Synthesis and Use of a Novel Biotinylated Probe for the Chemiluminescent Detection of Proteins That Bind 15-Hydroxyeicosatetraenoic Acid

Kang, Li-Ta; Vanderhoek, Jack Y.

doi:10.1006/abio.1997.2202

Cited by 4 publications

(4 citation statements)

References 14 publications

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“…The synthesis of biotinylated OA, LA, OA-NO 2 , and LNO 2 was performed as described for the synthesis of biotinylated (15S)-hydroxyeicosatetraenoic acids (16). Briefly, 1-methylpiperidine (5.3 l, 48 mol) was added to a solution of either OA, LA, OA-NO 2 , or LNO 2 (15 mg in 1.5 ml of methylene chloride).…”

Section: Fatty Acid Biotinylationmentioning

confidence: 99%

Nitrated Fatty Acids: Endogenous Anti-inflammatory Signaling Mediators

Cui

Schöpfer

Zhang³

et al. 2006

Journal of Biological Chemistry

319

371

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Reactive oxygen species and NO-derived oxidizing, nitrosating and nitrating products mediate diverse cell signaling and pathologic processes in cardiovascular, pulmonary, and neurodegenerative diseases (1, 2). These reactive inflammatory mediators chemically modify carbohydrates, DNA bases, amino acids, and unsaturated fatty acids to form oxidized, nitrosated and nitrated derivatives. For example, accumulation of inflammatory-induced protein tyrosine nitration products represents a shift from the physiological signal-transducing actions of NO to an oxidative, nitrative, and potentially pathogenic pathway (1).Recently, it has been reported that nitration products of unsaturated fatty acids (nitroalkenes) are formed via NO-dependent oxidative reactions (3-5). These derivatives were initially viewed to be, like nitrotyrosine, a "footprint" of NO-dependent redox reactions (3, 6). More recently, we have observed that nitrolinoleate (LNO 2 ) 7 mediates pluripotent cell signaling actions, since it induces relaxation of phenylephrine-preconstricted rat aortic rings, inhibits thrombin-induced Ca 2ϩ elevations and aggregation of human platelets, and attenuates human neutrophil superoxide generation, degranulation, and integrin expression. These cell responses are mediated by both cGMP-and cAMP-dependent and -independent mechanisms (7-9).LNO 2 positional isomers, including 9-, 10-, 12-, and 13-nitro-9,12-cis-octadecadienoic acids, have been identified as free acids in human plasma and red blood cells and as esterified components of plasma lipoproteins and red blood cell membranes (10). In addition, plasma and red cell free and esterified nitrooleate (OA-NO 2 , isomers 9-and 10-nitro-9-cis-octadecenoic acid) was also identified in healthy human blood (11).Current knowledge indicates that enzymatically oxidized unsaturated fatty acid-derived products, such as prostaglandins, thromboxanes, leukotrienes, epoxyeicosatrienoic acids, hydroxyeicosatetraenoic acids, lipoxins, and resolvins serve as lipid mediators or autacoids. These signaling mediators act within a local microenvironment to orchestrate both physiological and pathological events, including platelet aggregation, constriction of vascular smooth muscle, neonatal development, wound healing, and resolution of inflammation (12, 13). In this context, endogenous nitrated fatty acid derivatives, such as * This work was supported by National Institutes of Health Grants HL068878, HL075397, and S06GM08248 (to Y. E. C.), HL70146 (to R. P. P.), and HL58115 and HL64937 (to B. A. F.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. □ S The on-line version of this article (available at http://www.jbc.org) contains supplemental Table S1, Fig. 1, and Movie 1. 1 Supported by American Diabetes Association Grant JFA 7-05-JF-12. 2 These authors contributed equally to this work. 3 Supported by the postdoctoral f...

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Section: Fatty Acid Biotinylationmentioning

confidence: 99%

Nitrated Fatty Acids: Endogenous Anti-inflammatory Signaling Mediators

Cui

Schöpfer

Zhang³

et al. 2006

Journal of Biological Chemistry

319

371

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“…Also, lipid hydroperoxides may be decomposed by antioxidants to produce DNA-reactive electrophiles [41]. Relevant to this study, biotinylated derivatives of 15-HETE have been shown to form complexes with proteins that are stable enough to withstand SDS-PAGE and electrophoretic transfer [42]. This suggests that some lipid-protein associations may be very stable.…”

Section: Discussionmentioning

confidence: 93%

Capturing Proteins that Bind Polyunsaturated Fatty Acids: Demonstration Using Arachidonic Acid and Eicosanoids

