[show abstract][hide abstract] ABSTRACT: Fatty acid nitration by nitric oxide-derived species yields electrophilic products that adduct protein thiols, inducing changes in protein function and distribution. Nitro-fatty acid adducts of protein and reduced glutathione (GSH) are detected in healthy human blood. Kinetic and mass spectrometric analyses reveal that nitroalkene derivatives of oleic acid (OA-NO2) and linoleic acid (LNO2) rapidly react with GSH and Cys via Michael addition reaction. Rates of OA-NO2 and LNO2 reaction with GSH, determined via stopped flow spectrophotometry, displayed second-order rate constants of 183 M(-1)S(-1) and 355 M(-1)S(-1), respectively, at pH 7.4 and 37 degrees C. These reaction rates are significantly greater than those for GSH reaction with hydrogen peroxide and non-nitrated electrophilic fatty acids including 8-iso-prostaglandin A2 and 15-deoxy-Delta(12,14)-prostaglandin J2. Increasing reaction pH from 7.4 to 8.9 enhanced apparent second-order rate constants for the thiol reaction with OA-NO2 and LNO2, showing dependence on the thiolate anion of GSH for reactivity. Rates of nitroalkene reaction with thiols decreased as the pKa of target thiols increased. Increasing concentrations of the detergent octyl-beta-d-glucopyranoside decreased rates of nitroalkene reaction with GSH, indicating that the organization of nitro-fatty acids into micellar or membrane structures can limit Michael reactivity with more polar nucleophilic targets. In aggregate, these results reveal that the reversible adduction of thiols by nitro-fatty acids is a mechanism for reversible post-translational regulation of protein function by nitro-fatty acids.
Journal of Biological Chemistry 11/2007; 282(42):31085-93. · 4.65 Impact Factor
[show abstract][hide abstract] ABSTRACT: A bifunctional biaryl lactone has been synthesized that should be capable of iterative unidirectional aryl-aryl bond rotation via: (1) a diastereoselective lactone ring opening, (S)-1 to (P,S)-2 or (M,S)-2; (2) a chemoselective lactonization, (P,S)-2 or (M,S)-2 to (S)-3; and (3) a chemoselective hydrolysis, (S)-3 to (S)-1. Preliminary results of a racemic sample have indicated unidirectional 180 degrees rotation with very high directional selectivity per individual artificial molecular motor molecule through the first two steps of this sequence. [reaction: see text]
[show abstract][hide abstract] ABSTRACT: Unsaturated fatty acids are nitrated endogenously to produce nitrated lipids. Recent studies have shown that these nitrated lipids have high chemical reactivity and profound biological implications. We report an efficient, scalable synthesis which is regiospecific and stereoselective for all possible isomers of nitrated oleic acid: (E)- and (Z)-, 9- and 10-octadec-9-enoic acids.
[show abstract][hide abstract] ABSTRACT: Nitric oxide ((*)NO)-derived reactive species nitrate unsaturated fatty acids, yielding nitroalkene derivatives, including the clinically abundant nitrated oleic and linoleic acids. The olefinic nitro group renders these derivatives electrophilic at the carbon beta to the nitro group, thus competent for Michael addition reactions with cysteine and histidine. By using chromatographic and mass spectrometric approaches, we characterized this reactivity by using in vitro reaction systems, and we demonstrated that nitroalkene-protein and GSH adducts are present in vivo under basal conditions in healthy human red cells. Nitro-linoleic acid (9-, 10-, 12-, and 13-nitro-9,12-octadecadienoic acids) (m/z 324.2) and nitro-oleic acid (9- and 10-nitro-9-octadecaenoic acids) (m/z 326.2) reacted with GSH (m/z 306.1), yielding adducts with m/z of 631.3 and 633.3, respectively. At physiological concentrations, nitroalkenes inhibited glyceraldehyde-3-phosphate dehydrogenase (GAPDH), which contains a critical catalytic Cys (Cys-149). GAPDH inhibition displayed an IC(50) of approximately 3 microM for both nitroalkenes, an IC(50) equivalent to the potent thiol oxidant peroxynitrite (ONOO(-)) and an IC(50) 30-fold less than H(2)O(2), indicating that nitroalkenes are potent thiol-reactive species. Liquid chromatography-mass spectrometry analysis revealed covalent adducts between fatty acid nitroalkene derivatives and GAPDH, including at the catalytic Cys-149. Liquid chromatography-mass spectrometry-based proteomic analysis of human red cells confirmed that nitroalkenes readily undergo covalent, thiol-reversible post-translational modification of nucleophilic amino acids in GSH and GAPDH in vivo. The adduction of GAPDH and GSH by nitroalkenes significantly increased the hydrophobicity of these molecules, both inducing translocation to membranes and suggesting why these abundant derivatives had not been detected previously via traditional high pressure liquid chromatography analysis. The occurrence of these electrophilic nitroalkylation reactions in vivo indicates that this reversible post-translational protein modification represents a new pathway for redox regulation of enzyme function, cell signaling, and protein trafficking.
