Surface lysine and tyrosine residues are required for interaction of the major herpes simplex virus type 1 DNA-binding protein with single-stranded DNA. (9/78)

Modification of the herpes simplex virus type 1 major DNA-binding protein (ICP8) with reagents and conditions specific for arginine, lysine, and tyrosine residues indicates that surface lysine and tyrosine residues are required for the interaction of this protein with single-stranded DNA. Modification of either of these two amino acids resulted in a loss and/or modification of binding activity as judged by nitrocellulose filter assays and gel shift. Modification specific for arginine residues did not affect binding within the limits of the assays used. Finally, quenching of the intrinsic tryptophan fluorescence of ICP8 in the presence of single-stranded DNA either suggests involvement of this amino acid in the binding reaction or reflects a conformational change in the protein upon binding.  (+info)

Syntheses of DNA adducts of two heterocyclic amines, 2-amino-3-methyl-9H-pyrido[2,3-b]indole (MeAalphaC) and 2-amino-9H-pyrido[2,3-b]indole (AalphaC) and identification of DNA adducts in organs from rats dosed with MeAalphaC. (10/78)

2-Amino-3-methyl-9H-pyrido[2,3-b]indole (MeAalphaC) and 2-amino-3-methyl-9H-pyrido[2,3-b]indole (AalphaC) are mutagenic and carcinogenic heterocyclic amines formed during ordinary cooking. MeAalphaC and AalphaC are activated to mutagenic metabolites by cytochrome P450-mediated N-oxidation to the corresponding N2-OH derivatives. The proximate mutagenic N2-OH derivatives of MeAalphaC and AalphaC did not react with deoxynucleosides or DNA. However, upon acetylation with acetic anhydride both reacted with 2'-deoxyguannosine and 3'-phospho-2'-deoxyguanosine, resulting in one adduct each, but not with other nucleosides or nucleotides. The adducts were identified as N2-(2'-deoxyguanosin-8-yl)-MeAalphaC, N2-(2'-deoxyguanosin-8-yl)-AalphaC, N2-(3'-phospho-2'-deoxyguanosin-8-yl)-MeAalphaC and N2-(3'-phospho-2'-deoxyguanosin-8-yl)-AalphaC by comparison with adducts of known structure obtained by reaction of the parent amines with acetylated guanine N3-oxide. N2-OH-MeAalphaC and N2-OH-AalphaC reacted with calf thymus DNA after addition of acetic anhydride. 32P-postlabelling analysis of modified DNA showed one major adduct co-migrating with N2-(3',5'-diphospho-2'-deoxyguanosin-8-yl)-MeAalphaC and N2-(3',5'-diphospho-2'-deoxyguanosin-8-yl)-AalphaC, respectively. Some minor adducts presumed to be undigested oligomers were also detected. 32P-postlabelling analysis of DNA from several organs of rats dosed orally with MeAalphaC showed that in vivo N2-(2'-deoxyguanosin-8-yl)-MeAalphaC also was the major adduct formed. Relative adduct level in DNA isolated from the liver of the rats was about 50.40 adducts/10(9) nt. The adduct levels were approximately 4-fold lower in the colon and the heart and approximately 12-fold lower in the kidney of the rats.  (+info)

Inhibitors of sterol synthesis. A highly efficient and specific side-chain oxidation of 3 beta-acetoxy-5 alpha-cholest-8(14)-en-15-one for construction of metabolites and analogs of the 15-ketosterol. (11/78)

