This study investigates why hex2 mutants of the yeast Saccharomyces cerevisiae are inhibited by maltose, despite lacking defects in maltose permease or maltase enzyme levels. When maltose is added to hex2 cells growing on ethanol, it is rapidly taken up and hydrolyzed to glucose at rates exceeding glycolytic capacity. Using radioactive maltose labeling, NMR spectroscopy, and pH measurements, the researchers demonstrate that glucose accumulates intracellularly to toxic levels (up to 0.3 M) while glycolysis becomes blocked. Surprisingly, neither maltose permease nor maltase are overexpressed in hex2 cells compared to wild-type, indicating that enzyme abundance alone does not explain the toxicity. The intracellular pH remains stable, ruling out acidification as the cause. Instead, the massive glucose accumulation appears responsible for growth inhibition, protein synthesis arrest, and eventual cell death. The authors propose two mechanisms: osmotic damage from high cytoplasmic glucose concentration and non-specific glycosylation of proteins via Schiff base formation. Additionally, they show that accumulated glucose is excreted into the medium, providing evidence that at least one glucose transporter in yeast operates bidirectionally. These findings reveal an unknown regulatory mechanism controlled by HEX2 that coordinates maltose uptake with glycolytic capacity.

Key findings

• hex2 mutants accumulate intracellular glucose to toxic levels (up to 0.3 M) when exposed to maltose despite normal maltose permease and maltase expression levels
• Glucose accumulation causes metabolic paralysis including cessation of glycolysis, protein synthesis blockade, and growth inhibition in hex2 cells
• An unknown regulatory mechanism controlled by HEX2 coordinates maltose uptake and hydrolysis with glycolytic flux, operating independently of catabolic enzyme abundance
• Accumulated glucose is excreted from hex2 cells into the medium, demonstrating that at least one yeast glucose transporter functions bidirectionally
• Intracellular pH remains stable during maltose-induced toxicity, ruling out acidification as the primary mechanism of cell damage