This study examined glucose metabolism in Candida utilis CBS 621 grown in glucose-limited chemostat cultures to determine the contribution of the hexose monophosphate (HMP) pathway. Using radiorespirometric analysis with specifically labeled glucose (at positions 1 and 6), researchers tracked carbon flow through different metabolic pathways under varying nitrogen sources. When nitrate served as the nitrogen source, significantly more glucose flowed through the HMP pathway compared to ammonium, reflecting the higher NADPH requirement for nitrate reduction to ammonia. The HMP pathway contributed approximately 7% of glucose metabolism with ammonium and 21% with nitrate. Notably, increased recycling of hexose phosphates occurred with nitrate as the nitrogen source. The experimental findings closely matched theoretical minimum NADPH requirements for biosynthesis, suggesting the HMP pathway operates efficiently without excess capacity. Both steady-state substrate feeding and pulse-labeling methods confirmed these observations, with pulse-labeling on cell suspensions offering a faster alternative to continuous culture experiments.

Key findings

• HMP pathway flux through Candida utilis glucose metabolism increased approximately threefold during growth with nitrate versus ammonium, correlating with higher NADPH demands for nitrate reduction
• The HMP pathway contributed 7% of glucose metabolism with ammonium and 21% with nitrate, representing near-minimal levels required for biosynthetic NADPH
• Hexose phosphate recycling through the HMP pathway increased significantly with nitrate, confirmed by differential CO₂ release patterns from labeled substrates at different positions
• Pulse-labeling experiments on cell suspensions proved effective for estimating HMP pathway activity and provided comparable results to steady-state chemostat analysis with shorter experimental duration
• The C₁/C₆ ratio of ¹⁴CO₂ yields did not reliably indicate HMP pathway activity when glucose was diverted toward biosynthesis, as with glucose plus formate supplementation