Mutational analysis of Arabidopsis thaliana plant uncoupling mitochondrial protein

In this study, point mutations were introduced in plant uncoupling mitochondrial protein AtUCP1, a typical member of the plant uncoupling protein (UCP) gene subfamily, in amino acid residues Lys147, Arg155 and Tyr269, located inside the so-called UCP-signatures, and in two more residues, Cys28 and His83, specific for plant UCPs. The effects of amino acid replacements on AtUCP1 biochemical properties were examined using reconstituted proteoliposomes. Residue Arg155 appears to be crucial for AtUCP1 affinity to linoleic acid (LA) whereas His83 plays an important role in AtUCP1 transport activity. Residues Cys28, Lys147, and also Tyr269 are probably essential for correct protein function, as their substitutions affected either the AtUCP1 affinity to LA and its transport activity, or sensitivity to inhibitors (purine nucleotides). Interestingly, Cys28 substitution reduced ATP inhibitory effect on AtUCP1, while Tyr269Phe mutant exhibited 2.8-fold increase in sensitivity to ATP, in accordance with the reverse mutation Phe267Tyr of mammalian UCP1.     

Production of transgenic tropical maize with cryIAb and cryIAc genes via microprojectile bombardment of immature embryos

To enhance the level of resistance to insects in tropical maize germplasm we have developed techniques to successfully transform elite tropical maize inbred based on the activity of specific cryI proteins against four major maize pests – corn earworm, fall armyworm, southwestern corn borer and sugarcane borer. Constructs containing cryIAb or cryIAc synthetic genes were used. To generate transgenic plants we have established methods for biolistic bombardment and the selection and regeneration of immature embryos and calli from the elite tropical lines CML72, CML216, CML323, CML327 and hybrids. Transgenic plants resistant to the herbicide Basta^TM contained the bands for the cry, bar and gus genes as detected by Southern blot analyses. A simple leaf bioassay presented varying levels of resistance to Southwestern corn borer of transgenic tropical maize carrying the cryIAc gene. Analyses of the progenies confirmed the sexual transmission of the introduced genes and their stable expression. Received: 25 September 1998 / Accepted: 27 October 1998     

The Schizosaccharomyces pombe Pzh1 protein phosphatase regulates Na+ ion influx in a Trk1-independent fashion.

We have previously shown that fission yeast encodes a PPZ-like phosphatase, designated Pzhl, which is an important determinant of cation homeostasis. pzh1 delta mutants display increased tolerance to Na+ ions, but they are hypersensitive to KC1 [Balcells, L., Gómez, N., Casamayor, A., Clotet, J. & Ari?o, J. (1997) Eur. J. Biochem. 250, 476-483]. We have immunodetected Pzh1 in yeast extracts and found that this phosphatase is largely associated with particulate fractions. Cells defective in Pzh1 do not show altered efflux of Na+ or Li+ ions, but they accumulate these cations more slowly than wild-type cells. K+ ion content of pzh1 delta cells is about twice that of wild-type cells, and this can be explained by decreased efflux of K+. Therefore, Pzh1 may regulate both Na+ influx and K+ efflux in fission yeast. To test the possible relationship between K+ uptake, Na+ tolerance and Pzh1 function, we deleted the trk1+ gene, which encodes a putative high-affinity transporter of K+ ions. trkl delta mutants grew well even at relatively low concentrations of KCl and did not show significantly altered content or influx of K+ ions. However, they showed a Na(+)-sensitive phenotype which was greatly intensified by deletion of the sod2+ gene (which encodes the major determinant for efflux of Na+ ions), and clearly ameliorated by deletion of the pzh1 phosphatase, as well as by moderate concentrations of KCl in the medium. These results suggest that Trk1 does not mediate the effect of Pzh1 on NaCl tolerance and that fission yeast contains efficient systems, other than Trk1, for uptake of K+ ions.     

