Related Pyruvate Aldolases

has broad substrate specificity for aldoses while pyruvate was found to be irreplaceable. As a notable distinction, KdoA was also active on smaller acceptors such as glyceraldehyde. Preparative applications, for example, for the synthesis of KDO (enf-6) and its homologs or analogs (16)/(17), suffer from an unfavorable equilibrium constant of 13 in direction of synthesis [34]. The stereochemical course of aldol additions generally seems to adhere to a re-face attack on the aldehyde carbonyl, which is complementary to the stereoselectivity of NeuA. On the basis of the results published so far, it may be concluded that a (31 )-configuration is necessary (but not sufficient), and that stereochemical requirements at C-2 are less stringent [71]. 

2-keto-3-deoxy-6-phospho-D-galactonate (20) (KDPGa] aldolase EC 4.1.2.21), was also studied for synthetic applications [83]. 

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Wang, W. J., Baker, R, and Seah, S. Y. K., Comparison of two metal-dependent pyruvate aldolases related by convergent evolution Substrate specificity, kinetic mechanism, and substrate charmeling. Biochemistry 2010,49 (17), 3774-3782. 

Commercial A -acetylneuraminic acid aldolase from Clostridium perfringens (NeuAcA EC 4.1.3.3) catalyzes the addition of pyruvate to A-acetyl-D-mannosamine. A number of sialic acid related carbohydrates are obtained with the natural substrate22"24 or via replacement by aldose derivatives containing modifications at positions C-2, -4, or -6 (Table 4)22,23,25 26. Generally, a high level of asymmetric induction is retained, with the exception of D-arabinose (epimeric at C-3) where stereorandom product formation occurs 25 2t The unfavorable equilibrium constant requires that the reaction must be driven forward by using an excess of one of the components in order to achieve satisfactory conversion (preferably 7-10 equivalents of pyruvate, for economic reasons). 

Comparable to the situation for the sialic acid and KDO lyases (vide supra), sets of stereochemically complementary pyruvate lyases are known, e,g. in Pseudomonas strains, which act on related 2-keto-3-deoxy-aldonic acids [112]. The enzymes cleaving six-carbon sugar acid phosphates—the KdgA and 2-keto-3-deoxy-6-phospho-D-galactonate (20) aldolases (KDPGal aldolase EC 4.1.2.21) [139] — are typified as class I enzymes, whereas those acting on non-phosphorylated five-carbon substrates — 2-keto-3-deoxy-L-arabonate (21) (KDAra aldolase EC 4.1.2,18) [140, 141] and 2-keto-3-deoxy-D-xylonate (22)... 

In considering the application of enzyme catalysis to DCC, we were encouraged by the thermodynamic resolution of a dynamic mixture of aldol products by Whitesides and co-workers through the use of a broad-specificity aldolase to lead to reversible formation of carbon-carbon bonds under mild conditions.35 For the current investigation36 we chose a related enzyme, N-acetylneuraminic acid aldolase (NANA aldolase, EC 4.1.3.3), which catalyzes the cleavage of N-acetylneuraminic acid (sialic acid, 27a) to A-acetylmannosamine (ManNAc, 28a), and sodium pyruvate 29 in the presence of excess sodium pyruvate, aldol products 27a-c are generated from... 

In a related protocol, the acetaldehyde trimer 54 from the generic RibA oligomerization was found to be a substrate for the N-acetylneuraminic acid aldolase (NeuA EC 4.1.3.3) which catalyzed the addition of pyruvate. By this means, a tetradeoxy-L-arahi o-2-nonulosonic acid 56 was obtained in 55% yield [116]. A one-pot, tandem operation was complicated by the fact that temperature requirements for optimum activity and stability of the two catalysts were not compatible. 

The aldolases which have been investigated for their synthetic utility can be classified on the basis of the donor substrate accepted by the enzyme. For the synthesis of 3-deoxy-2-ulosonic acids pyruvate- and phosphoenolpyruvate dependent aldolases are the most desirable enzymes as they are involved in the metabolism of sialic acids (or structurally related ones) in vivo. They use pyruvate or phosphoenolpyruvate as a donor to form 3-deoxy-2-keto acids (Table 1). Both of them add a three-carbons ketone fragment onto a carbonyl group of an aldehyde. The pyruvate dependent aldolases have a catabolic function in vivo, whereas the phosphoenolpyruvate dependent aldolases are involved in the biosynthesis of the keto acids. For synthetic purpose the equilibrium of the pyruvate dependent aldolases is shifted toward the condensation products through the use of an excess of pyruvate. 

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