Chain Cleavage Products Research Articles

Detection of Unprecedented CYP74 Enzyme in Mammal: Hydroperoxide Lyase CYP74C44 of the Bat Sturnira hondurensis.

The genome of the neotropical fruit bat Sturnira hondurensis was recently sequenced, revealing an unexpected gene encoding a plant-like protein, CYP74C44, which shares ca. 90% sequence identity with the putative CYP74C of Populus trichocarpa. The preparation and properties of the recombinant CYP74C44 are described in the present work. The CYP74C44 enzyme was found to be active against the 13- and 9-hydroperoxides of linoleic and α-linolenic acids (13-HPOD, 13-HPOT, 9-HPOD, and 9-HPOT, respectively), as well as the 15-hydroperoxide of eicosapentaenoic acid (15-HPEPE). All substrates studied were specifically transformed into chain cleavage products that are typical for hydroperoxide lyases (HPLs). The HPL chain cleavage reaction was validated by the identification of NaBH4-reduced products (Me/TMS) of 15-HPEPE and 13- and 9-hydroperoxides as (all-Z)-14-hydroxy-5,8,11-tetradecatrienoic, (9Z)-12-hydroxy-9-dodecenoic, and 9-hydroxynonanoic acids (Me/TMS), respectively. Thus, CYP74C44 possessed the HPL activity that is typical for the CYP74C subfamily proteins.

Direct and nitrite-sensitized indirect photolysis of effluent-derived phenolic contaminants under UV254 irradiation.

UV254 photolysis has increasingly been utilized for disinfection of water-born pathogens in wastewater. During disinfection, wastewater-derived trace organic contaminants, such as pharmaceuticals and personal care products (PPCPs), may be subjected to direct photolysis and indirect photolysis sensitized by wastewater constituents such as nitrite (NO2-). Herein, we reported the direct photolysis and NO2--sensitized indirect photolysis of four phenolic contaminants commonly observed in wastewaters (i.e., bisphenol A (BPA), acetaminophen (ATP), salbutamol (SAL), and 2,4-dihydroxybenzophenone (BP1)). Spectroscopic characterization and quantum yield measurement were carried out to evaluate the photochemical reactivity of these phenolic compounds. In NO2--sensitized photolysis, the relative contribution of direct and indirect photolysis was quantified by light screening factor calculation and radical quenching studies. The experimental results highlight the important roles of HO˙ and NO2˙ in the NO2--sensitized photolysis of phenolic compounds. A series of intermediate products, including hydroxylated, nitrated, nitrosated, dimerized, and alkyl chain cleavage products, were identified by solid phase extraction (SPE) combined with high-resolution mass spectrometry (HRMS) analyses. On the basis of identified products, the underlying mechanisms and transformation pathways for NO2--sensitized photolysis of these phenolic compounds were elucidated. The second-order rate constants of BPA, SAL, BP1 with NO2˙ were calculated to be 2.25 × 104, 1.35 × 104 and 2.44 × 104 M-1 s-1, respectively, by kinetic modeling. Suwanee River natural organic matter (SRNOM) played complex roles in the direct and NO2--sensitized photolysis of phenolic compounds by serving as a photosensitizer, light screening and radical quenching agent. Wastewater constituents, such as NO3- and EfOM, could accelerate direct and NO2--sensitized photolysis of BPA, SAL, and BP1 in the wastewater matrix. Our results suggest that NO2- at the WWTP effluent-relevant level can sensitize the photolysis of effluent-derived phenolic contaminants during the UV254 disinfection process; however, the formation of potentially carcinogenic and mutagenic nitrated/nitrosated derivatives should be scrutinized.

Photo-induced fragmentation of tyrosine side chains in IgG4-Fc: Effect of protein sequence, conformation and glycan structure

This account summarizes recent work on photo-induced side chain cleavage reactions of Tyr in IgG4-Fc. These processes are initiated via photo-ionization of Trp to a Trp radical cation, followed by one-electron oxidation of Tyr to a Tyr radical cation, where fragmentation of the Cα-Cβ bond leads to an intermediary glycyl radical. The latter converts into Gly via hydrogen atom transfer, or into various backbone cleavage products subsequent to the addition of oxygen. Site-directed mutation of Trp381 to Phe381 significantly reduces the yields of Tyr side chain cleavage products, underlining a critical role of Trp for these reactions. Likewise, conformational changes of IgG4-Fc induced by increasing contents of 1-propanol in the solution diminishes the yields of Tyr side chain cleavage products. Variations of the glycan structure at Asn297 had an effect on Tyr side chain cleavage, where larger glycan structures led to lower yields. This result is in interesting contrast to Trp side chain cleavage in IgG1-Fc, where larger glycan structures led to higher yields. The opposite trends for Trp and Tyr side chain cleavage as a function of glycan structure are consistent with a connection between the precursors for both reactions, radical cations of Trp and Tyr, related by electron transfer. The potential significance of these reactions for protein integrity in formulations of pharmaceutical proteins will be discussed.

