Inhibition of transketolase by hexacyanoferrate(III)

Area of Science:

• Biochemistry
• Enzymology

Background:

• Transketolase is a key enzyme in the pentose phosphate pathway.
• Enzyme active sites are crucial for catalytic activity and substrate binding.
• Understanding enzyme active site properties is vital for drug development and metabolic research.

Purpose of the Study:

• To investigate the effect of hexacyanoferrate(III) on transketolase catalytic activity.
• To determine the selectivity of hexacyanoferrate(III) in targeting transketolase active sites.
• To explore the functional equivalence or nonequivalence of transketolase active sites.

Main Methods:

• Enzyme kinetics assays were performed to measure transketolase activity.
• Hexacyanoferrate(III) was used as an oxidant to probe active site accessibility and function.
• Comparative analysis of enzyme activity before and after oxidant treatment was conducted.

Main Results:

• Hexacyanoferrate(III) inactivated only one of the two active sites in transketolase.
• The inactivated active site demonstrated a higher affinity for the coenzyme, thiamine diphosphate.
• The second active site retained its full catalytic activity after exposure to the oxidant.

Conclusions:

• The active sites of holotransketolase are functionally nonequivalent, despite appearing identical in X-ray crystallographic data.
• This functional asymmetry suggests distinct roles or regulatory mechanisms for each active site.
• The findings provide new insights into the structural and functional complexity of enzymes.