The Calvin cycle enzyme pentose-5-phosphate 3-epimerase is encoded within the cfx operons of the chemoautotroph Alcaligenes eutrophus.

Several genes (cfx genes) encoding Calvin cycle enzymes in Alcaligenes eutrophus are organized in two highly homologous operons comprising at least 11 kb. One cfx operon is located on the chromosome; the other is located on megaplasmid pHG1 of the organism (B. Bowien, U. Windh?vel, J.-G. Yoo, R. Bednarski, and B. Kusian, FEMS Microbiol. Rev. 87:445-450, 1990). Corresponding regions of about 2.7 kb from within the operons were sequenced. Three open reading frames, designated cfxX (954 bp), cfxY (765 bp), and cfxE (726 bp), were detected at equivalent positions in the two sequences. The nucleotide identity of the sequences amounted to 94%. Heterologous expression of the subcloned pHG1-encoded open reading frames in Escherichia coli suggested that they were functional genes. The observed sizes of the gene products CfxX (35 kDa), CfxY (27 kDa), and CfxE (25.5 kDa) closely corresponded to the values calculated on the basis of the sequence information. E. coli clones harboring the cfxE gene showed up to about 19-fold-higher activities of pentose-5-phosphate 3-epimerase (PPE; EC 5.1.3.1) than did reference clones, suggesting that cfxE encodes PPE, another Calvin cycle enzyme. These data agree with the finding that in A. eutrophus, PPE activity is significantly enhanced under autotrophic growth conditions which lead to a derepression of the cfx operons. No functions could be assigned to CfxX and CfxY.     

Induction of ovulation and oocyte maturation of amphibian (Rana dybowskii) ovarian follicles by protein kinase C activation in vitro.

We previously reported that protein kinase C (PKC) activation induced meiotic maturation (germinal vesicle breakdown, GVBD) of Rana dybowskii follicular oocytes cultured in vitro without hormone treatment. The experiments reported here were carried out to establish whether ovarian follicles ovulated in response to PKC activation during culture. A phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA), was used for PKC activation. TPA addition (10 microM) to cultured ovarian fragments induced ovulation and maturation of the oocytes similar to that seen following addition of frog pituitary homogenate (FPH, 0.05 pituitary/ml) or progesterone (0.5 microgram/ml). Such changes were not observed when ovarian fragments were treated with inactive phorbol ester. The time course of TPA-induced ovulation was similar to that produced by FPH-stimulated ovulation. Both TPA- and FPH-stimulated ovulation and maturation were blocked by treatment with cycloheximide, forskolin (an adenylate cyclase stimulator), and 1-(5-isoquinolinylsulfonyl)-2-methyl-piperazine (H-7; a PKC inactivator). FPH treatment markedly increased progesterone levels in the medium during ovarian fragment culture whereas TPA treatment failed to elevate progesterone levels. Thus, TPA treatment mimics FPH and progesterone in inducing ovulation and meiotic maturation in cultured amphibian ovarian fragments. The data strongly suggest that PKC plays an important role in regulating ovulation as well as in modulating amphibian oocyte maturation during follicular differentiation.     

Expression of glucoamylase gene usingSUC2 promoter inSaccharomyces cerevisiae

Summary The cloning of glucoamylase geneSTA using theSUC2 promoter intoSaccharomyces cerevisiae was performed. The signal sequence ofSTA gene was used for the secretion of glucoamylase protein. The plasmid constructed in this way was named YEpSUCSTA and its expression was identified. The expression of YEpSUCSTA was repressed in the presence of glucose in growth medium, but derepressed when glucose became depleted. YEpSUCSTA showed the similar efficiency of glucoamylase secretion as YEpSTA-F which has the entireSTA gene. Glucoamylase activity in starch-glucose medium was largely increased because cell mass and plasmid stability were high in biosynthesis phase compared to extracellular glucoamylase activities in media which starch or glucose was the only carbon source.     

Ferrous Iron and Sulfur Oxidation and Ferric Iron Reduction Activities of Thiobacillus ferrooxidans Are Affected by Growth on Ferrous Iron, Sulfur, or a Sulfide Ore

