2 −). Reactivation of Ered, considerably slower than dioxygenase activity, is then required to complete the catalytic cycle and leads to a net inhibition of rates of (13S)-HPODE formation. This reaction of ⋅NO with 15-LOX inhibited ⋅NO-dependent activation of soluble guanylate cyclase and consequent cGMP production. Since accelerated ⋅NO production, enhanced 15-LOX gene expression, and 15-LOX product formation occurs in diverse inflammatory conditions, these observations indicate that reactions of ⋅NO with lipoxygenase peroxyl radical intermediates will result in modulation of both ⋅NO bioavailability and rates of production of lipid signaling mediators. Lipoxygenases are a family of ubiquitously expressed non-heme iron-containing enzymes that oxidize the unsaturated fatty acids arachidonate and linoleate to bioactive hydroperoxides and other metabolites (Scheme FS1). For example, 5-LOX 1The abbreviations LOXlipoxygenaseLDLlow density lipoproteinHPLChigh performance liquid chromatographyPBSphosphate-buffered salineHPODEhydroperoxyoctadecadienoic acidETYAeicosatetraynoic acidDTPAdiethylenetriamine pentaacetic acidDEA-NONOate2-(N,N-diethylamino)-diazenolate-2-oxide 1The abbreviations LOXlipoxygenaseLDLlow density lipoproteinHPLChigh performance liquid chromatographyPBSphosphate-buffered salineHPODEhydroperoxyoctadecadienoic acidETYAeicosatetraynoic acidDTPAdiethylenetriamine pentaacetic acidDEA-NONOate2-(N,N-diethylamino)-diazenolate-2-oxidegenerates precursors for leukotrienes, products involved in inflammation and allergic responses (1Yamomoto S. Smith W.L. J. Lipid Med. Cell Signal. 1995; 12: 195Crossref PubMed Scopus (40) Google Scholar). 12-Lipoxygenases, present in vascular endothelium, smooth muscle cells, platelets, and leukocytes (2Hamberg M. Samuellsson B. Proc. N. Y. Acad. Sci. U. S. A. 1974; 71: 3400-3404Crossref Scopus (1321) Google Scholar, 3Yokoyama C. Shinjo F. Yoshimoto S. J. PubMed Google to vascular and 15-LOX is involved in and in 15-LOX of is a for (2Hamberg M. Samuellsson B. Proc. N. Y. Acad. Sci. U. S. A. 1974; 71: 3400-3404Crossref Scopus (1321) Google Scholar, Lipid PubMed Scopus Google Scholar, Cell for 15-LOX in of in the of 15-LOX activity, and the of 15-LOX products that in human and rabbit J. S. S. J. Med. PubMed Scopus Google Scholar, S. S. J. PubMed Scopus Google Scholar, J. 1995; PubMed Scopus Google Scholar, J. J. PubMed Scopus Google Scholar). and of density by and the of and cells in the vascular is a of that and oxidize and S. Proc. Acad. Sci. U. S. A. PubMed Scopus Google Scholar, S. J. Lipid PubMed Google Scholar, A. Med. PubMed Scopus Google Scholar). that gene of 15-LOX to and with in of and of vascular S. J. J. 1995; PubMed Scopus Google Scholar, A. Google Scholar). to these of 15-LOX is the that of rabbit 15-LOX J. J. Scopus Google Scholar). in is inhibited by of a 15-LOX B. N. A. J. PubMed Scopus Google Scholar). these observations of the of 15-LOX, 15-LOX and vascular cells with of vascular and in ⋅NO a by peroxyl and in purified or by cells or U. PubMed Scopus Google Scholar, N. A. PubMed Scopus Google Scholar, N. 1995; PubMed Scopus Google Scholar, N. B. 1995; PubMed Scopus Google Scholar, PubMed Scopus Google Scholar, N. B. J. A. S. PubMed Scopus Google Scholar, M. PubMed Scopus Google Scholar, M. B. S. M. J. PubMed Google Scholar). of both and lipid by of nitric oxide (⋅NO) was to formation of J. PubMed Scopus Google Scholar, J. J. PubMed Scopus Google Scholar, J. S. PubMed Scopus Google Scholar). oxide form a nitrosyl with the of 15-LOX, a to J. J. PubMed Scopus Google that is by spectroscopy of the soybean 15-LOX the to either ferric by of to or of to ⋅NO J. PubMed Scopus Google Scholar, PubMed Scopus Google Scholar, J. PubMed Google Scholar). for