Epoxyalcohol synthase activity of the CYP74B enzymes of higher plants

The CYP74B subfamily of fatty acid hydroperoxide transforming cytochromes P450 includes the most common plant enzymes. All CYP74Bs studied yet except the CYP74B16 (flax divinyl ether synthase, LuDES) and the CYP74B33 (carrot allene oxide synthase, DcAOS) are 13-hydroperoxide lyases (HPLs, synonym: hemiacetal synthases). The results of present work demonstrate that additional products (except the HPL products) of fatty acid hydroperoxides conversion by the recombinant StHPL (CYP74B3, Solanum tuberosum), MsHPL (CYP74B4v1, Medicago sativa), and CsHPL (CYP74B6, Cucumis sativus) are epoxyalcohols. MsHPL, StHPL, and CsHPL converted the 13-hydroperoxides of linoleic (13-HPOD) and α-linolenic acids (13-HPOT) primarily to the chain cleavage products. The minor by-products of 13-HPOD and 13-HPOT conversions by these enzymes were the oxiranyl carbinols, 11-hydroxy-12,13-epoxy-9-octadecenoic and 11-hydroxy-12,13-epoxy-9,15-octadecadienoic acid. At the same time, all enzymes studied converted 9-hydroperoxides into corresponding oxiranyl carbinols with HPL by-products. Thus, the results showed the additional epoxyalcohol synthase activity of studied CYP74B enzymes. The 13-HPOD conversion reliably resulted in smaller yields of the HPL products and bigger yields of the epoxyalcohols compared to the 13-HPOT transformation. Overall, the results show the dualistic HPL/EAS behaviour of studied CYP74B enzymes, depending on hydroperoxide isomerism and unsaturation.

Photo-oxidation of IgG1 and Model Peptides: Detection and Analysis of Triply Oxidized His and Trp Side Chain Cleavage Products.

Triply oxidized histidine in an IgG1 monoclonal antibody was noticed when exposed to ICH light conditions. In order to understand the role of light source, irradiation wavelengths and primary sequence, specifically those of a nearby tryptophan, we synthesized and exposed several peptides to ICH light conditions and analyzed the products using LC-MS analysis. Protein and peptide samples were photo-irradiated under ICH conditions as well as with monochromatic light at λ = 254nm and analyzed using either LTQ Orbitrap or a LTQ-FT ion cyclotron resonance mass spectrometer respectively. A triply oxidized His residue was detected along with a second doubly oxidized His residue in an IgG1. Both of these oxidized His residues are located near Trp residues. In order to investigate the role of Trp photosensitization in His oxidation we synthesized model peptides and Ala mutants. Peptides exposed to ICH light stress conditions revealed a small percent of triply oxidized His in the Trp-containing peptide sequences but not in their corresponding Ala mutants. The differences in product formation under different photo-irradiation conditions underline the importance of light source, irradiation wavelengths and primary sequence in the photosensitivity of proteins.

Surface chain cleavage behavior of PBIA fiber induced by direct fluorination

The surface chain cleavage behavior of PBIA fiber induced by direct fluorination was reported based on the analysis of physical and chemical changes on the fiber surface. The chain cleavage product was obtained to evaluate the chemical reaction during the fluorination process, and its impact on composites performance was also involved. DSC, FTIR spectra, UV–vis absorption spectra and H1NMR were utilized to analyze the chemical structure and composition of the chain cleavage product. The results show gaseous fluorine is most likely to attack the benzimidazole and amide bond in PBIA unit, which was also demonstrated by molecular simulation. Owing to the polar groups contained in chain cleavage products, the wettability of epoxy resin to fiber has been improved, leading to an 11.5% increase of adhesive strength of fiber-epoxy composite.