Eight strains of Thiobacillus ferrooxidans (laboratory strains Tf-1 [= ATCC 13661] and Tf-2 [= ATCC 19859] and mine isolates SM-1, SM-2, SM-3, SM-4, SM-5, and SM-8) and three strains of Thiobacillus thiooxidans (laboratory strain Tt [= ATCC 8085] and mine isolates SM-6 and SM-7) were grown on ferrous iron (Fe²⁺), elemental sulfur (S⁰), or sulfide ore (Fe, Cu, and Zn). The cells were studied for their aerobic Fe²⁺ - and S⁰-oxidizing activities (O₂ consumption) and anaerobic S⁰-oxidizing activity with ferric iron (Fe³⁺) (Fe²⁺ formation). Fe²⁺-grown T. ferrooxidans cells oxidized S⁰ aerobically at a rate of 2 to 4% of the Fe²⁺ oxidation rate. The rate of anaerobic S⁰ oxidation with Fe³⁺ was equal to the aerobic oxidation rate in SM-1, SM-3, SM-4, and SM-5, but was only one-half or less that in Tf-1, Tf-2, SM-2, and SM-8. Transition from growth on Fe²⁺ to that on S⁰ produced cells with relatively undiminished Fe²⁺ oxidation activities and increased S⁰ oxidation (both aerobic and anaerobic) activities in Tf-2, SM-4, and SM-5, whereas it produced cells with dramatically reduced Fe²⁺ oxidation and anaerobic S⁰ oxidation activities in Tf-1, SM-1, SM-2, SM-3, and SM-8. Growth on ore 1 of metal-leaching Fe²⁺-grown strains and on ore 2 of all Fe²⁺-grown strains resulted in very high yields of cells with high Fe²⁺ and S⁰ oxidation (both aerobic and anaerobic) activities with similar ratios of various activities. Sulfur-grown Tf-2, SM-1, SM-4, SM-6, SM-7, and SM-8 cultures leached metals from ore 3, and Tf-2 and SM-4 cells recovered showed activity ratios similar to those of other ore-grown cells. It is concluded that all the T. ferrooxidans strains studied have the ability to produce cells with Fe²⁺ and S⁰ oxidation and Fe³⁺ reduction activities, but their levels are influenced by growth substrates and strain differences.     

Evidence for a role of specific isoacceptor species of tRNA in amino acid transport.

Soybean leghemoglobins ā and b?were compared by microscale peptide mapping after heme removal with acid-acetone. Maps generated by trypsin or the combined action of trypsin and thermolysin indicated a large amount of homology between the proteins with the only variations detected being the N-terminal peptides. The N-terminal tryptic peptide of leghemoglobin b? was found to be both blocked and to lack the first amino acid of the corresponding leghemoglobin ā peptide. Nuclear magnetic resonance and gas chromatography/mass spectroscopy studies showed that the N-terminal of leghemoglobin b? was N-acetyl-alanine. It is possible that leghemoglobin b? arises from leghemoglobin ā by a two-stage modification involving cleavage of the N-terminal valyl residue and subsequent acetylation of the exposed alanyl residue.     

Electron microscopic localization of monoamine oxidase in the rat liver

Summary Two methods have been employed to localize monoamine oxidase activity in the cells of rat liver, using either 2-(2′-benzothiazolyl)-5-stryl-3-(4′-phtalhydrazidyl) tetrazolium chloride (BSPT) or ferricyanide as electron acceptor. With both methods monoamine oxidase activity was found both in the inner and the outer mitochondral membrane, although the outer membrane appeared the most probable location. In addition the BSPT method but not the ferricyanide method, revealed monoamine oxidase activity in the endoplasmatic reticulum. The results obtained by the two methods have been compared and are discussed in view of available biochemical data on monoamine oxidase. Supported by research grants from the National Research Council of Canada (A 3651), The Swedish Medical Research Council (4145) and M. Bergwall's Foundation, Stockholm.     

Allyl Isothiocyanate Ameliorates Angiogenesis and Inflammation in Dextran Sulfate Sodium-Induced Acute Colitis

Allyl isothiocyanate (AITC) is a phytochemical found in cruciferous vegetables that has known chemopreventive and chemotherapeutic activities. Thus far, the antiangiogenic activity of AITC has not been reported in in vivo studies. Herein, we investigated the effect of AITC on angiogenesis and inflammation in a mouse model of colitis. Experimental colitis was induced in mice by administering 3% dextran sulfate sodium via drinking water. To monitor the activity of AITC in this model, we measured body weight, disease activity indices, histopathological scores, microvascular density, myeloperoxidase activity, F4/80 staining, inducible nitric oxide synthase (iNOS) expression, cyclooxygenase-2 (COX-2) expression, and vascular endothelial growth factor (VEGF)-A/VEGF receptor 2 (VEGFR2) expression in the mice. We found that AITC-treated mice showed less weight loss, fewer clinical signs of colitis, and longer colons than vehicle-treated mice. AITC treatment also significantly lessened the disruption of colonic architecture that is normally associated with colitis and repressed the microvascularization response. Further, AITC treatment reduced both leukocyte recruitment and macrophage infiltration into the inflamed colon, and the mechanism these activities involved repressing iNOS and COX-2 expression. Finally, AITC attenuated the expression of VEGF-A and VEGFR2. Thus, AITC may have potential application in treating conditions marked by inflammatory-driven angiogenesis and mucosal inflammation.     

Taxonomic revision of the genus Jassidophaga Aczél (Diptera: Pipunculidae) in Korea

In this paper, three Korean species of the big‐head fly, genus Jassidophaga Aczél were treated: J. chiiensis (Ouchi), J. pala (Morakote) and J. villosa (Roser). Of these, J. pala is reported in Korea for the first time. A key to Korean species and photographs on external features are given.