As part of a program directed towards the chemical syntheses of potential metabolites and analogs of 3 beta-hydroxy-5 alpha-cholest-8(14)-en-15-one (I), a potent regulator of cholesterol metabolism, several routes have been explored for the preparation of 3 beta-hydroxy-15-keto-5 alpha-chol-8(14)-en-24-oic acid (IV). These investigations led to a remarkably specific and efficient side-chain oxidation of I. For example, treatment of the acetate of I with a mixture of trifluoroacetic anhydride, hydrogen peroxide, and sulfuric acid for 3.5 h at -2 degrees C gave a crude product consisting of 3 beta-acetoxy-24-trifluoroacetoxy-5 alpha-chol-8(14)-en-15-one (XI), 3 beta-acetoxy-24-hydroxy-5 alpha-chol-8(14)-en-15-one (XII), and 3 beta, 24-diacetoxy-5 alpha-chol-8(14)-en-15-one (XIII) in yields of 58%, 8%, and 3%, respectively, by HPLC analysis. XI was readily hydrolyzed to XII upon treatment with triethylamine in methanol at room temperature. Oxidation of XII with Jones reagent gave 3 beta-acetoxy-15-keto-5 alpha-chol-8(14)-en-24-oic acid (XVIII) from which its methyl ester (IX) was prepared by treatment with diazomethane. Mild alkaline hydrolysis of XVIII gave the 3 beta-hydroxy-delta 8(14)-15-keto C24 acid (IV). Hydrolysis of the crude product of the side-chain oxidation with K2CO3 in methanol gave 3 beta,24-dihydroxy-5 alpha-chol-8(14)-en-15-one (XIV) which was oxidized with Jones reagent to yield 3,15-diketo-5 alpha-chol-8(14)-en-24-oic acid (XV). Treatment of XV with diazomethane gave its methyl ester (XVI) which, upon controlled reduction with NaBH4, yielded methyl 3 beta-hydroxy-15-keto-5 alpha-chol-8(14)-en-24-oate (XVII). Compound IX was also prepared by an independent route. Full 1H and 13C NMR assignments are presented for 12 new compounds. IV caused a approximately 56% reduction of the level of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity in CHO-K1 cells at a concentration of 2.5 microM. In contrast, the corresponding 3,15-diketo acid XV had no detectable effect on reductase activity under the same conditions.  (+info)

Cyclization of 1-benzyl-1,2-dihydro-2-(substituted methylene)quinolines to pyrrolo[1,2-a]quinoline derivatives. (12/78)

1-Alkyl-2-alkylthioquinolinium salts were prepared from 1-alkyl-2(1H)-quinolones via 1-alkyl-2(1H)-thioquinolones in two steps. Under mild conditions, the reaction of 1-alkyl-2-alkylthioquinolinium iodides with active methylene compounds in the presence of sodium hydride afforded 1-alkyl-1,2-dihydro-2-(substituted methylene)quinolines in good yields. The cyclization of 1-benzylquinolines using acetic anhydride produced the corresponding pyrrolo[1,2-a]quinoline derivatives.  (+info)

Solid-phase column chromatographic and gas chromatographic-mass spectrometric determination of heptaminol in human urine and related pharmacokinetic profiles. (13/78)

Heptaminol is an antihypotensive drug and is one of the stimulants banned in sport competitions. When heptaminol was fortified to a drug-free urine sample and subjected to solid-phase extraction, trifluroacetic anhydride derivatization, and gas chromatography-mass spectrometry analysis, the results indicated three chromatographic peaks, with one major peak [peak 1 (P1) as heptaminol-2TFA], appearing at retention time 7.17 min, and two minor peaks [peak 2 (P2) and peak 3 (P3) as heptaminol-TFA], appearing at RT 5.87 and 5.81 min, respectively. The characteristic ions of peak mass spectra were m/z 322, 224, and 140 for P1, m/z 223 (molecular ion), 208, 140, and 110 for P2, and m/z 208, 140, and 110 for P3. The urine samples collected from healthy male volunteers who orally ingested a single dose (100 mg) of heptaminol were similar to the analytical results shown in the heptaminol-spiked control urine samples. This result suggested that the unchanged heptaminol was the sole form found in urine. The unchanged parent compound was completely eliminated in urine within 24 h and an average of approximately 97% of the dose was excreted through the renal pathway.  (+info)

Modification of potassium channel kinetics by amino group reagents. (14/78)

