Enzymatic desulfurization of dibenzothiophene by a cell-free system of Rhodococcus erythropolis D-1

Abstract Pseudomonas syringae cells were exposed to Cu²⁺ alone or in the precence of acetate, proline or cysteine, at concentrations that reduced free Cu²⁺ to 1/10 of the total copper. Ligand concentrations (designated as isoeffective) were determined experimentally using a Cu²⁺-selective electrode and confirmed by computer calculations using published stability constants. Exposure of P. syringae cells to Cu²⁺ alone resulted in rapid and pronounced cell death, and binding of most of the copper in solution. The addition of acetate, proline or cysteine, a few minutes after Cu²⁺ treatment, resulted in a significant reduction in cell death, and in the amount of copper bound to the cells. For short exposures to Cu²⁺, cysteine was more effective than acetate or proline, but after 60 min of treatment, similar results were observed with these ligands. The addition of ligands before Cu²⁺ resulted in even more reduced copper toxicity. The results showed that, at isoeffective concentrations, weak and moderate copper-ligands can effectively antagonize copper toxicity, and that this protective effect does not require previously equilibrated copper-ligand solutions and is not very dependent of the nature of the ligand.     

In vitro mutated beta subunits from the F1-ATPase of the thermophilic bacterium, PS3, containing glutamine in place of glutamic acid in positions 190 or 201 assembles with the alpha and gamma subunits to produce inactive complexes

Using site-directed mutagenesis, Glu-190 or Glu-201 of the beta subunit of the F1-ATPase from the thermophilic bacterium PS3 were replaced with glutamine. It was possible to reconstitute complexes of the mutated beta subunits with alpha and gamma subunits, but the complexes did not have ATPase activity. It is concluded that carboxylic acid side chains of Glu-190 and Glu-201 of the beta subunit are essential for catalytic activity of F1-ATPase.     

NADPH production from NADP+ with a glucose dehydrogenase system involving whole cells and immobilized cells of Gluconobacter suboxydans

Summary A convenient and efficient method of NADPH production from NADP⁺ has been established using a glucose dehydrogenase system involving whole cells and immobilized cells of Gluconobacter suboxydans IFO 3172. Using airdried cells of the bacterium, the optimum conditions for NADPH production were examined, including the cell and glucose concentrations, NADP⁺ concentration, pH, buffer and reaction temperature. Under suitable conditions, the conversion ratio and the amount of NADPH accumulated reached about 100% and 73 mg/ml of the reaction mixture, respectively, after 1-h reaction. Intact cells of the bacterium also showed high NADPH production even in the reaction mixture without a surfactant. The addition of Triton X-100 to the reaction mixture and freeze-thawing treatment of intact cells enhanced the production. The NADPH production method was further improved by using cells of the bacterium immobilized by entrapment in a -carrageenan gel lattice. The immobilized cells had almost the same enzymatic properties as the air-dried cells. The conditions for the continuous production of NADPH with an immobilized cell column were also investigated. NADPH was produced in a good yield (about 95%) with this continuous process.     

Structure of human phenylethanolamine n-methyltransferase gene: existence of two types of mRNA with different transcription initiation sites

The chromosomal gene for human phenylethanolamine N-methyltransferase (PNMT; EC 2.1.1.28) was isolated from a human genomic library using a cloned human PNMT cDNA as a probe, and the nucleotide sequence was determined. PNMT is encoded in a single gene which consists of three exons. We observed newly the presence of minor PNMT mRNA (type B) besides the major mRNA (type A) as reported previously (Kaneda et al., J. Biol. Chem. 263, 7672–7677, 1988) by Northern hybridization. Type B mRNA carries an approximately 700 nucleotide-long untranslated region in the 5′ terminus. This suggests that two types of mRNA are produced from a single gene through the use of two alternative promoters. A TATA-like sequence locates 30 base pair upstream from the cap site of type A mRNA. Upstream of the cap site, there are several sequences resembling Spl binding sites and glucocorticoid responsive elements, with the latter also found in the first intron.     

