Dynamic¹³C-tracer study of storage carbohydrate pools in aerobic glucose-limitedSaccharomyces cerevisiaeconfirms a rapid steady-state turnover and fast mobilization during a modest stepup in the glucose uptake rate

Abstract

In this research, two dynamic (13)C-labeling experiments confirmed turnover and rapid mobilization of stored glycogen and trehalose in an aerobic glucose-limited chemostat (D=0.05 h(-1)) culture of Saccharomyces cerevisiae. In one experiment, the continuous feed to an aerobic glucose-limited chemostat culture of S. cerevisiae was instantaneously switched from naturally labeled to fully (13)C labeled while maintaining the same feed rate before and after the switch. The dynamic replacements of naturally labeled intracellular glycolytic intermediates and CO(2) (in the off-gas) with their (13)C-labeled equivalents were measured. The data of this experiment suggest that the continuous turnover of glycogen and trehalose is substantial (c. 1/3 of the glycolytic flux). The second experiment combined the medium switch with a shiftup in the glucose feeding rate (dilution rate shiftup from 0.05 to 0.10 h(-1)). This experiment triggered a strong but transient mobilization of storage carbon, that was channelled into glycolysis, causing a significant disruption in the dynamic labeling profile of glycolytic intermediates. The off-gas measurements in the shiftup experiment confirmed a considerable transient influx of (12)C-carbon into glycolysis after the combined medium switch and dilution rate shiftup. This study shows that for accurate in vivo kinetic interpretation of rapid pulse experiments, glycogen and trehalose metabolism must be taken into account.

Quantitative Metabolomics and Instationary 13C-Metabolic Flux Analysis Reveals Impact of Recombinant Protein Production on Trehalose and Energy Metabolism in Pichia pastoris

“…The glycolytic intermediates reached isotopic steady states after about 10–15 minutes, which is high compared to the turnover time (data not shown) but a commonly observed delay [25,36,37]. The delay can be explained by exchange fluxes with big pools of trehalose and storage polymers such as glycogen.…”

Carbon Flux Analysis by 13 C Nuclear Magnetic Resonance To Determine the Effect of CO 2 on Anaerobic Succinate Production by Corynebacterium glutamicum

“…C. glutamicum can store glycogen and maltodextrins during growth (45), as well as trehalose (46), and the catabolism of these components concomitantly with the metabolism of added labeled glucose leads to 13 C dilution, as demonstrated by the presence of unlabeled end products when uniformly labeled glucose was provided. This phenomenon has been observed in bacteria and yeast, which showed fast mobilization of carbohydrate storage pools (47,48). Moreover, the reverse flux from labeled glucose to reserves was apparent from the buildup of labeled glycogen and trehalose (see Fig.…”

Fast “Feast/Famine” Cycles for Studying Microbial Physiology Under Dynamic Conditions: A Case Study with Saccharomyces cerevisiae

“…It is frequently reported that the storage carbohydrates react rapidly to dynamic conditions, even under mild perturbations [33], thus during the feast/famine it might be expected these metabolites to exhibit a strong dynamic response. The metabolism of two major storage carbohydrates, trehalose and glycogen, was monitored through their intermediates (Figure 7).…”