Brock

2007

Lipids

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Polyunsaturated fatty acids (PUFA) and their biological derivatives, including the eicosanoids, have numerous roles in physiology and pathology. Although some eicosanoids are known to act through receptors, the molecular actions of many PUFA remain obscure. As the three-dimensional structure of eicosanoids allows them to specifically bind and activate their receptors, we hypothesized that the same structure would allow other proteins to associate with PUFA and eicosanoids. Here, we demonstrate that biotinylation of arachidonic acid and its oxygenated derivatives 5-hydroxyeicosatetraenoic acid (5-HETE) and leukotriene (LT) B(4) can be used to pull down associated proteins. Separation of proteins by two-dimensional gel electrophoresis indicated that a large number of proteins bound each lipid and that proteins could distinguish between two enantiomers of 5-HETE. Individual proteins, identified by matrix assisted laser desorption/ionization-time of flight mass spectrometry, included proteins that are known to bind lipids, including albumin and phosphatidylethanolamine-binding protein, as well as several novel proteins. These include cytoskeletal proteins, such as actin, moesin, stathmin and coactosin-like protein, and G protein signaling proteins, such as Rho GDP dissociation inhibitor 1 and nucleoside diphosphate kinase B. This method, then, represents a relatively simple and straightforward way to screen for proteins that directly associate with, and are potentially modulated by, PUFA and their derivatives.

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“…Linoleic acid, oleic acid, stearic acid, A23187, phenylsulfonyl fluoride, and bacitracin were bought from Sigma (St. Louis, MO). 15(S)-HETE, 15(S)-HETE methyl ester, and 15(S)-HETE biotin hydrazide were prepared as previously described (7,10,11 The preparation of cytosol fractions of RBL-1 cells, the 5lipoxygenase inhibition assay utilizing RBL-1 homogenates and the binding studies involving cytosolic fractions, fatty acids, and [ 3 H]15(S)-HETE were carried out by previously described procedures (9).…”

Section: Methodsmentioning

confidence: 99%

“…The beads were washed three times with phosphate-buffered saline and then treated with 200 l citric acid (pH 2) followed by removal of the beads. The resulting supernatant protein solution was neutralized and analyzed by SDS-PAGE and Western blotting as previously described (11). The 43 kD protein was then excised and collected for amino acid sequence analysis.…”

Section: Methodsmentioning

confidence: 99%

Mono (S) hydroxy fatty acids: novel ligands for cytosolic actin

Kang

Vanderhoek

1998

Journal of Lipid Research

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The ubiquitous hydroxylated fatty acids derived from arachidonic acid (HETEs) or linoleic acid (HODEs) exhibit diverse biological effects including chemotaxis, cell proliferation, and modulation of several enzymatic pathways, including the 5-lipoxygenase leading to the inflammatory leukotrienes. It was observed that 12(S)-and 15(S)-HETE and 13(S)-HODE (12-and 15-lipoxygenase-derived metabolites, respectively) inhibited the 5-lipoxygenase present in rat basophilic leukemia (RBL -1) cell homogenates whereas the 15(R) chiral enantiomer and the nonhydroxylated linoleic, oleic, and stearic acids were either less potent or ineffective. In examining the mechanism of this inhibition, the relative effectiveness of several fatty acids in displacing [ 3 H]15-HETE bound to cytosol preparations were compared and the results indicated that these (S) hydroxy fatty acids and 5(S)-HETE were significantly more potent than either the 15(R) enantiomer, 15(S)-HETE methyl ester, arachidonic acid, or prostaglandin F 2 ␣ . In order to identify the protein(s) that specifically binds HETEs, 15(S)-HETE biotin hydrazide was used as a probe to detect any HETEprotein complexes as this compound both inhibited the 5lipoxygenase and interfered with the binding of [ 3 H]15-HETE to cytosol preparations. SDS-PAGE analysis and chemiluminescent detection revealed that the major cytosolic proteins that bound this biotinylated probe had molecular masses of 43 and 51 kD. Fatty acid competition experiments indicated that the order of effectiveness in displacing this probe from these proteins was 13(S)-HODE Ͼ 5(S)-HETE Ϸ 15(S)-HETE Ͼ Ͼ stearic acid Ϸ arachidonic acid Ϸ 15(R)-HETE. Amino acid sequence analysis showed that the 43 kD protein was actin. These findings suggest the possibility that actin may play a major role in the biological effects of monohydroxylated metabolites derived from cellular 5-, 12-, and 15-lipoxygenases.-Kang, L-T., and J. Y. Vanderhoek. Mono (S) hydroxy fatty acids: novel ligands for cytosolic actin.

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Synthesis and Use of a Novel Biotinylated Probe for the Chemiluminescent Detection of Proteins That Bind 15-Hydroxyeicosatetraenoic Acid

Cited by 4 publications

References 14 publications

Nitrated Fatty Acids: Endogenous Anti-inflammatory Signaling Mediators

Nitrated Fatty Acids: Endogenous Anti-inflammatory Signaling Mediators

Capturing Proteins that Bind Polyunsaturated Fatty Acids: Demonstration Using Arachidonic Acid and Eicosanoids

Mono (S) hydroxy fatty acids: novel ligands for cytosolic actin

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