Studies on Geobacillus stearothermophilus-Part V1: Transformation of 17α-hydroxyprogesterone and 21-hydroxyprogesterone

Side chain cleavage by biotransformation of 17α-hydroxyprogesterone 1 and 21-hydroxyprogesterone 2 by the thermophile, Geobacillus stearothermophilus was investigated. The side chain cleavage product, androst-4-ene-3,17-dione 1a & 2a, was a common product formed from both substrates. 17α-hydroxy-5α-pregnane-3,6,20-trione 1b and 17α-20α-dihydroxyprogesterone 1c were formed from 1. A rare 17-carboxylic acid analogue of androstene 2b, a transformation product of 2, was also identified. G. stearothermophilus also facilitated hydroxylation of 17α-hydroxyprogesterone at 6α-and 6β-positions resulting in 6β,17α-dihydroxyprogesterone 1d, 6α,17α-dihydroxyprogesterone 1e, 6β,21-dihydroxyprogesterone 2c and 6α,21-dihydroxyprogesterone 2d. All transformation products were identified through their spectral data and comparison with reference compounds.

Hydroperoxide lyases (CYP74C and CYP74B) catalyze the homolytic isomerization of fatty acid hydroperoxides into hemiacetals

The conversion of linoleic acid 9-hydroperoxide (9-HPOD) by recombinant melon ( Cucumis melo L.) hydroperoxide lyase (HPL, CYP74C subfamily) was studied. Short (5 s–1 min) incubations at 0 °C followed by rapid extraction and trimethylsilylation made it possible to trap a new unstable ( t 1/2 < 30 s) product, i.e. the hemiacetal (1′ E,3′ Z)-9-hydroxy-9-(1′,3′-nonadienyloxy)-nonanoic acid. Identification was performed by GC-MS analysis and substantiated by the formation of trimethylsilyl 9-trimethylsilyloxy-9-nonyloxy-nonanoate upon catalytic hydrogenation and by 2H-labelling experiments. Both 18O atoms of [ 18O 2-hydroperoxy]9-HPOD were incorporated into the hemiacetal. Along with the hemiacetal, three chain-cleavage products, i.e. the enol (1 E,3 Z)-nonadienol and the hydrates of 3( Z)-nonenal and 9-oxononanoic acid, were trapped as their trimethylsilyl derivatives. The kinetics of 18O incorporation from [ 18O 2]9-HPOD provided strong evidence that the cleavage products originated in the hemiacetal. Linolenic and linoleic acid 13-hydroperoxides served as substrates for recombinant HPLs of melon, alfalfa ( Medicago sativa) and guava ( Psidium guajava), and in each case hemiacetals and enols were detectable by the trapping technique. The data obtained demonstrated that CYP74C and CYP74B HPLs act as isomerases performing a homolytic rearrangement of fatty acid hydroperoxides into short-lived hemiacetals which upon decomposition produce 3( Z)-nonenal, 3( Z)-hexenal and other short chain aldehydes.

Chain cleavage and sulfoxidation of thiastearoyl-ACP upon reaction with stearoyl-ACP desaturase.

The fatty acid analogues 9- and 10-thiastearate were converted to acyl-ACP derivatives by in vitro enzymatic synthesis and reacted with the reconstituted soluble stearoyl-ACP Delta9 desaturase complex. Electrospray ionization mass spectral analysis of the acyl chains purified from the reaction mixtures showed that 10-thiastearoyl-ACP was converted to the 10-sulfoxide as the sole product. In the presence of (18)O(2), the sulfoxide oxygen was found to be derived exclusively from O(2). This result confirms the ability of the soluble stearoyl-ACP desaturase to catalyze O atom transfer in the presence of the appropriate substrate analogue. Inhibition studies showed that 10-thiastearoyl-ACP was a mixed-type inhibitor of 18:0-ACP, with an apparent K(I) of approximately 10 microM. Comparable reactions of the stearoyl-ACP desaturase complex with 9-thiastearoyl-ACP gave the 9-sulfoxide as approximately 5% of the total products, with the O atom again exclusively derived from O(2). The remaining 95% of the total products arose from an acyl chain cleavage reaction between S-9 and C-10. Matrix-assisted laser desorption ionization time-of-flight mass spectral analysis showed that 9-thiastearoyl-ACP had a mass of 9443 amu while the acyl chain cleavage product had a mass of 9322 amu, corresponding to the calculated mass of 8-mercaptooctanoyl-ACP. Recovery of the acyl chain from the ACP product gave the disulfide of 8-mercaptooctanoate (mass of 349.1 amu), arising from the dimerization of 8-mercaptooctanoate during product workup. Gas chromatography-mass spectral analysis also showed the accumulation of nonanal in sealed reaction vials, accounting for the other product of the acyl chain cleavage reaction. The reactivity at both the 9 and 10 positions of the thia-substituted acyl-ACPs is consistent with the proximity of both positions to the diiron center oxidant in the enzyme-substrate complex. Moreover, the differential reactivity of the 9- and 10-thiastearoyl-ACPs also suggests position-dependent consequences of the reaction within the Delta9D active site. Mechanisms accounting for both sulfoxidation and acyl cleavage reactions by the stearoyl-ACP Delta9 desaturase are proposed.

