Underestimation of D-glucose phosphorylation as measured by 3H2O production from D-[2-3H]glucose

Area of Science:

• Biochemistry
• Cell Biology
• Metabolic Research

Background:

• Hexose phosphorylation is a critical initial step in glucose metabolism.
• Assessing hexose phosphorylation rates is vital for understanding cellular energy production and metabolic disorders.
• Tritiated water (3H2O) production from labeled glucose has been used as a proxy for hexose phosphorylation.

Purpose of the Study:

• To investigate the reliability of using D-[2-3H]glucose to measure hexose phosphorylation rates.
• To explore the mechanism behind observed discrepancies in 3H2O production from different labeled glucose isotopes.

Main Methods:

• Incubation of rat pancreatic islets, tumoral islet cells (RINm5F line), parotid gland, and human erythrocytes with D-[2-3H]glucose and D-[5-3H]glucose.
• Measurement of 3H2O production.
• Analysis of tritiated lactic acid formation.

Main Results:

• 3H2O production from D-[2-3H]glucose was 20-30% lower than from D-[5-3H]glucose in rat pancreatic islets, RINm5F cells, parotid gland, and human erythrocytes.
• This difference was not observed in rat hepatocytes.
• Tritiated lactic acid was produced from D-[2-3H]glucose, suggesting intramolecular hydrogen transfer during the phosphoglucoisomerase reaction.

Conclusions:

• The production of 3H2O from D-[2-3H]glucose is not a universally reliable method for assessing the total rate of hexose phosphorylation.
• An intramolecular hydrogen transfer in the phosphoglucoisomerase reaction likely contributes to the observed lower 3H2O yield from D-[2-3H]glucose.
• Alternative methods or careful consideration of isotopic labeling are necessary for accurate hexose phosphorylation measurements in specific cell types.