International Journal Abstracts

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International Journal Abstracts

The following research results, of which the abstracts are provided here, have been published in various International Journals during 2006.

The effect of polysaccharide-degrading wine yeast transformants on the efficiency of wine processing and wine flavour
C.T. Louw ¹, D. la Grange ², I.S. Pretorius ^1,3 & P. van Rensburg ¹.
1. Institute for Wine Biotechnology, Department of Viticulture & Oenology, Stellenbosch University, 7602 Matieland, South Africa
2. Department of Microbiology, Stellenbosch University, Private Bag X1, 7602 Matieland, South Africa
3. The Australian Wine Research Institute, PO Box 197, Glen Osmond (Adelaide), SA 5064, Australia
Commercial polysaccharase preparations are applied to winemaking to improve wine processing and quality. Expression of polysaccharase-encoding genes in Saccharomyces cerevisiae allows for the recombinant strains to degrade polysaccharides that traditional commercial yeast strains cannot. In this study, we constructed recombinant wine yeast strains that were able to degrade the problem-causing grape polysaccharides, glucan and xylan, by separately integrating the Trichoderma reesei XYN2 xylanase gene construct and the Butyrivibrio fibrisolvens END1 glucanase gene cassette into the genome of the commercial wine yeast strain S. cerevisiae VIN13. These genes were also combined in S. cerevisiae VIN13 under the control of different promoters.
The strains that were constructed were compared under winemaking conditions with each other and with a recombinant wine yeast strain expressing the endo-?-1,4-glucanase gene cassette (END1) from B. fibrisolvens and the endo-?-1,4-xylanase gene cassette (XYN4) from Aspergillus niger, a recombinant strain expressing the pectate lyase gene cassette (PEL5) from Erwinia chrysanthemi and the polygalacturonase-encoding gene cassette (PEH1) from Erwinia carotovora. Wine was made with the recombinant strains using different grape cultivars. Fermentations with the recombinant VIN13 strains resulted in significant increases in free-flow wine when Ruby Cabernet must was fermented. After 6 months of bottle ageing significant differences in colour intensity and colour stability could be detected in Pinot Noir and Ruby Cabernet wines fermented with different recombinant strains. After this period the volatile composition of Muscat d’Alexandria, Ruby Cabernet and Pinot Noir wines fermented with different recombinant strains also showed significant differences. The Pinot Noir wines were also sensorial evaluated and the tasting panel preferred the wines fermented with the recombinant strains.
Journal of Biotechnology 125 2006 447 - 461

Rapid screening of the fermentation profiles of wine yeasts by Fourier transform infrared spectroscopy
H.H. Nieuwoudt ^1,2, I.S. Pretorius ³, F.F. Bauer ², D.G. Nel ⁴ & B.A. Prior ¹
1. Department of Microbiology, Stellenbosch University, Private Bag X1, 7602 Matieland, South Africa
2. Institute for Wine Biotechnology, Department of Viticulture and Oenology, Stellenbosch University, Private Bag X1, 7602 Matieland, South Africa
3. The Australian Wine Research Institute, PO Box 197, Glen Osmond, Adelaide, SA 5064, Australia
4. Centre for Statistical Consultation, Department of Statistics, Stellenbosch University, Private Bag X1, 7602 Matieland, South Africa
A rapid screening method for the evaluation of the major fermentation products of Saccharomyces wine yeasts was developed using Fourier transform infrared spectroscopy and principal component factor analysis. Calibration equations for the quantification of volatile acidity, glycerol, ethanol, reducing sugar and glucose concentrations in fermented Chenin blanc and synthetic musts were derived from the Fourier transform infrared spectra of small-scale fermentations. The accuracy of quantification of volatile acidity in both Chenin blanc and synthetic must was excellent, and the standard error of prediction was 0.07 g l?1 and 0.08 g l?1, respectively. The respective standard error of prediction in Chenin blanc and synthetic musts for ethanol was 0.32% v/v and 0.31% v/v, for glycerol was 0.38 g l?1 and 0.32 g l?1, for reducing sugar in Chenin blanc must was 0.56 g l?1 and for glucose in synthetic must was 0.39 g l?1. These values were in agreement with the accuracy obtained by the respective reference methods used for the quantification of the components. The screening method was applied to quantify the fermentation products of glycerol over-producing hybrid yeasts and commercial wine yeasts. Principal component factor analysis of the fermentation data facilitated an overall comparison of the fermentation profiles (in terms of the components tested) of the strains. The potential of Fourier transform infrared spectroscopy as a tool to rapidly screen the fermentative properties of wine yeasts and to speed up the evaluation processes in the initial stages of yeast strain development programs is shown.
Journal of Microbiological Methods 67 2006 248 - 256

