The stereochemistry at carbon 3 of pyruvate lyase condensation products. Aldolases forming condensation products racemic at carbon 4.

Both the Schiff base-mediated 2-keto-4-hydroxyglutarate aldolase of liver and the metal-activated a-keto-/3-deoxyhexarate aldolase of Escherichia coli condense pyruvate and glyoxylate to form (4-RS)-Z-keto-4-hydroxyglutarate. Both enzymes catalyzed a condensation between [3-3H3]pyruvate and glyoxylate. It was found that C-C synthesis occurred 6 to 7 times faster than C-T bond cleavage. Thus, pyruvate deprotonation appeared to be partially rate-limiting in C-C synthesis. Condensations were carried out between (3R)[3-3H,2H,Hlpyruvate or (3S)-[3-3H,2H,H]pyruvate to form (4-RS)-2-keto-4-hydroxy-[3-3H, 2H]glutarate which was decarboxylated to DL-[3-3H, 2H]malate. The prochirality of malate’s C-3 was analyzed by determining tritium release catalyzed by fumarase and maleate hydratase. It was found that malate derived from (3R)-[3-3H,2H,H]pyruvate was of (3R)-33H, 2H-configuration while that derived from (3S)[3-3H, 2H,H]pyruvate was of (3S)-3-3H,2H-configuration. Comparison with stereochemical models shows that the configuration at C-3 of pyruvate is thus retained during C-C synthesis catalyzed by the two aldolases studied, i.e. both the exchanging proton and aldehyde approach the same face of bound enolpyruvate. A stereochemical mechanism explaining the synthesis of racemic condensation products is discussed.