Non-enzymic enolisation

1 Non-enzymic Enolisation. The carbanion chemistry of aldoses and ketoses themselves is masked by the ring opening step and mechanistic work is limited. However, a detailed examination has been made of the enolisation of L-glyceraldehyde 3-phosphate, which cannot form monomeric rings. The use of the unnatural (l) enantiomer enabled any racemisation to the natural 

D-isomer to be monitored by enzyme-linked oxidation of NADH. The same technique was used to monitor the formation of dihydroxyacetone phosphate. The major reaction on formation of the enediolate, however, is loss of phosphate to give the enol of methylglyoxal. The formation of the enediolate is independent of pH between pH 6 and 10 because of intramolecular proton abstraction by the phosphate dianion. Above pH 10 and below pH 6, the pH-rate profile has a gradient of +1.0, in the former case a reflection of direct attack by OH on the phosphate dianion, in the latter a reflection of the proportion of reactive dianionic form of the substrate present. 

5-phospho-D-ribose, as bound to ribose-5-phosphate isomerase 

A single base mechanism, with glutamate as the base, is common to most aldose-ketose phosphate isomerases, but the electrophilic machinery to stabilise the enolate appears to differ from enzyme to enzyme. TIM uses His95 in its 

Ribose-5-phosphate isomerase, which catalyses the interconversion of ribose-5-phosphate and ribulose-5-phosphate, activates the carbonyl differently. The aldose substrate can form a furanose (but not a pyranose) ring, but the furanose forms are so disfavoured (see Table 1.1) that there are likely to be appreciable amounts of open-chain sugar present at equilibrium. Moreover, this equilibrium, involving only furanose forms whose opening is possibly assisted by general acid catalysis from the 5-phosphate, is likely to be achieved rapidly. Although the enzyme from other sources crystallise with ring forms of the substrate bound, that from Thermus thermophilus crystallises with both ribose- 

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