Effect of Kainic Acid, Glutamate, and Aspartate on CO2 Production by Goldfish Tectal Slices

For a study of the excitatory effect of kainate, glutamate, and aspartate in the goldfish optic tectum, these substances were tested on the production of CO2 from radioactive glucose in tectal slices incubated in Krebs-Ringer medium for fish. Kainate increased the rate of CO2 production for up to 30 min in a dose-related manner, the effect being maximum at 0.1 mM concentration and decreasing at higher doses. The effect was blocked by ouabain (1 mM) as well as by the substitution of choline for Na+ in the incubation medium. Glutamate and aspartate exerted a less pronounced excitatory effect on CO2 production at higher concentration than kainate. This effect was also abolished by ouabain. Glutamate, added to the medium at a concentration at least 100-fold higher than kainate, partially reversed the increase in CO2 production induced by kainic acid. No similar effect was noticed for aspartate. The supposed glutamate antagonists glutamic acid diethylester (1 mM) and proline (5 mM) did not affect the excitatory action of kainic acid or exert an antagonistic effect towards glutamate. At higher concentration (10 mM) glutamic acid diethylester increased CO2 production, an effect that was, however, ouabain insensitive. Methyltetrahydrofolic acid (1 mM), a substance reported to compete for the kainate receptor, did not inhibit the effect of kainic acid or increase CO2 production.     

Recognition of human minor alloantigen(s) by cytotoxic lymphocytes in vitro

Lymphocytes from an extensively transfused patient with aplastic anemia were induced to cytotoxicity against target cells from several HLA-matched siblings by in vitro stimulation with allogeneic cells. Effective stimulating cells shared HLA-B7 with the patient, but not all B7 individuals were effective. An additional factor, which was found to segregate in both the patient's and an unrelated sibship, was also necessary. Segregation of this minor alloantigen, W, was also revealed among the patient's HLA-matched sibs by differential susceptibility to lysis by effectors from the patient. The ratio of six positive to four negative siblings suggests that the antigen difference might be coded by a single locus. Lymphocytes from a normal sib, who like the patient is lacking the minor antigen, could not be induced to cytotoxicity against positive targets. Thus in vivo sensitization of the donor of the responding cells appears to be necessary for the demonstration of the cytotoxic response to the minor antigen in vitro. No correlation was observed between the segregation pattern of W and of known blood group antigens, and no cytotoxic antibody to W was detected in the patient's serum in several trials.Abbreviations used in this paper MHC major histocompatibility complex - Tc thymus dependent cells capable of mediating cytotoxicity in the absence of Immoral antibody - GVHD graft versus host disease - CML cell mediated lympholysis - MLC unidirectional mixed lymphocyte culture - ADCC antibody-dependent cell mediated cytotoxicity     

Exploring encapsulation strategies as a protective mechanism to avoid amensalism in mixed populations of Pseudomonas taetrolens and Lactobacillus casei

Bioprocess and Biosystems Engineering - Pseudomonas taetrolens constitutes an efficient platform for the biosynthesis of lactobionic acid, a potentially prebiotic compound. Unfortunately, an...     

A new kumamolisin-like protease from Alicyclobacillus acidocaldarius: an enzyme active under extreme acidic conditions

A new serine-carboxyl proteinase, called kumamolisin-ac, was purified from the thermoacidophilic bacterium Alicyclobacillus acidocaldarius. The enzyme is a monomeric protein of 45?kDa, active over a wide temperature range (5.0–70°C) and extremely acidic pHs (1.0–4.0), showing maximal proteolytic activity at pH?2.0 and 60°C. Interestingly, kumamolisin-ac displayed a significant proteolytic activity even at 5°C, thus suggesting a sort of cold-adaptation for this enzyme. The protease was remarkably stable at high temperatures (t_1/2 at 80°C, 10?h, pH?2.0) and over a broad range of pH (2.0–7.0). Substrate analysis indicated that kumamolisin-ac was active on a variety of macromolecular substrates, such as haemoglobin, hide powder azure, and azocoll. In particular, a high specific activity was detected towards collagen. The corresponding gene was cloned, expressed and the recombinant protease, was found to be homologous to proteases of the ‘S53’ family. From the high identity with kumamolisin and kumamolisin-As, known as collagenolytic proteases, kumamolisin-ac can be considered as the third collagenolytic affiliate within the ‘S53’ family. Cleavage specificity investigation of kumamolisin-ac revealed a unique primary cleavage site in bovine insulin B-chain, whereas a broad specificity was detected using bovine α-globin as substrate. Thus, kumamolisin-ac could represent an attractive candidate for industrial-scale biopeptide production under thermoacidophilic conditions.     

