Oxidative degradation of thymine with O2 promoted by L-ascorbic acid and Cu(II) ion

Oxidation of thymine with O2 was promoted by copper(I) ion generated from reaction of L-ascorbic acid (AA) with copper (II) ion. The main oxidation products were thymine glycol (TG) and N-formyl-N'-pyruvylurea (FPU). At higher concentration of O2, formation of FPU was favored, while TG was dominant at higher Cu(II) ion and lower O2 concentrations. Reaction mechanism involving hydroxy thyminyl radical was proposed.     

High performance liquid chromatography of nucleic acids and the related compounds on a fluorinated silica gel column.

Separations of nucleic acids and the related compounds were investigated by HPLC on a new fluorinated bonded silica gel column. Polyadenylate (Poly (A)) enzymatic partial hydrolysate sample and the mixture of various polynucleotide samples were sufficiently separated by the reversed-phase mode using a gradient elution with aqueous ammonium acetate/acetonitrile system. Mixed-mode separation on the fluorinated bonded phase coated with a tert-alkylammonium salt was also examined for the separation of the various polynucleotides including tRNAs.     

Mutagenicity of ptaquiloside, the carcinogen in bracken, and its related illudane-type sesquiterpenes II. Chromosomal aberration tests with cultured mammalian cells

The chromosomal aberration test using a Chinese hamster lung cell line (CHL) was carried out on ptaquiloside and its related compounds, hypoloside B, hypoloside C, illudin M and illudin S. Ptaquiloside induced chromosomal aberrations at doses as low as 4.5 μg/ml (0.0113 mM). The clastogenic effect was ph-dependent. The same activity was observed at a 90-fold higher dose at pH 5.3 in the culture medium compared with the activity at pH 74. or pH 8.0. Both hypoloside B and hypoloside C were also clastogenic at almost the same dose levels as that of ptaquiloside. Illudin M and illudin S were also potet clastogens and induced aberrations at much lower doses than ptaquiloside. These results suggest that the clastogenic effect is involved in the mechanism of carcinogenic potency of ptaquiloside in animals.     

The low C5 convertase activity of the C4A6 allotype of human complement component C4.

We have compared the C5-convertase-forming ability of different C4 allotypes, including the C4A6 allotype, which has low haemolytic activity and which has previously been shown to be defective in C5-convertase formation. Recent studies suggest that C4 plays two roles in the formation of the C5 convertase from the C3 convertase. Firstly, C4b acts as the binding site for C3 which, upon cleavage by C2, forms a covalent linkage with the C4b. Secondly, C4b with covalently attached C3b serves to form a high-affinity binding site for C5. Purified allotypes C4A3, C4B1 and C4A6 were used to compare these two activities of C4. Covalently linked C4b-C3b complexes were formed on sheep erythrocytes with similar efficiency by using C4A3 and C4B1, indicating that the two isotypes behave similarly as acceptors for covalent attachment of C3b. C4A6 showed normal efficiency in this function. However, cells bearing C4b-C3b complexes made from C4A6 contained only a small number of high-affinity binding sites for C5. Therefore a lack of binding of C5 to the C4b C3b complexes is the reason for the inefficient formation of C5 convertase by C4A6. The small number of high-affinity binding sites created, when C4A6 was used, were tested for inhibition by anti-C3 and anti-C4. Anti-C4 did not inhibit C5 binding, whereas anti-C3 did. This suggests that the sites created when C4A6 is used to make C3 convertase may be C3b-C3b dimers, and hence the low haemolytic activity of C4A6 results from the creation of low numbers of alternative-pathway C5-convertase sites.     

A newly established human acute lymphoblastic leukemia cell line with characteristics of the earliest B-cell maturation

A new human acute lymphoblastic leukemia (ALL) cell line, designated HBL-3, was established from the bone marrow of a patient with non-T-ALL. The HBL-3 cell line expressed B4 (CD 19), BA-1 (CD 24) and HLA-DR antigens, but not surface immunoglobulin (SIg) or cytoplasmic immunoglobulin (CIg). The cell line lacked the common acute lymphoblastic leukemia antigen (CALLA) and antigenic markers characteristic of T-cell and myeloid cell lineages. The HBL-3 cells had structural rearrangements of both the homologous chromosome 9s, including a translocation with chromosome 1 which has been reported in a patient with common ALL. The cell line had rearranged immunoglobulin heavy chain genes but retained germ-line κ light chain genes and germ-line T-cell receptorβ- and γ-chain genes. The HBL-3 cell line was strongly positive for terminal deoxynucleotidyl transferase (TdT). These findings indicate that the HBL-3 cell line is derived from the earliest B-cell committed to B-cell lineage.     

Structures of the tetrahydrogenated menaquinones fromActinomadura angiospora,Faenia rectivirgula,andSaccharothrix australiensis

The structures of the tetrahydrogenated menaquinones fromActinomadura angiospora, Faenia rectivirgula, andSaccharothrix australiensis were determined by mass spectrometry and proton nuclear magnetic resonance spectrometry. The positions of saturation of the tetrahydrogenated menaquinones fromFaenia rectivirgula andSaccharothrix australiensis were units II plus III (counting from the ring system), whereas that ofActinomadura angiospora had units III and VIII hydrogenated. The tetrahydrogenated menaquinones fromFaenia rectivirgula andSaccharothrix australiensis are similar to those characterized from other Gram-positive taxa to date, whereas that fromActinomadura angiospora represents a hitherto unknown isomer.     