Regulation of respiratory growth by Ras: The glyoxylate cycle mutant, cit2, is suppressed by RAS2
J.H. Swiegers ¹, I.S. Pretorius ¹ & F.F. Bauer ²
1. The Australian Wine Research Institute, PO Box 197, Glen Osmond, Adelaide, SA 5064, Australia
2. Institute for Wine Biotechnology, Department of Oenology and Viticulture, Faculty of Agricultural and Forestry Sciences, University of Stellenbosch, 7600 Stellenbosch, South Africa
In Saccharomyces cerevisiae the Ras/cAMP/ PKA signalling pathway controls multiple metabolic pathways, and alterations in the intracellular concentrations of cAMP through modiWcation of signalling pathway factors can be lethal or result in severe growth defects. In this work, the important role of Ras2p in metabolic regulation during growth on the non-fermentable carbon source glycerol is further investigated. The data show that the over-expression of RAS2 suppresses the growth defect of the glyoxylate cycle citrate synthase mutant, cit2. The over-expression results in enhanced proliferation and biomass yield when cells are grown on glycerol as sole carbon source, and increases citrate synthase activity and intracellular citrate concentration. Interestingly, the suppression of cit2 and the enhanced proliferation and biomass yield are only observed when RAS2 is over-expressed and not in strains containing the constitutively active allele RAS2val19. However, both RAS2 and RAS2^val19 upregulated citrate synthase activity. We propose that the RAS2 over-expression results in a combination of general upregulation of respiratory growth capacity and an increase in mitochondrial citrate/citrate synthases, which together, complement the metabolic requirements of the cit2 mutant. The data therefore provide new evidence for the role of Ras2p as a powerful modulator of metabolism during growth on a non-fermentable carbon source.
Current Genetics 50 2006 161 - 171

High-performance liquid chromatography profiling of the major carotenoids in Arabidopsis thaliana leaf tissue
K.L. Taylor ¹, A.E. Brackenridge ¹, M.A. Vivier ^1,2 & A. Oberholster ²
1. Institute for Wine Biotechnology, Stellenbosch University, Stellenbosch 7600, South Africa
2. Department of Viticulture and Oenology, Stellenbosch University, Stellenbosch 7600, South Africa
Carotenoids are extremely sensitive to a variety of physico-chemical attacks which may have a profound effect on their characteristic properties, thereby influencing the accurate identification and quantification of individual compounds. In this light, a comprehensive summary of the pitfalls encountered and precautions to be administered during handling and storage of authentic standards and samples was found to be incomplete. Furthermore, acceptable baseline separation of trans-lutein from trans-zeaxanthin and between the cis- and trans-forms of neoxanthin and violaxanthin has not been satisfactorily demonstrated. Hence the most optimal sample preparation and analytical steps were determined and a sensitive and reproducible method for the quantitative HPLC profiling of the principal carotenoids found in plant leaf tissue was developed. A reverse-phase C30 column with a binary mobile solvent system was used for the baseline separation of eight of the major carotenoids and the two chlorophylls (a and b) within 18 min. These compounds were identified via the use of authentic standards, their spectral characteristics and HPLC-atmospheric pressure chemical ionization (APCI)-mass spectrometry (MS) confirmation. This method has been successfully applied for the quantification of plant pigments in Arabidopsis thaliana wild-type (WT) leaf tissue and in two A. thaliana non-photochemical mutants, namely npq1 and npq2. These mutants have previously been well-characterised and provided valuable reference data as well as acting as internal controls for the assessment of our new method.
Journal of Chromatography A 2006 1121: 83 - 91

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