2-KETO-3-DEOXY-6-PHOSPHOGLUCONATE ALDOLASES AS CATALYSTS FOR STEREOCONTROLLED CARBON-CARBON BOND FORMATION

The pyruvate aldolases use pyruvate as the nucleophilic component in s tereoselective aldol condensations, producing a 4-hydroxy-2-ketobutyra te framework. We have examined the 2-keto-3-deoxy-6-phosphogluconate ( KDPG) aldolases from Pseudomonas putida, Escherichia coli, and Zymomon as mobilis for utility as synthetic reagents. Unlike other pyruvate al dolases examined to date, the KDPG aldolases accept short-chain, non-c arbohydrate electrophilic aldehydes as substrates, providing a general methodology for the construction of the 4-hydroxy-2-ketobutyrate skel eton. The three aldolases differ markedly with respect to enzyme stabi lity, pH optima, stability in organic cosolvent mixtures, substrate sp ecificity, and diastereoselectivity during aldol condensation. All thr ee enzymes show broad substrate specificity with regard to the electro philic component. The primary requirements for substrate activity appe ar to be minimal steric hindrance and the presence of electron-withdra wing substituents at C2. The aldolases from Pseudomonas and Escherichi a are also specific for the D-stereochemical configuration at C2, whil e the enzyme from Zymomonas displays no stereochemical discrimination with regard to the electrophilic substrate. Nucleophiles other than py ruvate are accepted as nucleophilic substrates by all three enzymes, p rovided the electrophile is sufficiently reactive, In preparative scal e reactions with three unnatural electrophiles, the three enzymes show varying degrees of stereochemical fidelity, In most cases, a single d iastereomer of the aldol adduct was produced, although in one case, a diastereomeric excess of 50% was observed. In all cases, the diastereo selectivity is exclusively kinetic in origin, despite the reversibilit y of some reactions. The enzymes are remarkably tolerant of added coso lvent: all three showed >60% of native activity in 30% DMSO and DMF. B y appropriate choice of enzyme, the KDPG aldolases offer exceptional u tility for stereocontrolled carbon-carbon bond formation under a wide range of experimental conditions.