ROLE OF THREE SF-1 BINDING SITES IN THE EXPRESSION OF THE MVDP/AKR1-B7 ISOCAPROALDEHYDE REDUCTASE IN Y1 CELLS

Mvdp/akr1-b7 encodes an aldose-reductase-like enzyme expressed in the zona fasciculata of the adrenal cortex, the function of which is essential for the detoxification of the cholesterol side chain cleavage product, isocaproaldehyde. The −510/+41 akr1-b7 promoter fragment is able to reproduce the endogenous gene zona fasciculata restricted, ACTH-controlled expression, in transgenic mice adrenals. Here, we report that three response elements contained within this promoter (positions −102, −458, −503) are able to bind SF-1, the essential regulator of steroidogenesis, although the low affinity site at −503 retains some other specific proteins present in Y1 nuclear extracts. Mutation of the −102 site results in a lowering of the activity of the −510/+41 promoter in Y1 cells, whereas mutation of the −458 site induces a reduction both in the global activity and forskolin sensitivity of the promoter. Interestingly, differential mutations of the −503 site nucleotides either induce an increase or a decrease in the basal and forskolin-induced activity.

Reactions of beta-carotene with cigarette smoke oxidants. Identification of carotenoid oxidation products and evaluation of the prooxidant/antioxidant effect.

Recent intervention trials reported that smokers given dietary beta-carotene supplementation exhibited an increased risk of lung cancer and overall mortality. beta-Carotene has been hypothesized to promote lung carcinogenesis by acting as a prooxidant in the smoke-exposed lung. We have examined the interactions of cigarette smoke with beta-carotene in model systems. Both whole smoke and gas-phase smoke oxidized beta-carotene in toluene to several products, including carbonyl-containing polyene chain cleavage products and beta-carotene epoxides. A major product of the reaction was identified as 4-nitro-beta-carotene, which was formed by nitrogen oxides in smoke. Both cis and all-trans isomers of 4-nitro-beta-carotene were detected. The hypothesis that smoke-driven beta-carotene autoxidation exerts prooxidant effects was tested in a liposome system. Lipid peroxidation in dilinoleoylphosphatidylcholine liposomes exposed to gas-phase smoke was modestly inhibited by the incorporation of 0.1 mol % beta-carotene. Both the lipid soluble antioxidant alpha-tocopherol and the water soluble antioxidant ascorbate were oxidized more slowly by gas-phase smoke exposure in liposomes containing beta-carotene. These data indicate that beta-carotene exerts weak antioxidant effects against smoke-induced oxidative damage in vitro. It is unlikely that a prooxidant effect of beta-carotene occurs under biologically relevant conditions or is responsible for an increased incidence of lung cancer observed in smokers who consume beta-carotene supplements.

Factor VIIa/tissue factor generates a form of factor V with unchanged specific activity, resistance to activation by thrombin, and increased sensitivity to activated protein C.

Factor VIIa, in complex with tissue factor (TF), is the serine protease responsible for initiating the clotting cascade. This enzyme complex (TF/VIIa) has extremely restricted substrate specificity, recognizing only three previously known macromolecular substrates (serine protease zymogens, factors VII, IX, and X). In this study, we found that TF/VIIa was able to cleave multiple peptide bonds in the coagulation cofactor, factor V. SDS-PAGE analysis and sequencing indicated the factor V was cleaved at Arg679, Arg709, Arg1018, and Arg1192, resulting in a molecule with a truncated heavy chain and an extended light chain. This product (FVTF/VIIa) had essentially unchanged activity in clotting assays when compared to the starting material. TF reconstituted into phosphatidylcholine vesicles was ineffective as a cofactor for the factor VIIa cleavage of factor V. However, incorporation of phosphatidylethanolamine in the vesicles had little effect over the presence of 20% phosphatidylserine. FVTF/VIIa was as sensitive to inactivation by activated protein C (APC) as thrombin activated factor V as measured in clotting assays or by the appearance of the expected heavy chain cleavage products. The FVTF/VIIa could be further cleaved by thrombin to release the normal light chain, albeit at a significantly slower rate than native factor V, to yield a fully functional product. These studies thus reveal an additional substrate for the TF/VIIa complex. They also indicate a new potential regulatory pathway of the coagulation cascade, i.e., the production of a form of factor V that can be destroyed by APC without the requirement for full activation of the cofactor precursor.