Effect of arbuscular mycorrhizal fungi on the acclimatization of micropropagated banana plantlets

 The effect of arbuscular mycorrhizal fungi (AMF) on micropropagated banana plantlets was evaluated during the acclimatization period. Plants were inoculated with Acaulospora scrobiculata, Glomus clarum or Glomus etunicatum. After cultivation in a greenhouse for 3 months, height, leaf area, fresh weight and dry matter of root and shoots, level of AMF colonization, nutrient level, photosynthesis and transpiration rate, water potential and stomatal conductance were measured. The number of AMF spores produced in each treatment was also determined. Plantlets inoculated with AMF had greater height, leaf area and fresh weight of shoots and roots, as well as higher rates of photosynthesis and transpiration than controls. Plants inoculated with Glomus were superior in most of the evaluated parameters. Accepted: 24 May 1999     

Experimental design to enhance the production of l-(+)-lactic acid from steam-exploded wood hydrolysate using Rhizopus oryzae in a mixed-acid fermentation

The fermentation of hemicellulosic hydrolysate from Pinus taeda chips, using the fungal culture Rhizopus oryzae, was carried out to produce l-(+)-lactic acid and to optimize and enhance the biological conversion of reducing sugar into l-(+)-lactic acid using the experimental design to evaluate the culture conditions. The first factorial design based on surface response with five factors (agitation level, substrate concentration, CaCO₃ concentration, C/N and C/P ratios) at low levels and one medium point was performed to optimize culture conditions. The second study tested two factors (substrate concentration and C/N ratio) at three levels. The statistical analysis of the data obtained from the factorial study showed that a C/N ratio of 35 and substrate concentration of 90 g/litre were the best conditions to produce l-(+)-lactic acid with R. oryzae on P. taeda hydrolysate, but in this case the statistical projection was not correct and the real optimized conditions were C/N ratio of 55 and substrate concentration of 75 g/litre of reducing sugar.     

Water activity and temperature effects on germination and growth of Eurotium amstelodami, E. chevalieri and E. herbariorum isolates from bakery products

This study investigated the effect of temperature (5-30 degrees C), water activity (0.775-0.90 aw) and their interactions on the temporal rates of germination and mycelial growth of three species of Eurotium on flour wheat sucrose medium. Germination was quite rapid at aw >0.85, with an almost linear increase with time for all isolates. However, under more extreme water stress, germination was slower. The aw minima for germination were usually lower than those for growth and varied with temperature. The effect of aw x temperature interactions on the lag phases (h) prior to germination and on the germination rates (h-1) were predicted using the Gompertz model modified by Zwietering. Eurotium spp. had shown short lag times at 0.90 aw over a wide range of temperatures. At marginal temperatures, lag phases were significantly longer, especially at >15 degrees C. The temperature x aw profiles for mycelial growth varied between species in terms of rates (mm d(-1)). Predictions of the effect of important environmental factors, such as temperature, aw and their interactions on lag times to germination, germination rates and mycelial growth, are important in the development of hurdle technology approaches to predict fungal spoilage in food products.     

Effect of antifoam addition on gas-liquid mass transfer in stirred fermenters

The influence of three well-known antifoaming agents (polypropylene glycol, silicone and soybean oil) on gas-liquid mass transfer in stirred tanks is studied, both in model and in fermentation media. The effect of antifoam concentration, ionic strength, viscosity, agitation speed and gas flow rate are investigated. It is found that antifoam addition at low concentrations markedly decreases the gas-liquid volumetric mass transfer coefficient, k_La, for the three antifoam agents tested. Although the major effect is on the film coefficient k_L, some effect is also detected on the specific area, a. It is found that the influence of viscosity and antifoam addition are not cumulative: each tends to attenuate the other's effect on mass transfer. Both for silicone and for soybean oil, but not for PPG in the concentration range studied, there is an antifoam concentration above which further antifoam addition starts to improve k_La.     

Acetate Utilization in Lactococcus lactis Deficient in Lactate Dehydrogenase: a Rescue Pathway for Maintaining Redox Balance

Acetate was shown to improve glucose fermentation in Lactococcus lactis deficient in lactate dehydrogenase. 13C and 1H nuclear magnetic resonance studies using [2-13C]glucose and [2-(13)C]acetate as substrates demonstrated that acetate was exclusively converted to ethanol. This novel pathway provides an alternative route for NAD+ regeneration in the absence of lactate dehydrogenase.