Journal of Biological Chemistry 08/2006; 281(29):20450-63. · 4.65 Impact Factor
[show abstract][hide abstract] ABSTRACT: Mass spectrometric analysis of human plasma and urine revealed abundant nitrated derivatives of all principal unsaturated fatty acids. Nitrated palmitoleic, oleic, linoleic, linolenic, arachidonic and eicosapentaenoic acids were detected in concert with their nitrohydroxy derivatives. Two nitroalkene derivatives of the most prevalent fatty acid, oleic acid, were synthesized (9- and 10-nitro-9-cis-octadecenoic acid; OA-NO2), structurally characterized and determined to be identical to OA-NO2 found in plasma, red cells, and urine of healthy humans. These regioisomers of OA-NO2 were quantified in clinical samples using 13C isotope dilution. Plasma free and esterified OA-NO2 concentrations were 619 +/- 52 and 302 +/- 369 nm, respectively, and packed red blood cell free and esterified OA-NO2 was 59 +/- 11 and 155 +/- 65 nm. The OA-NO2 concentration of blood is approximately 50% greater than that of nitrated linoleic acid, with the combined free and esterified blood levels of these two fatty acid derivatives exceeding 1 microm. OA-NO2 is a potent ligand for peroxisome proliferator activated receptors at physiological concentrations. CV-1 cells co-transfected with the luciferase gene under peroxisome proliferator-activated receptor (PPAR) response element regulation, in concert with PPARgamma, PPARalpha, or PPARdelta expression plasmids, showed dose-dependent activation of all PPARs by OA-NO2. PPARgamma showed the greatest response, with significant activation at 100 nm, while PPARalpha and PPARdelta were activated at approximately 300 nm OA-NO2. OA-NO2 also induced PPAR gamma-dependent adipogenesis and deoxyglucose uptake in 3T3-L1 preadipocytes at a potency exceeding nitrolinoleic acid and rivaling synthetic thiazo-lidinediones. These data reveal that nitrated fatty acids comprise a class of nitric oxide-derived, receptor-dependent, cell signaling mediators that act within physiological concentration ranges.
Journal of Biological Chemistry 01/2006; 280(51):42464-75. · 4.65 Impact Factor
[show abstract][hide abstract] ABSTRACT: A new radical-sensitive caging technique releases a caged molecule under lipid peroxidation conditions. In a competitive oxidation study with a model lipid, methyl linoleate, the oxidatively releasable xanthenyl caging groups is found to be 1.93+/-0.55 times more reactive than the lipid model compound.