Expression of active alpha-3 subunit of rat brain Na+,K+-ATPase from the messenger RNA injected into Xenopus oocytes

Cloned cDNA encoding the so-far uncharacterized alpha-3 subunit of rat brain Na+,K+-ATPase (Hara et al. (1987) J. Biochem. 102, 43-58, Shull et al. (1986) Biochemistry 25, 8125-8132) was incorporated into a vector carrying the SP6 promoter. The mRNA produced in vitro was injected into Xenopus oocytes with the mRNA encoding the Na+,K+-ATPase beta subunit of Torpedo electroplax. Increased Na+,K+-ATPase activity in the oocyte membrane was observed. This newly expressed activity was inhibited by ouabain (Ki = 1.5 x 10(-7) M), suggesting that the alpha-3 subunit of rat brain Na+,K+-ATPase is a highly ouabain-sensitive catalytic subunit.     

Induction of tumoricidal activated macrophages by a liposome-encapsulated glycolipid, trehalose 2,3,6'-trimycolate from Gordona aurantiaca

A mycolic acid-containing glycolipid, trehalose 2,3,6'-trimycolate, prepared from a non-pathogenic acid-fast bacterium Gordona aurantiaca, was shown to induce strong tumoricidal activity in peritoneal exudate cells by intravenous or intraperitoneal injection of liposome-encapsulated preparations. The mycolic acid derivative containing a high proportion of unsaturated fatty acids rendered macrophages cytotoxic against syngeneic mastocytoma cells in the absence of endotoxin, for over 14 days after the injection. The macrophages were ascertained to be at low intracellular levels of a lysosomal enzyme beta-galactosidase and an ectoenzyme alkaline phosphodiesterase, a specific pattern as previously described for "primed macrophages". However the culture supernatants of the peritoneal exudate cells were not cytotoxic.     

Peri-operative immunotherapy with OK-432

Pre- and postoperative intradermal administration of OK-432 enhanced the SU-PS skin reaction in patients with gastric cancer, but failed to prevent a fall in the NK activity induced by the operation.The change in NK activity was not associated with a change in the proportion of Leu 7-positive cells, but was related to Leu 11a-positive cells. Intradermal injection of OK-432 increased the proportion of Leu 7-positive cells in the patients in whom they accounted for less than 20% of lymphocyte population. The case was the same with Leu 11a-positive cells.Intravenous injection of OK-432 tended to increase suppressor-inducer T cells (CD4⁺2HA⁺ cells), B cells and Leu 7-positive cells. Particularly, the proportions of OK-M₁-positive cells and MHC class II antigen-positive cells increased in all patients. Immunotherapy with OK-432 given intravenously at a dose of 0.1 KE appeared to be safe because no side effects were essentially observed.     

The oriC unwinding by dam methylation in Escherichia coli.

It has been shown that dam methylation is important in the regulation of initiation of DNA replication in E.coli. The question then arises as to whether dam methylation in the oriC region mediates any structural changes in DNA involved in the regulation of initiation of DNA replication. We demonstrate that the thermal melting temperature of the oriC region is lowered by adenine methylation at GATC sites. The regulation of initiation of DNA replication by dam methylation may be attributed to the ease of unwinding at GATC sites in oriC.     

Distribution of six species of bitterlings in a creek in Fukuoka Prefecture,Japan

Comparative study on the distribution of six species of bitterling in a creek in Fukuoka Prefecture was carried out. The upper part of the creek was fluvial, and the lower part stagnant.Acheilognathus lanceolatus, A. tabira subsp. (b) of Nakamura (1969),A. rhombeus andRhodeus atremius showed wide distribution in the creek throughout their life.A. limbatus seemed to be a fluvial species, andR. ocellatus smithii a lentic species. In the spawning season, however, all the species of bitterling, exceptingR. o. smithii, probably possess a common spawning ground in which mussels occur at a relatively high density. Adult females ofA. lanceolatus, A. tabira subsp. (b) andR. atremius without ripe eggs aggregated in the lower, stagnant parts of the creek. Most juveniles of these species and ofA. rhombeus seemed to grow in the lower part of the creek, and then migrate upward untill their spawning season. No species among the same phylogenetic group has the same set of the life modes (spawning season, spawning sites, aggregation of females without ripe eggs in the lower part of the creek and probability of upstream migration of 0-year-old fish toward spawning sites).