Fructose-1,6-bisphosphate Aldolase from Rabbit Muscle. Kinetic Resolution of the Enamine Phosphate from the Enamine-Aldehyde Intermediate at Low Temperature

At or below −12 °C and in the presence of 40% ethylene glycol, only two out of the four dihydroxyacetone phosphate binding sites of aldolase are catalytically active. At these same temperatures and at pH* 8.3, the equilibrium between the pre-enamine and the enamine plus the post-enamine intermediates is largely shifted in favor of the latter. The enamine phosphate and the enamine-aldehyde phosphate intermediates have been resolved by studying the rate of their formation at −13 °C and pH* 5.28 and the trapping by dl-glyceraldehyde 3-phosphate at −24 °C and pH* 5.24.