formation of is for soybean 15-LOX at J. PubMed Google Scholar). of the to a and ⋅NO of at for PubMed Scopus Google Scholar). of that of the to enzyme formation of the nitrosyl of ⋅NO to anaerobic 15-LOX in of a to that of the ferric enzyme native enzyme with J. PubMed Scopus Google Scholar). and of rabbit 15-LOX showed that with ferric J. PubMed Scopus Google Scholar). required for formation of the required to inhibit soybean 15-LOX catalytic activity S. S. M. B. 1995; PubMed Scopus Google and ⋅NO in Proc. Acad. Sci. U. S. A. PubMed Scopus Google Scholar, J. J. PubMed Scopus Google that 15-LOX inhibition does not formation. other are to in the inhibition of lipid to the form 15-LOX that with ⋅NO and to enzyme enzyme-bound lipid and reactions of ⋅NO with are at rates Scopus Google Scholar, S. PubMed Scopus Google Scholar). indicate that ⋅NO with in N. A. PubMed Scopus Google Scholar, N. B. J. A. S. PubMed Scopus Google Scholar, S. PubMed Scopus Google Scholar). Since ⋅NO the 15-LOX that enzyme-bound are to ⋅NO of ⋅NO by nitric oxide is in of vascular present in with and ⋅NO in the vascular J. PubMed Scopus Google is the that ⋅NO in the of ⋅NO in is J. Google to for by that consumption consumption with complete of the activation of soluble guanylate cyclase S. PubMed Scopus Google Scholar). of the inhibition of the ⋅NO signaling is the reaction of with to S. PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar, M. J. Proc. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). Since reactions of not for complete signaling to smooth muscle cells M. J. Proc. Acad. Sci. U. S. A. PubMed Scopus Google other that to ⋅NO consumption are are the radical intermediates the of enzymes During of in impairment of the vascular to or ⋅NO is a J. PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar, M. J. Proc. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). Since 15-LOX is to present in the in and ⋅NO in Proc. Acad. Sci. U. S. A. PubMed Scopus Google was of to reactions of lipid by 15-LOX at a rate to alter ⋅NO and the reactions of soybean and 15-LOX with ⋅NO at and indicate that there are for ⋅NO reaction 15-LOX and that ⋅NO consumption occurs inhibition of 15-LOX. was 15-LOX of lipid lipid radical reactions with ⋅NO in activation of soluble guanylate cyclase and the formation of these observations that lipoxygenase reactions with ⋅NO inhibit both lipoxygenase catalytic activity and ⋅NO-dependent that the vascular actions of both ⋅NO and lipoxygenase products ⋅NO rates of 15-LOX product formation in 15-LOX catalytic activity ⋅NO and guanylate cyclase activation. in the and actions of ⋅NO that in of ⋅NO or with PubMed Scopus Google Scholar, PubMed Scopus Google Scholar, J. 1995; PubMed Scopus Google Scholar, PubMed Google Scholar). to of ⋅NO with lipid and to that in the vascular For example, the of 15-LOX by cells transfected with the human 15-LOX gene is the M. M. N. A. S. J. Scopus Google the 15-LOX rates consumption are to rates of production in or are and Scopus Google Scholar, S. J. 1995; PubMed Scopus Google than the rates of ⋅NO consumption that the of 15-LOX in and a ⋅NO for soluble cyclase activation and This was by the that soluble guanylate cyclase formation of cGMP was the catalytic of arachidonate by the of ⋅NO reaction with and consumption by 15-LOX, purified rabbit reticulocyte and soybean 15-LOX. For both reversible inhibition was of coincided with ⋅NO and activity was ⋅NO was For ⋅NO of linoleate not (13S)-HPODE was required and ⋅NO consumption was that or product of the dioxygenase cycle with ⋅NO to inhibit 15-LOX intermediates that with ⋅NO are in Since linoleate was required the enzyme is the of ⋅NO reaction is ⋅NO anaerobic peroxidase activity a for consumption is the of ⋅NO in these