Isolation and characterization of extragenic mutations affecting the expression of the uroporphyrinogen decarboxylase gene (HEM12) in Sacharomyces cerevisiae

Uroporphyrinogen decarboxylase (Uro-d; EC 4.1.1.37), the fifth enzyme in the heme biosynthetic pathway, which catalyzes the sequential decarboxylation of uroporphyrinogen to coproporphyrinogen, is encoded by the HEM12 gene in Saccharomyces cerevisiae. The HEM12 gene is transcribed into a major short mRNA and a minor longer one, approximately 1.35 and 1.55 kb, respectively, in size, and that differ in the 5′ untranslated region. “Uroporphyric” mutants, which have no mutations in the HEM12 gene but accumulate uroporphyrinogen, a phenotype chracteristic of partial Uro-d deficiency, were investigated. Genetic analysis showed that the mutant phenotype depends on the combined action of two unlinked mutations, udt1 and either ipa1, ipa2, or ipa3. ipa1 is tightly linked to HEM12 The mutation udt1 apparently acts specifically on the HEM12 gene, and causes a six to tenfold decrease in the levels of the short HEM12 mRNA, in the β-galactosidase activity of a HEM12-lacZ fusion, in immunodetectable protein and enzyme activity. But heme synthesis is normal and porphyrin accumulation was modest. The mutations ipa1, ipa2, and ipa3 had no phenotype on their own, but they caused an increase in porphyrin accumulation in a udt1 background. This multiplicity of genetic factors leading to uroporphyric yeast cells closely resembles the situation in human porphyria cutanea tarda.     

Glycerolysis of olive oil: batch operational stability of Candida rugosa lipase immobilized in hydrophilic polyurethane foams

The operational stability of the Candida rugosa lipase immobilized in a hydrophilic polyurethane foam was evaluated in consecutive batches for the glycerolysis of olive oil in n-hexane, aimed at the production of monoglycerides.Glycerol controlled the glycerolysis in the system under study, since it is both a substrate and a powerful water binder that reduces the water activity of the reaction medium and of the microenvironment. Two sets of experiments were carried out under different glycerol/triglyceride ratios. After 345 hours of consecutive 23 hours batches no lipase inactivation was observed.List of Symbols a_w thermodynamic activity of water - DG diglyceride (s) - FAME fatty acid methyl ester (s) - FFA free fatty acid (s) - FID flame ionization detector Gly glycerol - MG monoglyceride (s) - TG triglyceride (s) - TLC thin layer chromatography The authors are grateful to Profs. P. Adlercreutz, Technical University of Lund, Sweden, and J.M.S. Cabral, Instituto Superior Técnico, Lisbon, Portugal, for inspiring discussions and advice, to Prof. Helena Pereira, Instituto Superior de Agronomia (ISA), Lisbon, Portugal, for the use of GC equipment and to Mrs. Marlene Dionísio, ISA, for invaluable help with some of the experimental work.     

Adsorption studies for the separation ofl-tryptophan froml-serine and indole in a bioconversion medium

l-tryptophan was produced froml-serine and indole by immobilized Escherichia coli cells in organic-aqueous systems. Selective adsorption was the method chosen to enable both product separation andl-serine reutilization. Amongst various adsorbents tested activated carbons and neutral polymeric resins (XAD-4 and XAD-7) showed good performance. The neutral resins could selectively concentrate thel-tryptophan from dilute aqueous solutions and adsorbed only 5% of the unconvertedl-serine. High separation factors (l-tryptophan/l-serine and indole/l-tryptophan) were obtained with these adsorbents. Despite a lower capacity, the XAD-7 resin had the advantage of desorbingl-tryptophan with basic or acidic solutions, while organic solvents were required to desorb, at the same concentration levels, this compound from XAD-4.In a packed bed column filled with XAD-4 resin or activated carbon, totall-tryptophan adsorption and recovery were achieved at linear velocities up to 5.0 cm/min and 3.2 cm/min respectively. Successive sorbent reutilization, following continuous sorption and elution steps, was carried out in packed bed columns with the neutral resins and activated carbon.Thel-form of tryptophan, after crystallization, was identified by HPTLC.List of Symbols HPLC High Performance Liquid Chromatography - HPTLC High Performance Thin Layer Chromatography - Trp tryptophan - Ser Serine - A amount of sorbent(g) - c equilibrium solute concentration in the aqueous phase (g/dm³) - c _i initial (before adding the sorbent) liquid phase concentration (g/dm³) - C _T tryptophan concentration in the inlet solution (g/dm³) - C _To tryptophan concentration in the outlet solution (g/dm³) - E _z axial dispersion coefficient (m²/s) - k experimental constant (Eq. 1, 2 and 3) - K ₁ rate constant of adsorption (min^–1) - L column length(m) - n experimental constant (eq. 1, 2 and 3) - q equilibrium solid phase concentration (g solute/g sorbent) - q _max maximum capacity of sorbent (g solute/g sorbent) - t time(s) - v liquid velocity (m/s) - V volume of liquid phase(dm³) - V _e eluted volume(dm³) - V _r volume needed to saturate the column (dm³)     