Human Protein S Cleavage and Inactivation by Coagulation Factor Xa

Human factor Xa specifically cleaves the anticoagulant protein S within the thrombin-sensitive domain. Amino-terminal amino acid sequencing of the heavy chain cleavage product indicates cleavage of protein S by factor Xa at Arg60, a site that is distinct from those utilized by alpha-thrombin. Cleavage by factor Xa is unaffected by the presence of hirudin and is completely blocked by tick-anticoagulant-peptide and D-Glu-Gly-Arg-chloromethyl ketone, the latter two being specific inhibitors of factor Xa. The cleavage requires the presence of phospholipid and Ca2+, and is markedly inhibited by the presence of factor Va. Factor Xa-cleaved protein S no longer possesses its activated protein C-dependent or -independent anticoagulant activity, as measured in a factor VIII-based activated partial thromboplastin time clot assay. The apparent binding constant for protein S binding to phospholipid (Kd approximately 4 nM +/- 1.0) is unaffected by factor Xa or thrombin cleavage, suggesting that the loss of anticoagulant activity resulting from cleavage is not primarily due to the loss of membrane binding ability. Cleavage and inactivation of protein S by factor Xa may be an additional way in which factor Xa exerts its procoagulant effect, after the initial stages of clot formation.

Effect of the side-chain structure on the specificity of beta-oxidation in bile acid biosynthesis in rat liver homogenates

3Alpha, 7alpha, 12alpha-trihydroxy-5beta-cholestan-26-oic acid (C27-5beta-cholestanoic acid) derivatives with different carbon-number side chains were incubated with rat liver 800 g supernatant to study the effect of the side-chain length on the beta-oxidation system in bile acid biosynthesis. The intermediate alpha, beta-unsaturated and beta-hydroxylated bile acids, and the corresponding degradation products, were quantitatively determined by gas chromatography. The longer side-chained derivatives (C28- and C29-5beta-cholestanoic acids) were converted into corresponding sidechain degradation products, and the alpha,beta-unsaturated and beta-hydroxylated intermediates were also produced. On the other hand, the shorter side-chained derivative (C26-5beta-cholestanoic acid) only gave alpha,beta-unsaturated intermediate. The total formation of intermediates and degradation products from corresponding substrates was in the order of C27- > C28- > C29- > C26-5beta-cholestanoic acids. In the case of clofibrate-treated rat liver 800 g supernatant, the formation of intermediates and final degradation products from C28- and C29-5beta-cholestanoic acids increased significantly. These longer side-chained analogues seemed to be subjected to beta-oxidation system(s) induced with clofibrate treatment. The effect of a terminal methyl group in the side chain of 5beta-cholestanoic acid on the oxidation system was also investigated using 3alpha, 7alpha, 12alpha-trihydroxy-27-nor-5beta-cholestanoic acid derivatives as enzymatic substrates. These derivatives gave corresponding side chain degradation products, but the formation of intermediates was not detected. The formation of side chain cleavage products from 27-nor-5beta-cholestanoic acid derivatives increased to 10 to 25-fold that of the controls by treatment with clofibrate. The results suggested that the beta-oxidation system for 27-nor-5beta-cholestanoic acid derivatives was different from that for C27-5beta-cholestanoic acid, despite their bile acid steroidal structure.

A rapid method for profiling the products of antioxidant reactions by negative ion chemical ionization mass spectrometry.

The antioxidant nutrients beta-carotene and alpha-tocopherol are thought to prevent oxidation of important biomolecules in vivo by trapping reactive free radicals. Conventional analytical methods for analyzing antioxidants and their products rely on time consuming chromatographic isolation followed by spectroscopic characterization. This approach is complicated by the instability of many of these products and the large amount of time required for isolation and characterization of multiple products. We have developed a negative ion chemical ionization mass spectrometry method to detect beta-carotene, alpha-tocopherol, and their reaction products. The method involves minimal sample handling and does not require compound isolation. Direct probe negative ion chemical ionization produces molecular anions with little or no fragmentation. Each ion signal in the resulting mass spectrum represents a compound or group of isomeric compounds in the original reaction mixture. Thus, a rapid "snapshot" of the reaction product profile is obtained within seconds. Application of this methodology to the analysis of beta-carotene oxidation in model chemical reaction systems and in microsomal membranes in vitro identified intact chain beta-carotene oxygenation products, carbonyl-containing chain cleavage products, and their oxygenation products. Similar analyses of alpha-tocopherol oxidations identified 8a-substituted tocopherones, alpha-tocopherolquinone and alpha-tocopherolquinone epoxides. Full-scan detection limits for the antioxidants and their products are in the picogram range. This method offers the first means for comprehensively profiling the fate of antioxidants in tissue samples.