are to with present at for reaction with at the the that (13S)-HPODE was the product that ⋅NO occurs 15-LOX and is by the During dioxygenase in a that by of a of a nitrosyl with the of 15-LOX J. PubMed Scopus Google Scholar, PubMed Scopus Google Scholar, J. PubMed Google Scholar). is that ⋅NO with (13S)-HPODE for reaction with 15-LOX flow of ⋅NO either the rate or extent of 15-LOX tryptophan fluorescence quenching was A. Scopus Google Scholar, A. J. PubMed Scopus Google Scholar). native 15-LOX did not ⋅NO and the of ⋅NO inhibition was of (13S)-HPODE was and that ⋅NO the of 15-LOX activity J. PubMed Scopus Google Scholar). of 15-LOX did not of ⋅NO the of ⋅NO inhibition is the ⋅NO was either or enzyme This is not 15-LOX activation and that enzyme inhibition reaction with a dioxygenase did not ⋅NO in the of nitrosyl complexes with 15-LOX metal centers did not This is not a at for the and soybean 15-LOX, in of ⋅NO in and was by reactions J. PubMed Google Scholar). Since formation of required at is a of 15-LOX inhibition that of ⋅NO oxidize native 15-LOX, to either activation or formation of a to activation J. PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar). both occurred formation of the nitrosyl was by of ⋅NO did not 15-LOX. for formation of the that conditions, ⋅NO will than both the formation of complexes and activation of 15-LOX by ⋅NO is During activation by consumption of 2 mol of ⋅NO/mol of 15-LOX was of ⋅NO to by the by reactions of that ⋅NO these ⋅NO 15-LOX tryptophan fluorescence ⋅NO uptake activation is to enzyme inhibition ⋅NO at dioxygenase that was not or is of ⋅NO with in is J. J. Scopus Google Scholar, N. 1995; PubMed Scopus Google Scholar, and J. PubMed Google Scholar). Since ⋅NO the of in a reaction of ⋅NO with EredLOO⋅ is ⋅NO with EredLOO⋅ to form LOONO, of product the enzyme (Ered) that by for of the catalytic cycle of soybean 15-LOX is of the rate of linoleate J. J. PubMed Scopus Google activation of rabbit 15-LOX is of the of dioxygenase activity A. J. PubMed Scopus Google Scholar). formation of the enzyme Ered by ⋅NO reaction with EredLOO⋅ will the rate of dioxygenase activity and product (Scheme product of reaction ⋅NO and at in and S. PubMed Scopus Google to form the either to form or to which with of and of of these to is Since of either in the or of ⋅NO and or was of at the 15-LOX to and showed that ⋅NO reaction with soybean 15-LOX of and S. S. M. B. 1995; PubMed Scopus Google Scholar). to that ⋅NO is reaction with 15-LOX intermediates and enzyme did not and formation of and that with ⋅NO to form products Scopus Google Scholar, PubMed Scopus Google Scholar, PubMed Scopus Google Scholar). of with and of which at will to form M. S. 12: PubMed Scopus Google that at ⋅NO and 15-LOX in 15-LOX is 15-LOX catalytic activity ⋅NO-dependent activation of soluble guanylate and 15-LOX ⋅NO activation of 15-LOX, 2 mol of ⋅NO/mol of 15-LOX are reaction of ⋅NO with of the dioxygenase leads to reversible enzyme inhibition by formation of the ⋅NO reaction 15-LOX leads to in product of occurred in with consumption of of modulation of 15-LOX activity by ⋅NO occurs at ⋅NO of formation and ⋅NO consumption is in for ⋅NO activation of soluble cyclase is these will cGMP production and smooth muscle revealed in consumption of ⋅NO by the lipoxygenase present in a of vascular then to responses to ⋅NO J. PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar, M. J. Proc. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). Lipoxygenases are a family of ubiquitously expressed non-heme iron-containing enzymes that oxidize the unsaturated fatty acids arachidonate and linoleate to bioactive hydroperoxides and other metabolites (Scheme FS1). For example, 5-LOX 1The abbreviations LOXlipoxygenaseLDLlow density lipoproteinHPLChigh performance liquid chromatographyPBSphosphate-buffered salineHPODEhydroperoxyoctadecadienoic acidETYAeicosatetraynoic acidDTPAdiethylenetriamine pentaacetic acidDEA-NONOate2-(N,N-diethylamino)-diazenolate-2-oxide 1The abbreviations LOXlipoxygenaseLDLlow density lipoproteinHPLChigh performance liquid chromatographyPBSphosphate-buffered salineHPODEhydroperoxyoctadecadienoic acidETYAeicosatetraynoic acidDTPAdiethylenetriamine pentaacetic acidDEA-NONOate2-(N,N-diethylamino)-diazenolate-2-oxidegenerates precursors for leukotrienes, products involved in inflammation and allergic responses (1Yamomoto S. Smith W.L. J. Lipid Med. Cell Signal. 1995; 12: 195Crossref PubMed Scopus (40) Google Scholar). 12-Lipoxygenases, present in vascular endothelium, smooth muscle cells, platelets, and leukocytes (2Hamberg M. Samuellsson B. Proc. N. Y. Acad. Sci. U. S. A. 1974; 71: 3400-3404Crossref Scopus (1321) Google Scholar, 3Yokoyama C. Shinjo F. Yoshimoto S. J. PubMed Google to vascular and 15-LOX is involved in and in 15-LOX of is a for (2Hamberg M. Samuellsson B. Proc. N. Y. Acad. Sci. U. S. A. 1974; 71: 3400-3404Crossref Scopus (1321) Google Scholar, Lipid PubMed Scopus Google Scholar, Cell Scholar). lipoxygenase density performance liquid pentaacetic lipoxygenase density performance liquid pentaacetic for 15-LOX in of in the of 15-LOX activity, and the of 15-LOX products that in human and rabbit J. S. S. J. Med. PubMed Scopus Google Scholar, S. S. J. PubMed Scopus Google Scholar, J. 1995; PubMed Scopus Google Scholar, J. J. PubMed Scopus Google Scholar). and of density by and the of and cells in the vascular is a of that and oxidize and S. Proc. Acad. Sci. U. S. A. PubMed Scopus Google Scholar, S. J. Lipid PubMed Google Scholar, A. Med. PubMed Scopus Google Scholar). that gene of 15-LOX to and with in of and of vascular S. J. J. 1995; PubMed Scopus Google Scholar, A. Google Scholar). to these of 15-LOX is the that of rabbit 15-LOX J. J. Scopus Google Scholar). in is inhibited by of a 15-LOX B. N. A. J. PubMed Scopus Google Scholar). these observations of the of 15-LOX, 15-LOX and vascular cells with of vascular and in ⋅NO a by peroxyl and in purified or by cells or U. PubMed Scopus Google Scholar, N. A. PubMed Scopus Google Scholar, N. 1995; PubMed Scopus Google Scholar, N. B. 1995; PubMed Scopus Google Scholar, PubMed Scopus Google Scholar, N. B. J. A. S. PubMed Scopus Google Scholar, M. PubMed Scopus Google Scholar, M. B. S. M. J. PubMed Google Scholar). of both and lipid by of nitric oxide (⋅NO) was to formation of J. PubMed Scopus Google Scholar, J. J. PubMed Scopus Google Scholar, J. S. PubMed Scopus Google Scholar). oxide form a nitrosyl with the of 15-LOX, a to J. J. PubMed Scopus Google that is by spectroscopy of the soybean 15-LOX the to either ferric by of to or of to ⋅NO J. PubMed Scopus Google Scholar, PubMed Scopus Google Scholar, J. PubMed Google Scholar). for formation of is for soybean 15-LOX at J. PubMed Google Scholar). of the to a and ⋅NO of at for PubMed Scopus Google Scholar). of that of the to enzyme formation of the nitrosyl of ⋅NO to anaerobic 15-LOX in of a to that of the ferric enzyme native enzyme with J. PubMed Scopus Google Scholar). and of rabbit 15-LOX showed that with ferric J. PubMed Scopus Google Scholar). required for formation of the required to inhibit soybean 15-LOX catalytic activity S. S. M. B. 1995; PubMed Scopus Google and ⋅NO in Proc. Acad. Sci. U. S. A. PubMed Scopus Google Scholar, J. J. PubMed Scopus Google that 15-LOX inhibition does not formation. other are to in the inhibition of lipid to the form 15-LOX that with ⋅NO and to enzyme enzyme-bound lipid and reactions of ⋅NO with are at rates Scopus Google Scholar, S. PubMed Scopus Google