The Pectinase produced by Tubercularia vulgaris in submerged culture using pectin or orange-pulp pellets as inducer

The growth of four strains of the shiitake mushroom Lentinus edodes in solid substrate fermentation in synthetic oak sawdust logs was studied over a 14-week period. Total extracellular phenol oxidase activity and soluble protein were monitored and biomass estimated as the ergosterol content of the fermented sawdust. It was observed that two of the strains had a similar pattern of phenol oxidase activity with two cycles with maxima at 2 and 8 weeks of mycelial growth prior to fruiting. With the other two strains there was a maximum at week 4. For each strain, phenol oxidase activity increased with the cold shock used to induce fruiting. Phenol oxidase activity was not found to be correlated with either soluble protein or total fungal biomass in the fermented sawdust, which were correlated for each strain. Quantification of biomass from submerged liquid culture on the basis of dry weight and ergosterol contents showed that the strains fell into the same two groups with respect to the ergosterol to biomass ratio, which was markedly lower than that for a strain of L. lepideus.Correspondence to: B. C. Okeke     

Effect of ethanol and fatty acids on malolactic activity ofLeuconostoc oenos

Medium-chain fatty acids (C₆ to C₁₂), produced by yeast metabolism during alcoholic fermentation, are known to be inhibitory to lactic acid bacteria. The purpose of this work was to clarify the effect of both ethanol and decanoic and dodecanoic acids on the growth and malolactic activity of aLeuconostoc oenos strain isolated from Portuguese red wine. Ethanol in concentrations up to 12% had no significant effect on malolactic activity but strongly inhibited cell growth. The fatty acids decanoic acid, in concentrations up to 12.5 mg l^–1, and, dodecanoic acid up to 2.5 mg l^–1 seemed to act as growth factors stimulating also malolactic activity; at higher concentrations they exerted an inhibitory effect. We found clear pH dependence between pH 3.0 and pH 6.0, between decanoic acid concentration and its effect on malolactic activity, indicating that the undissociated molecule is the active form. At pH 3.0 the results can be explained by considering that fatty acids enter the cell as protonated molecules and dissociate in the cytoplasm due to the higher internal pH, leading to increased intracellular hydrogenous concentration. This may be the basis of two different effects that contribute to the observed inhibition: decrease in the intracellular pH and dissipation of the transmembrane proton gradient, thus inhibiting intracellular enzymes and ApH-dependent transport systems.     

Lead as an inductor of some morphological and functional changes in synaptosomes from rat brain

Summary 1. The effect of lead (in vivo) on the uptake of GABA, dopamine, and histidine as a precursor of histamine in synaptosomes obtained from chronically lead-treated rats was studied.2. Lead decreased the uptake of GABA, increased the uptake of dopamine, and did not change the uptake of histidine. These effects were independent of calcium concentration.3. Lead administration to the rat changed the morphology of the synaptosomes, as manifested in the decreased number of synaptic vesicles and disturbed mitochondrial structure.4. The results suggest the existence of several mechanisms of lead toxicity on uptake, related to individual neurotransmitters, which are not necessarily connected with a Pb²⁺/Ca²⁺ interaction.     

Basement membrane components of bronchial epithelium in humans suffering from chronic nonspecific lung diseases

Collagen IV and laminin are important constituents of the basement membrane (BM). By use of immunocytochemistry we examined the occurrence and distribution of these two components in the BM beneath normal, mucoid and metaplastic epithelium of large bronchi in 22 adults suffering from chronic nonspecific lung diseases. Both collagen IV and laminin were expressed as a thin and continuous layer beneath the epithelium in most tissue specimens with normal epithelium. In a few specimens the layer showed interruptions with a patchy distribution of the immunoreactivity. Three patterns of distribution of BM components were found under the metaplastic epithelium. Total absence of immunoreactive collagen IV and laminin was the most common variant. Weak and scarce staining for both proteins in the BM characterized the second pattern. The third variant showed strong collagen IV immunoreactivity but lack of laminin. The BM beneath the mucoid epithelium was characterized by irregular distribution of collagen IV and laminin. We suggest that the occurrence and distributional pattern of the BM components are related to the type of overlying epithelium and connected with an altered synthesis of these components.     

Existence and uniqueness of a sharp travelling wave in degenerate non-linear diffusion Fisher-KPP equations

In this paper we use a dynamical systems approach to prove the existence of a unique critical value c ^* of the speed c for which the degenerate density-dependent diffusion equation u _ct = [D(u)u _x ] _x + g(u) has: 1. no travelling wave solutions for 0 < c < c ^*, 2. a travelling wave solution u(x, t) = (x - c ^* t) of sharp type satisfying (– ) = 1, () = 0 ^*; '(^*–) = – c ^*/D'(0), '(^*+) = 0 and 3. a continuum of travelling wave solutions of monotone decreasing front type for each c > c ^*. These fronts satisfy the boundary conditions (– ) = 1, '(– ) = (+ ) = '(+ ) = 0. We illustrate our analytical results with some numerical solutions.     

Subject: Migration

Research Notes on Avian Biology 1994: Selected Contributions from the 21st International Ornithological CongressGeneral Biology: Migration

Subject: Migration