Cytotoxicity and genotoxicity of lipid-oxidation products

The autoxidation of unsaturated lipids contained in oils, fats, and food and the endogenous oxidative degradation of membrane lipids by lipid peroxidation result in the formation of a very complex mixture of lipid hydroperoxides, chain-cleavage products, and polymeric material. Experimental animal studies and biochemical investigations lend support to the hypothesis that lipid-oxidation products, ingested with food or produced endogenously, represent a health risk. The oral toxicity of oxidized lipids is unexpectedly low. Chronic uptake of large amounts of such materials increases tumor frequency and incidence of atherosclerosis in animals. 4-Hydroxynonenal, a chain-cleavage product resulting from omega 6 fatty acids, disturbs gap-junction communications in cultured endothelial cells and induces several genotoxic effects in hepatocytes and lymphocytes. Although the concentrations of the aldehyde needed to produce these effects are in the range expected to occur in vivo, their pathological significance is far from clear. Recent findings strongly suggest that in vivo modification of low-density lipoprotein by certain lipid-peroxidation products (eg, 4-hydroxynonenal and malonaldehyde) renders this lipoprotein more atherogenic and causes foam-cell formation. Proteins modified by 4-hydroxynonenal and malonaldehyde were detected by immunological techniques in atherosclerotic lesions.

Produits CD-ROM de l'Organisation mondiale de la propriété intellectuelle (OMPI)

The surface chain cleavage behavior of PBIA fiber induced by direct fluorination was reported based on the analysis of physical and chemical changes on the fiber surface. The chain cleavage product was obtained to evaluate the chemical reaction during the fluorination process, and its impact on composites performance was also involved. DSC, FTIR spectra, UV–vis absorption spectra and H1NMR were utilized to analyze the chemical structure and composition of the chain cleavage product. The results show gaseous fluorine is most likely to attack the benzimidazole and amide bond in PBIA unit, which was also demonstrated by molecular simulation. Owing to the polar groups contained in chain cleavage products, the wettability of epoxy resin to fiber has been improved, leading to an 11.5% increase of adhesive strength of fiber-epoxy composite.

Pharmacological significance of the species differences in bupropion metabolism

Bupropion provided a dose-dependent prevention of tetrabenazine-induced sedation in mice but not rats. Bupropion was extensively metabolized in mice, rats, dogs and man. About 85% of the dose was excreted in urine of rats and man. The predominant metabolites in rat urine were side chain cleavage products of bupropion (m-chlorobenzoic acid) with a minor fraction consisting of basic side chain hydroxylated metabolites. Mice, dogs and man form a major side chain hydroxylated product (BW 306U) which appeared in higher concentration than bupropion in plasma of these species but not rats. The relatively high plasma levels of BW 306U in mice but not rats may account for the species difference in pharmacological response observed with bupropion.

Shared Chemical Properties of Different Murine Thymus-Leukemia Antigens

Immunochemical studies of murine thymus-leukemia antigens (TLA) have confirmed that the subunit structure consists of a 45,000-dalton heavy chain and a beta 2 microglobulin (beta 2m) light chain. Similar structural features are exhibited by the TLA from thymocytes of Tlaa, Tlac, Tlad, and a leukemia cell derived from C57BL/6, a Tlab strain. In addition to the similar subunit structure from the four haplotypes, each TLA shows a similar pattern of trypsin proteolysis. This procedure yields a major heavy chain cleavage product of approximately 37,000 daltons that remains associated with beta 2m and retains most or all of the antigenic determinants of the intact TLA. Evidence is presented that TLA do not exhibit Fc receptor properties, nor do they adsorb to murine leukemia virus antigens under the conditions of isolation for analysis on polyacrylamide gel electrophoresis (PAGE) in sodium dodecyl sulfate (SDS). Taken together these findings strongly support the hypothesis that TLA comprise a family of chemically similar antigens belonging to a structurally and genetically related group that includes H-2D, H-2K, and Qa-2,3.