Kinetic analysis of a Saccharomyces cerevisiae strain adapted for improved growth on glycerol: Implications for the development of yeast bioprocesses on glycerol.

Glycerol is an agro-industrial residue generated in high amounts during the biodiesel production. The growing production of biodiesel is creating a worldwide glycerol surplus. Therefore, replacing sugar-based feedstock in bioprocesses by glycerol could be potentially attractive. Saccharomyces cerevisiae is one of the most commonly used microorganisms in the agri-food industry and therefore currently produced in large quantities from sugar-based feedstock. Unfortunately, growth of S. cerevisiae strains on glycerol is very low with reported µmax around 0.01 h(-1). This study demonstrates that successive growth of the S. cerevisiae CBS 8066, CEN.PK 113-7 D and Ethanol Red on glycerol as sole carbon source considerably improved the µmax from 0.01 up to 0.2 h(-1). The "adapted strain" CBS 8066-FL20 was kinetically characterized during aerobic and oxygen-limited cultivation in bioreactor and the results discussed in terms of their implication for developing glycerol-based S. cerevisiae bioprocesses.

Disparity between changes in mRNA abundance and enzyme activity in Corynebacterium glutamicum: implications for DNA microarray analysis.

The relationship between changes in mRNA abundance and enzyme activity was determined for three genes over a span of nearly 3 h during amino acid production in Corynebacterium glutamicum. Gene expression changes during C. glutamicum fermentations were examined by complementary DNA (cDNA) microarrays and by a second method for quantitating RNA levels, competitive reverse transcriptase-PCR (RT-PCR). The results obtained independently by both methods were compared and found to be in agreement, thus validating the quantitative potential of DNA microarrays for gene expression profiling. Evidence of a disparity between mRNA abundance and enzyme activity is presented and supports our belief that it is difficult to generally predict protein activity from quantitative transcriptome data. Homoserine dehydrogenase, threonine dehydratase, and homoserine kinase are enzymes involved in the biosynthesis of l-isoleucine and other aspartate-derived amino acids in C. glutamicum. Our data suggest that different underlying regulatory mechanisms may be connected with the expression of the genes encoding each of these three enzymes. Indeed, whereas in one case the increases in enzyme activity exceeded those in the corresponding mRNA abundance, in another case large increases in the levels of gene expression were not congruent with changes in enzyme activity.

Exopolysaccharide production by Propionibacterium acidi-propionici on milk microfiltrate.

AIMS: The aims of this work were to evaluate growth and exopolysaccharide (EPS) production properties of Propionibacterium acidi-propionici DSM 4900 on milk permeate. METHODS AND RESULTS: Anaerobic growth on milk permeate was only possible if supplemented with yeast extract (YE). Fermentation capacities of the strain were significantly improved by further increasing the supplemented YE. At 5 g l(-1) YE, consumption of 45 g l(-1) lactose to produce 9 g l(-1) biomass, 34 g l(-1) organic acids and 0.65 g l(-1) EPS was observed. From a kinetic point of view, EPS production occurred during the bacteria growth phase. At the excreted polysaccharide level, the medium showed shear-thinning behaviour with a relatively high apparent viscosity of up to 30 mPa.s (milli.Pascal.second) at a shear rate of 17 s(-1). CONCLUSION: EPS production by P. acidi-propionici DSM 4900 on milk permeate showed promising rheological behaviour of the milk-derived medium obtained, even at a low production level. SIGNIFICANCE AND IMPACT OF THE STUDY: A kinetic study on EPS production by a food-grade bacterium that could be used in situ in alimentation was carried out.

Study of the effects of temperature, pH and yeast extract on growth and exopolysaccharides production by Propionibacterium acidi-propionici on milk microfiltrate using a response surface methodology.

AIMS: To study the effects of temperature, pH and yeast extract (YE) concentration on growth and exopolysaccharide (EPS) production by Propionibacterium acidi-propionici DSM 4900 cultivated on milk microfiltrate. METHODS AND RESULTS: A multifactorial approach using a Response Surface Methodology (RSM) was followed. The results indicated that both growth, and EPS and organic acids production, were influenced by pH, temperature and YE concentration. Biomass and organic acids production occurred in all the tested domains, whereas EPS production was only possible in a narrow pH range (5.3-6.5). The results clearly showed that the optimal conditions for EPS production were different to those for optimal growth. The effect of YE on EPS production was not only due to an increase in growth but also to a direct effect on the production of EPS. The temperature played a major role. A decrease of temperature induced a slowing down of both growth and organic acids production, making the essential factors of the medium and the precursors of EPS biosynthesis more available and hence, leading to an increase in EPS production. CONCLUSION: The effects of pH, temperature and YE were determined, allowing the definition of favourable, though non-optimal, conditions for EPS production: 23 degrees C, pH 6 and 3 g l(-1) YE concentration. SIGNIFICANCE AND IMPACT OF THE STUDY: The use of a multifactorial approach for investigating the effect of fermentation conditions on EPS production has been demonstrated.

Metabolic redirection of carbon flow toward isoleucine by expressing a catabolic threonine dehydratase in a threonine-overproducing Corynebacterium glutamicum.

Carbon destined for lysine synthesis in Corynebacterium glutamicum ATCC 21799 can be diverted toward threonine by overexpression of genes encoding a feedback-insensitive homoserine dehydrogenase (hom(dr)) and homoserine kinase (thrB). We studied the effects of introducing two different threonine dehydratase genes into this threonine-producing system to gauge their effects on isoleucine production. Co-expression of hom(dr), thrB, and ilvA, which encodes a native threonine dehydratase, caused isoleucine to accumulate to a final concentration of 2.2+/-0.2 g l(-1), five-fold more than accumulates in the wild-type strain, and approximately twice as much as accumulates in the strain expressing only hom(dr) and thrB. Comparing these data with previous results, we found that overexpression of the three genes, hom(dr), thrB, and ilvA, in C. glutamicum ATCC 21799 is no better in terms of isoleucine production than the expression of a single gene, tdcB, encoding a catabolic threonine dehydratase from Escherichia coli. Co-expression of hom(dr), thrB, and tdcB, however, caused the concentration of isoleucine to increase 20-fold compared to the wild-type strain, about four times more than the corresponding ilvA-expressing strain. In this system, the apparent yield of isoleucine production was multiplied by a factor of two [2.1 mmol (g dry cell weight)(-1)]. While the balance of excreted metabolites showed that the carbon flow in this strain was completely redirected from the lysine pathway into the isoleucine pathway, it also showed that more pyruvate was diverted into amino acid synthesis.