C5 convertase of the alternative complement pathway: covalent linkage between two C3b molecules within the trimolecular complex enzyme

C5 convertase of the alternative C pathway is a complex enzyme consisting of three C fragments--one molecule of a major fragment of factor B (Bb) and two molecules of a major fragment of C3 (C3b). Within this C3bBbC3b complex, the first C3b binds covalently to the target surface, and Bb, which bears a catalytic site, binds noncovalently to the first C3b. In the present investigation, we studied the nature of the convertase that is assembled on E surfaces and obtained evidence that the second C3b binds directly to the alpha'-chain of the first through an ester bond rather than to the target surface. Thus, the alternative pathway C5 convertase could be described as a trimolecular complex in which Bb binds noncovalently to a covalently linked C3b dimer. We also obtained evidence that not only the second C3b but also the first C3b participates in binding C5, that is, covalently-linked C3b dimer acts as a substrate-binding site. Because of this two-site binding, the convertase has a much higher affinity for C5 than the surrounding monomeric C3b molecules. Based on this evidence, a new model of the alternative pathway C5 convertase is proposed. Covalent association of two subunits and the bivalent binding of the substrate are then common properties of the alternative and classical pathway C5 convertases.     

Monoclonal antibodies to mouse complement receptor type 1 (CR1). Their use in a distribution study showing that mouse erythrocytes and platelets are CR1-negative

mAb to murine C receptor type 1 (CR1) were produced and three of them were characterized. One antibody, designated as 8C12, immunoprecipitated a protein of 190,000 Mr from a detergent extract of surface-labeled spleen cells and stained spleen B but not T lymphocytes in fluorescent flow cytometry. It inhibited both CR1-mediated rosette formation and the cofactor activity of CR1 for factor I-mediated cleavage of C3b, suggesting that it recognizes the ligand-binding site of CR1. The two other antibodies, designated as 7G6 and 7E9, recognized different epitopes from that recognized by 8C12, and they cross-reacted with a protein of 150,000 Mr that is present in a spleen extract. The distribution of CR1 in murine hemopoietic cells was studied by binding experiments with radiolabeled 8C12 and fluorescent flow cytometry. When CR1 was not detected by 8C12 alone, the two other antibodies were used in combination with 8C12 to confirm the negative results. Almost all B lymphocytes from the spleen, lymph nodes, and peripheral blood were CR1 positive. Most of the Thy-1-positive lymphocytes from these tissues were CR1 negative. Thymus lymphocytes were also CR1 negative. Peritoneal macrophages and chemotactic factor stimulated but not unstimulated peripheral blood granulocytes were CR1 positive. In contrast to human E, mouse E were CR1 negative. This pattern of distribution was consistent with previous results obtained by rosette assays. Although mouse platelets cause immune adherence hemagglutination with C3b-bearing SRBC, they are CR1 negative. Three other lines of evidence also indicated that platelets are CR1 negative. First, no band of CR1 was demonstrated by immunoprecipitation with 8C12 of an extract of surface-labeled platelets. Second, 8C12, which inhibited rosette formation by lymphocytes, alone or in combination with 7G6 and 7E9, did not inhibit immune adherence between platelets and C3b-bearing SRBC. Third, polyclonal rabbit IgG prepared from anti-mouse CR1 antiserum did not inhibit immune adherence by platelets. These results strongly suggest that the C3b-binding factor(s) on mouse platelets is different from CR1 and that processing of C3b-bearing immune complexes in mouse blood may be mediated by a new and as yet unidentified C3b-binding factor(s).     

Ultrastructure of spermatogenesis in the sea star,Asterina minor

The ultrastructural features of spermatogenesis were investigated in the hermaphroditic sea star Asterina minor. The primordial germ cells in the genital rachis contain small clusters of electron-dense material (nuage material) and a stack of annulate lamellae. They also have a flagellum and basal body complex situated close to the Golgi complex. After the development of the genital rachis into the ovotestis, spermatogenic cells increase in number and differentiation begins. Nuage material is observed in spermatogonia, but it gradually disappears in spermatocytes. The annulate lamellae do not exist beyond the early spermatogonial stage. By contrast, a flagellum and basal body complex are found throughout spermatogenesis. The Golgi-derived proacrosomal vesicles appear in the spermatocyte and coalesce to form an acrosomal vesicle in the early spermatid. The process of acrosome formation is as follows: (1) a lamella of endoplasmic reticulum (ER) continuous with the outer nuclear membrane encloses the posterior portion of the acrosomal vesicle; (2) the vesicle attaches to the cell membrane with its anterior portion; (3) periacrosomal material accumulates in the space between the acrosomal vesicle and the ER; (4) the nucleus proper changes its features to surround the acrosome; (5) amorphous, electron-dense material is deposited under the electron-dense disk; and (6) the nucleus forms a hollow opposite the electron-dense material.