[show abstract][hide abstract] ABSTRACT: The aqueous decay and concomitant release of nitric oxide (*NO) by nitrolinoleic acid (10-nitro-9,12-octadecadienoic acid and 12-nitro-9,12-octadecadienoic acid; LNO2) are reported. Mass spectrometric analysis of reaction products supports a modified Nef reaction as the mechanism accounting for the generation of *NO by the aqueous reactions of fatty acid nitroalkene derivatives. Nitrolinoleic acid is stabilized by an aprotic milieu, with LNO2 decay and *NO release strongly inhibited by phosphatidylcholine/cholesterol liposome membranes and detergents when present at levels above their critical micellar concentrations. The release of *NO from LNO2 was induced by UV photolysis and triiodide-based ozone chemiluminescence reactions currently used to quantify putative protein nitrosothiol and N-nitrosamine derivatives. This reactivity of LNO2 complicates the qualitative and quantitative analysis of biological oxides of nitrogen when applying UV photolysis and triiodide-based analytical systems to biological preparations typically abundant in nitrated fatty acids. The results reveal that nitroalkene derivatives of linoleic acid are pluripotent signaling mediators that act not only via receptor-dependent mechanisms, but also by transducing the signaling actions of *NO via pathways subject to regulation by the relative distribution of LNO2 to hydrophobic versus aqueous microenvironments.
Journal of Biological Chemistry 06/2005; 280(19):19289-97. · 4.65 Impact Factor
[show abstract][hide abstract] ABSTRACT: Ether tethers allow Heck cyclizations to proceed in high yields. Ester tethers lead to low yields. Styrene trapping experiments indicate that ester reactions form viable organopalladium intermediates that cannot cyclize efficiently.
Tetrahedron Letters - TETRAHEDRON LETT. 01/2005; 46(42):7213-7215.
[show abstract][hide abstract] ABSTRACT: High-quality alloxazine (a flavin isomer) imprinted polymers have been made for the first time. A molecularly imprinted polymer (MIP) for the N1,N3-di-functionalized alloxazine template 2 was made. The MIP prepared for 2 exhibited excellent, highly selective molecular recognition for template 2, as determined by HPLC analysis using columns prepared with the MIP. This has also demonstrated that the core flavin structure can survive the imprinting process.
Tetrahedron Letters - TETRAHEDRON LETT. 01/2005; 46(34):5651-5654.
[show abstract][hide abstract] ABSTRACT: Net directed 180° bond rotation was achieved through diastereoselective ring-opening reactions in an achiral biaryl lactone using a chiral nucleophile followed by re-lactonization. The efficiency of the directed bond rotation has been determined by HPLC–MS to be 50% and 20% with two different chiral nucleophiles. These results demonstrate the potential for a prototype of a chemically driven synthetic molecular motor which has the advantages of both simplicity and flexibility in operation and is the first example of the use of a chiral auxiliary to induce transient axial chirality resulting in net directed bond rotation.
[show abstract][hide abstract] ABSTRACT: A class of chiral biaryl molecules that have been designed to undergo unidirectional rotation about the aryl–aryl bond may show promise for future application in the area of synthetic molecular motors. These asymmetric molecules should be capable of channeling chemical energy into repeated 360° rotation in one direction. Herein we report the synthesis and characterization of one such biaryl molecule (1).
Tetrahedron Letters - TETRAHEDRON LETT. 01/2004; 45(52):9599-9602.
[show abstract][hide abstract] ABSTRACT: We report a synthesis of the ammonium salt of the 8-carbon monosaccharide 3-deoxy-(d)-manno-2-octulosonic acid (KDO) starting from (d)-mannose. The key C-C bond construction in our synthetic strategy is a novel alkyl cobaloxime mediated radical alkyl-alkenyl cross coupling of α-ethoxyacrylonitrile with an alkyl cobaloxime derivative of D-mannose. KMnO4 unmasking of the latent functionality in the α-ethoxyacrylonitrile moiety provides the α-hydroxyester moiety necessary for completion of the KDO synthesis.
[show abstract][hide abstract] ABSTRACT: Processing of trans-2-phenylcyclopropylmethanols 5 and 6 by the monocopper/tyrosine radical enzyme galactose oxidase led to mechanism-based inactivation with a partition ratio, (kcat + kinact)kinact, of approximately 1 and a primary deuterium isotope effect, kinact(H)kinact(D), of 3.2. The data are consistent with a radical mechanism for galactose oxidase with a short lived ketyl radical anion intermediate.