Osservazioni Preliminari sui Sistemi Degradanti dell'RNA nei semi di Ricino

Abstract

Preliminary observations on the enzymatic degradation of RNA in castor bean seeds. — Cocucci, Maggio, Monroy and Marrè have shown the decrease of RNA content during ripening in castor bean seeds, and its increase during germination. Furthermore, these Authors have demonstrated that in the dry ripe seeds the ribosomes are undetectable, and that they increase rapidly during germination. Two peaks of ribosomes are easily detected upon ultracentrifugal analysis in germinating seeds (Cocucci and Sturani). These observations were the basis for our investigations of the enzymes of RNA metabolism in castor bean seeds. This paper deals with our preliminary observations on RNA degrading enzymes in these tissues. We have been able to measure RNase activity, phosphodiesterase, 3′-,5′- and 2′-nucleotidases in castor bean seeds at different stages of development. RNase activity (measured in crude extracts) changes little during the ripening process, its rate corresponding to 40–50 μMoles of nucleotides liberated from RNA per hour and per gram of fresh weight. In the dry seeds, RNase activity is 30–40 μMoles of nucleotides/h.g.f.w., and it increases to about 60–70 μMoles/h/g.f.w. after 72 hours of germination.

Phosphodiesterase activity is about 4–5 μMoles/h.g.f.w.

The following rates have been found in seeds almost completely ripe seeds for 3′-, 5′- and 2′-nucleotidase activities, respectively 45–50 μMoles/h.g.f.w.; 6–7 μMoles/h.g.f.w.; 8 μMoles/h.g.f.w.; ATP-ase activity was of about 80–100 μMoles of phosphate liberated /h.g.f.w. - The high activity of 3′-nucleotidase, of the same order of that of RNase, suggests that these two enzymes are responsible for degradation of RNA to nucleosides and inorganic phosphate. Further investigations are being carried on to define the biochemical properties of castor bean RN-ase.     

Specificity of Lipoxygenase Pathways Supports Species Delineation in the Marine Diatom Genus Pseudo-nitzschia

Oxylipins are low-molecular weight secondary metabolites derived from the incorporation of oxygen into the carbon chains of polyunsaturated fatty acids (PUFAs). Oxylipins are produced in many prokaryotic and eukaryotic lineages where they are involved in a broad spectrum of actions spanning from stress and defense responses, regulation of growth and development, signaling, and innate immunity. We explored the diversity in oxylipin patterns in the marine planktonic diatom Pseudo-nitzschia. This genus includes several species only distinguishable with the aid of molecular markers. Oxylipin profiles of cultured strains were obtained by reverse phase column on a liquid chromatograph equipped with UV photodiode detector and q-ToF mass spectrometer. Lipoxygenase compounds were mapped on phylogenies of the genus Pseudo-nitzschia inferred from the nuclear encoded hyper-variable region of the LSU rDNA and the plastid encoded rbcL. Results showed that the genus Pseudo-nitzschia exhibits a rich and varied lipoxygenase metabolism of eicosapentaenoic acid (EPA), with a high level of specificity for oxylipin markers that generally corroborated the genotypic delineation, even among genetically closely related cryptic species. These results suggest that oxylipin profiles constitute additional identification tools for Pseudo-nitzschia species providing a functional support to species delineation obtained with molecular markers and morphological traits. The exploration of the diversity, patterns and plasticity of oxylipin production across diatom species and genera will also provide insights on the ecological functions of these secondary metabolites and on the selective pressures driving their diversification.