Dynamic interactions of enzymes involved in triosephosphate metabolism

A steady‐state kinetic analysis of the coupled reactions catalysed by the three‐enzyme system, aldolase, glyceraldehyde‐3‐phosphate dehydrogenase and triosephosphate isomerase, was performed. The kinetic parameters of the progress curves of end‐product formation calculated for noninteracting enzymes were compared with those measured in the two‐enzyme and three‐enzyme systems. Changes in the fluorescence anisotrophy of labelled dehydrogenase upon addition of aldolase and/or isomerase were also measured.Glyceraldehyde‐3‐phosphate oxidation catalysed by glyceraldehyde‐3‐phosphate dehydrogenase in the presence of isomerase (which ensures rapid equilibration of the triosephosphates) follows single first‐order kinetics. The rate constant depends simply on the concentration of the dehydrogenase, indicating no kinetically significant isomerase‐dehydrogenase interaction. Fluorescence anisotropy measurements also fail to reveal complex formation between the two enzymes.The steady‐state velocity of 3‐phosphoglycerate formation from fructose 1,6‐bisphosphate in the reactions catalysed by aldolase and dehydrogenase is not increased twofold on addition of the isomerase, even though a 1:2 stoichi