We have examined the actions of several amino group reagents on delayed rectifier potassium channels in squid giant axons. Three general classes of reagents were used: (1) those that preserved the positive charge of amino groups; (2) those that neutralize the charge; and (3) those that replace the positive with a negative charge. All three types of reagents produced qualitatively similar effects on K channel properties. Trinitrobenzene sulfonic acid (TNBS) neutralizes the peptide terminal amino groups and the epsilon-amino group of lysine groups. TNBS (a) slowed the kinetics of macroscopic ionic currents; (b) increased the size of ionic currents at large positive voltages; (c) shifted the voltage-dependent probability of channel opening to more positive potentials but had no effect on the voltage sensitivity; and (d) altered several properties of K channel gating currents. The actions of TNBS on gating currents suggest the presence of multiple gating current components. These effects are not all coupled, suggesting that several amino groups on the external surface of K channels are important for channel gating. A simple kinetic model that considers the channel to be composed of independent heterologous subunits is consistent with most of the modifications produced by amino group reagents.  (+info)

The synthesis, structure and properties of N-acetylated derivatives of ethyl 3-amino-1H-pyrazole-4-carboxylate. (15/78)

Ethyl 3-amino-1H-pyrazole-4-carboxylate (1) was yielded through total synthesis and reacted with acetic anhydride to give the acetylated products 2-6. Compounds 1-6 were studied with HPLC, X-ray, FT-IR, (1)H-NMR, (13)C-NMR and MS. Acetylation was carried out in solvents of various polarity, namely; chloroform; dioxane; DMF; acetic anhydride, at room temperature and at boiling points; and in the presence and absence of DMAP. The acetylated products are mainly nitrogen atoms in the ring. The position of the ring proton in the solution was based on NOESY; multinuclear HMBC, HSQC spectra and calculations. For equivalent amounts (1-1.5 mol) of acetic anhydride at room temperature two products of monoacetylation are produced in the ring: 2 and 3, ca. 2 : 1 and at the same time only small amount of the third product of monoacetylated, 5 in DMF, as well the product diacetylated, 4. The greatest amount of the product 4 is produced during the reaction with chloroform. However, in this solvent and in dioxane no product 5 is produced. Compound 2 is, largely, formed in dimethylformamide, in the presence DMAP, 0.2 eq. In the presence of this catalytic base, for the first hour, there is a mixture 2 and 3 to the ratio ca. 95 : 5. With 8 eq of Ac(2)O at reflux, after another hour, the compounds 3, 4 and 6 appear about equal amounts. After a longer time, the compound, which appears most in this mixture is triacetylated derivative 6. The structural and spectroscopic characteristics of compounds 1-6 have been given and the methods for their preparation have been provided.  (+info)

Tissue distribution, intracellular localization, and in vitro expression of bovine macrophage scavenger receptors. (16/78)

Macrophage scavenger receptors are trimeric membrane proteins implicated in the pathologic deposition of cholesterol in atherogenesis. The authors have studied the tissue distribution and intracellular localization of bovine scavenger receptors using monoclonal antibovine receptor antibody IgG-D2. The receptor proteins were detectable in macrophages of various organs and tissues, particularly Kupffer cells, alveolar macrophages, and macrophages in the spleen and lymph nodes. In the brain, perivascular macrophages were immunoreactive with IgG-D2. Fibroblasts, endothelial cells, smooth muscle cells, and dendritic cells such as epidermal Langerhans cells, interdigitating cells, or follicular dendritic cells, however, showed no immunoreactivity to IgG-D2. Immunoelectron microscopy showed localization of reaction products for these receptors on the cell surface, vesicles, and endosomes of macrophages. Transient expression of bovine scavenger receptors on cultured cells shows that scavenger receptors are mainly expressed in the endoplasmic reticulum, nuclear envelope, and Golgi apparatus of nonmacrophage cells and moved to the cell surface and endosomes of macrophagelike cells. These results indicate that efficient intracellular transport of scavenger receptors in macrophages is mediated by a macrophage-specific transport system.  (+info)