Scholar). indicate that ⋅NO with in N. A. PubMed Scopus Google Scholar, N. B. J. A. S. PubMed Scopus Google Scholar, S. PubMed Scopus Google Scholar). Since ⋅NO the 15-LOX that enzyme-bound are to ⋅NO of ⋅NO by nitric oxide is in of vascular present in with and ⋅NO in the vascular J. PubMed Scopus Google is the that ⋅NO in the of ⋅NO in is J. Google to for by that consumption consumption with complete of the activation of soluble guanylate cyclase S. PubMed Scopus Google Scholar). of the inhibition of the ⋅NO signaling is the reaction of with to S. PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar, M. J. Proc. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). Since reactions of not for complete signaling to smooth muscle cells M. J. Proc. Acad. Sci. U. S. A. PubMed Scopus Google other that to ⋅NO consumption are are the radical intermediates the of enzymes During of in impairment of the vascular to or ⋅NO is a J. PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar, M. J. Proc. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). Since 15-LOX is to present in the in and ⋅NO in Proc. Acad. Sci. U. S. A. PubMed Scopus Google was of to reactions of lipid by 15-LOX at a rate to alter ⋅NO and the reactions of soybean and 15-LOX with ⋅NO at and indicate that there are for ⋅NO reaction 15-LOX and that ⋅NO consumption occurs inhibition of 15-LOX. was 15-LOX of lipid lipid radical reactions with ⋅NO in activation of soluble guanylate cyclase and the formation of these observations that lipoxygenase reactions with ⋅NO inhibit both lipoxygenase catalytic activity and ⋅NO-dependent that the vascular actions of both ⋅NO and lipoxygenase products ⋅NO rates of 15-LOX product formation in 15-LOX catalytic activity ⋅NO and guanylate cyclase activation. in the and actions of ⋅NO that in of ⋅NO or with PubMed Scopus Google Scholar, PubMed Scopus Google Scholar, J. 1995; PubMed Scopus Google Scholar, PubMed Google Scholar). to of ⋅NO with lipid and to that in the vascular For example, the of 15-LOX by cells transfected with the human 15-LOX gene is the M. M. N. A. S. J. Scopus Google the 15-LOX rates consumption are to rates of production in or are and Scopus Google Scholar, S. J. 1995; PubMed Scopus Google than the rates of ⋅NO consumption that the of 15-LOX in and a ⋅NO for soluble cyclase activation and This was by the that soluble guanylate cyclase formation of cGMP was the catalytic of arachidonate by the of ⋅NO reaction with and consumption by 15-LOX, purified rabbit reticulocyte and soybean 15-LOX. For both reversible inhibition was of coincided with ⋅NO and activity was ⋅NO was For ⋅NO of linoleate not (13S)-HPODE was required and ⋅NO consumption was that or product of the dioxygenase cycle with ⋅NO to inhibit 15-LOX intermediates that with ⋅NO are in Since linoleate was required the enzyme is the of ⋅NO reaction is ⋅NO anaerobic peroxidase activity a for consumption is the of ⋅NO in these are to with present at for reaction with at the the that (13S)-HPODE was the product that ⋅NO occurs 15-LOX and is by the During dioxygenase in a that by of a of a nitrosyl with the of 15-LOX J. PubMed Scopus Google Scholar, PubMed Scopus Google Scholar, J. PubMed Google Scholar). is that ⋅NO with (13S)-HPODE for reaction with 15-LOX flow of ⋅NO either the rate or extent of 15-LOX tryptophan fluorescence quenching was A. Scopus Google Scholar, A. J. PubMed Scopus Google Scholar). native 15-LOX did not ⋅NO and the of ⋅NO inhibition was of (13S)-HPODE was and that ⋅NO the of 15-LOX activity J. PubMed Scopus Google Scholar). of 15-LOX did not of ⋅NO the of ⋅NO inhibition is the ⋅NO was either or enzyme This is not 15-LOX activation and that enzyme inhibition reaction with a dioxygenase did not ⋅NO in the of nitrosyl