Methods in Enzymology 02/2002; 354:415-25. · 2.00 Impact Factor
[show abstract][hide abstract] ABSTRACT: Intermolecular additions of highly enantiomerically enriched episulfonium ions onto activated benzene rings are known to accomplish chiral Friedel–Crafts alkylations. Unactivated benzenes are either unreactive or can give low stereoselectivities. Intramolecular cyclizations should be faster than intermolecular additions; thus, cyclization reactions should be able to avoid undesired episulfonium ion racemization. The work reported here tests that hypothesis and demonstrates that cyclizations of enantiomerically enriched episulfonium ions onto unactivated benzene rings are highly stereoselective.
Tetrahedron Letters - TETRAHEDRON LETT. 01/2002; 43(3):351-353.
[show abstract][hide abstract] ABSTRACT: In previous work we discovered that reductive Pd-mediated aryl-enone cyclization of an advanced intermediate resulted in epimerization of a critical stereocenter. Analysis of conformational and steric effects could rationalize the result. Based on that analysis it was anticipated that a conformational lock could suppress the undesired epimerization. Results reported herein confirm that expectation.
Tetrahedron Letters - TETRAHEDRON LETT. 01/1999; 40(14):2691-2694.
[show abstract][hide abstract] ABSTRACT: N,N′-Bis(3,5-di-tert-butylsalicylidene)-1,2-cyclohexanediaminocobalt(II) (1) catalyzes the reductive deoxygenation of epoxides by Na(Hg) in THF, with 5–10 catalytic turnovers. The simpler N,N′-bis(salicylidene)-ethylenediaminocobalt(II) [Co(Salen)2] (16) failed to catalyze deoygenations in THF but did in DMF.
Tetrahedron Letters - TETRAHEDRON LETT. 01/1999; 40(50):8747-8749.
[show abstract][hide abstract] ABSTRACT: Photochemical cross coupling of alkyl cobaloximes with maleic anhydrides and PhSSPh led to addition of an alkyl moiety and an SPh moiety across the alkene. Oxidation of the sulfide to a sulfoxide followed by elimination of phenyl sulfenic acid provided substituted maleic anhydrides. Alkyl cobaloximes could also be coupled with maleic anhydrides in the absence of PhSSPh to provide substituted maleic anhydrides directly. These methods are applied to a short and efficient synthesis of the anhydride of the Ras farnesyl protein transferase inhibitor chaetomellic acid A and in preparations of C-glycosyl maleic anhydrides, in which the carbohydrate anomeric carbon is directly attached to a maleic anhydride alkene carbon, in modest yields (19−46%).
Journal of Organic Chemistry - J ORG CHEM. 05/1998; 63(11).
[show abstract][hide abstract] ABSTRACT: Studies are reported on the cyclization of (ω-hydroxy-β-hydroxyalkyl)cobaloximes (1, 2 and 3) to form 5, 6 and 7-membered ring cyclic ethers (4, 5 and 6). Reversible cyclization and eventual irreversible alkene decomplexation (to form 7, 8 and 9) varied as a function of ring size. The practical consequence is that cyclizations to form 5 and 6 membered rings are feasible whereas formation of a 7 membered ring is not.
Tetrahedron Letters - TETRAHEDRON LETT. 01/1998; 39(21):3447-3448.
[show abstract][hide abstract] ABSTRACT: Cyclization of (5-N-pyrrolyl-2-hydroxypentyl)cobaloxime (13) proceeds by intramolecular electrophilic aromatic substitution of a cobaloxime π-cation onto the pyrrole ring to provide 6-exo cyclization product (14) in 95% yield. This cyclization is highly enantioselective. It is applied to a synthesis of highly enantioenriched (−)-tashiromine, (−)-21, and a formal synthesis of (+)-tashiromine.