Metabolite profiles of the biocontrol yeast Pichia anomala J121 grown under oxygen limitation (original) (raw)

2004, Applied Microbiology and Biotechnology

The biocontrol yeast Pichia anomala prevents mould damage of moist cereal grain during malfunctioning airtight storage but it can also spoil food and feed. This thesis focuses on the physiology and metabolism of P. anomala, in particular during oxygen limitation, a condition relevant to airtight storage of cereal grain. P. anomala grew under strictly anaerobic conditions, at temperatures between 3ºC and 37ºC, pH values between 2.0 and 12.4, low water activity (0.85), and on many different nutrients. Accumulation of low-molecular compounds in living cells was analysed by HR MAS-NMR. Glycerol, arabitol, and trehalose accumulation increased with reduced oxygen availability, indicating a role during oxygen-limited growth. Regulation of the central aerobic and hypoxic metabolism of P. anomala was investigated under controlled fermentor conditions. Oxygen limitation induced alcoholic fermentation as well as activity of the key fermentative enzymes, ADH and PDC. Metabolic flux analysis revealed that the TCA pathway operated as a cycle during aerobic batch culture and as a twobranched pathway under oxygen limitation. Hypoxic conditions also increased the production of ethyl acetate, an ester involved in the biocontrol activity of P. anomala. Genes encoding the ADH and PDC enzymes were cloned, PaADH1, PaADH2, and PaPDC1, and their expression was analysed with real-time RT-PCR. PaADH1 and PaPDC1 were expressed during aerobic growth on glucose and ethanol and were up-regulated in response to oxygen limitation. PaADH2 expression was low during these growth conditions, i.e. <1% of the level of its isogene, PaADH1. In cells grown on succinate, the expression of the two ADH isogenes was the opposite, high expression of PaADH2 and low expression of PaADH1. The upregulation of gene expression and enzyme activity did not quantitatively correlate with glycolytic flux. Thus, additional regulatory phenomena at the posttranscriptional and posttranslational level are important in the distribution of carbon through the respiratory and fermentative pathways.