complexes with 15-LOX metal centers did not This is not a at for the and soybean 15-LOX, in of ⋅NO in and was by reactions J. PubMed Google Scholar). Since formation of required at is a of 15-LOX inhibition that of ⋅NO oxidize native 15-LOX, to either activation or formation of a to activation J. PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar). both occurred formation of the nitrosyl was by of ⋅NO did not 15-LOX. for formation of the that conditions, ⋅NO will than both the formation of complexes and activation of 15-LOX by ⋅NO is During activation by consumption of 2 mol of ⋅NO/mol of 15-LOX was of ⋅NO to by the by reactions of that ⋅NO these ⋅NO 15-LOX tryptophan fluorescence ⋅NO uptake activation is to enzyme inhibition ⋅NO at dioxygenase that was not or is of ⋅NO with in is J. J. Scopus Google Scholar, N. 1995; PubMed Scopus Google Scholar, and J. PubMed Google Scholar). Since ⋅NO the of in a reaction of ⋅NO with EredLOO⋅ is ⋅NO with EredLOO⋅ to form LOONO, of product the enzyme (Ered) that by for of the catalytic cycle of soybean 15-LOX is of the rate of linoleate J. J. PubMed Scopus Google activation of rabbit 15-LOX is of the of dioxygenase activity A. J. PubMed Scopus Google Scholar). formation of the enzyme Ered by ⋅NO reaction with EredLOO⋅ will the rate of dioxygenase activity and product (Scheme product of reaction ⋅NO and at in and S. PubMed Scopus Google to form the either to form or to which with of and of of these to is Since of either in the or of ⋅NO and or was of at the 15-LOX to and showed that ⋅NO reaction with soybean 15-LOX of and S. S. M. B. 1995; PubMed Scopus Google Scholar). to that ⋅NO is reaction with 15-LOX intermediates and enzyme did not and formation of and that with ⋅NO to form products Scopus Google Scholar, PubMed Scopus Google Scholar, PubMed Scopus Google Scholar). of with and of which at will to form M. S. 12: PubMed Scopus Google that at ⋅NO and 15-LOX in 15-LOX is 15-LOX catalytic activity ⋅NO-dependent activation of soluble guanylate and 15-LOX ⋅NO activation of 15-LOX, 2 mol of ⋅NO/mol of 15-LOX are reaction of ⋅NO with of the dioxygenase leads to reversible enzyme inhibition by formation of the ⋅NO reaction 15-LOX leads to in product of occurred in with consumption of of modulation of 15-LOX activity by ⋅NO occurs at ⋅NO of formation and ⋅NO consumption is in for ⋅NO activation of soluble cyclase is these will cGMP production and smooth muscle revealed in consumption of ⋅NO by the lipoxygenase present in a of vascular then to responses to ⋅NO J. PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar, M. J. Proc. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). that the vascular actions of both ⋅NO and lipoxygenase products ⋅NO rates of 15-LOX product formation in 15-LOX catalytic activity ⋅NO and guanylate cyclase activation. in the and actions of ⋅NO that in of ⋅NO or with PubMed Scopus Google Scholar, PubMed Scopus Google Scholar, J. 1995; PubMed Scopus Google Scholar, PubMed Google Scholar). to of ⋅NO with lipid and to that in the vascular For example, the of 15-LOX by cells transfected with the human 15-LOX gene is the M. M. N. A. S. J. Scopus Google the 15-LOX rates consumption are to rates of production in or are and Scopus Google Scholar, S. J. 1995; PubMed Scopus Google than the rates of ⋅NO consumption that the of 15-LOX in and a ⋅NO for soluble cyclase activation and This was by the that soluble guanylate cyclase formation of cGMP was the catalytic of arachidonate by 15-LOX. the of ⋅NO reaction with and consumption by 15-LOX, purified rabbit reticulocyte and soybean 15-LOX. For both reversible inhibition was of coincided with ⋅NO and activity was ⋅NO was For ⋅NO of linoleate not (13S)-HPODE was required and ⋅NO consumption was