In vivo analysis of glucose-induced fast changes in yeast adenine nucleotide pool applying a rapid sampling technique

Transition from glucose limitation to glucose excess in Saccharomyces cerevisiae leads to a fast change from complete to partial oxidative metabolism. To determine rapid changes of in vivo concentrations of yeast metabolites, occurring in the range of seconds after a glucose injection, a fast sampling technique was developed. A harvesting device with negligible dead space and highly efficient inactivation and extraction steps was applied to analyze the behavior of the physiological concentrations of adenine nucleotides as the dynamic response to fast changing glucose concentrations. Adenine nucleotides were selected as appropriate references due to their high turnover rates (in the range of seconds). During the first 5 s after a glucose pulse to a continuous chemostat culture, a significant decrease (30%) in ATP concentration and a coincident rise of ADP and AMP values were found. A further advantage of this technique is the high sampling frequency (5 s) and the long-term sterility ensuring recurrent harvesting.