that or product of the dioxygenase cycle with ⋅NO to inhibit 15-LOX. 15-LOX intermediates that with ⋅NO are in Since linoleate was required the enzyme is the of ⋅NO reaction is ⋅NO anaerobic peroxidase activity a for consumption is the of ⋅NO in these are to with present at for reaction with at the the that (13S)-HPODE was the product that ⋅NO occurs 15-LOX and is by the During dioxygenase in a that by of a of a nitrosyl with the of 15-LOX J. PubMed Scopus Google Scholar, PubMed Scopus Google Scholar, J. PubMed Google Scholar). is that ⋅NO with (13S)-HPODE for reaction with 15-LOX flow of ⋅NO either the rate or extent of 15-LOX tryptophan fluorescence quenching was A. Scopus Google Scholar, A. J. PubMed Scopus Google Scholar). native 15-LOX did not ⋅NO and the of ⋅NO inhibition was of (13S)-HPODE was and that ⋅NO the of 15-LOX activity J. PubMed Scopus Google Scholar). of 15-LOX did not of ⋅NO the of ⋅NO inhibition is the ⋅NO was either or enzyme This is not 15-LOX activation and that enzyme inhibition reaction with a dioxygenase 15-LOX did not ⋅NO in the of nitrosyl complexes with 15-LOX metal centers did not This is not a at for the and soybean 15-LOX, in of ⋅NO in and was by reactions J. PubMed Google Scholar). Since formation of required at is a of 15-LOX inhibition that of ⋅NO oxidize native 15-LOX, to either activation or formation of a to activation J. PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar). both occurred formation of the nitrosyl was by of ⋅NO did not 15-LOX. for formation of the that conditions, ⋅NO will than both the formation of complexes and activation of 15-LOX by ⋅NO is During activation by consumption of 2 mol of ⋅NO/mol of 15-LOX was of ⋅NO to by the by reactions of that ⋅NO these Since ⋅NO 15-LOX tryptophan fluorescence ⋅NO uptake activation is to enzyme inhibition ⋅NO at dioxygenase that was not or is of ⋅NO with in is J. J. Scopus Google Scholar, N. 1995; PubMed Scopus Google Scholar, and J. PubMed Google Scholar). Since ⋅NO the of in a reaction of ⋅NO with EredLOO⋅ is ⋅NO with EredLOO⋅ to form LOONO, of product the enzyme (Ered) that by for of the catalytic cycle of soybean 15-LOX is of the rate of linoleate J. J. PubMed Scopus Google activation of rabbit 15-LOX is of the of dioxygenase activity A. J. PubMed Scopus Google Scholar). formation of the enzyme Ered by ⋅NO reaction with EredLOO⋅ will the rate of dioxygenase activity and product (Scheme product of reaction ⋅NO and at in and S. PubMed Scopus Google to form the either to form or to which with of and of of these to is Since of either in the or of ⋅NO and or was of at the 15-LOX to and showed that ⋅NO reaction with soybean 15-LOX of and S. S. M. B. 1995; PubMed Scopus Google Scholar). to that ⋅NO is reaction with 15-LOX intermediates and enzyme did not and formation of and that with ⋅NO to form products Scopus Google Scholar, PubMed Scopus Google Scholar, PubMed Scopus Google Scholar). of with and of which at will to form M. S. 12: PubMed Scopus Google Scholar). that at ⋅NO and 15-LOX in 15-LOX is 15-LOX catalytic activity ⋅NO-dependent activation of soluble guanylate and 15-LOX ⋅NO activation of 15-LOX, 2 mol of ⋅NO/mol of 15-LOX are reaction of ⋅NO with of the dioxygenase leads to reversible enzyme inhibition by formation of the ⋅NO reaction 15-LOX leads to in product of occurred in with consumption of of Since modulation of 15-LOX activity by ⋅NO occurs at ⋅NO of formation and ⋅NO consumption is in for ⋅NO activation of soluble cyclase is these will cGMP production and smooth muscle revealed in consumption of ⋅NO by the lipoxygenase present in a of vascular then to responses to ⋅NO J. PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar, M. J. Proc. Acad. Sci. U. S. A. PubMed Scopus Google Scholar).