Clinton R. Smith

Intramolecular catalysis of amide hydrolysis by the carboxy-group. Rate determining proton transfer from external general acids in the hydrolysis of substituted maleamic acids

The highly efficient intramolecular catalysis by the carboxy-group of the hydrolysis of simple dialkylmaleamic acids is itself subject to external general acid catalysis. The kinetic characteristics of the general acid catalysed reaction are those expected for a diffusion-controlled proton transfer. At high concentrations of general acid, external catalysis disappears. This is shown to result from a change in rate-determining step, and is thus evidence for an intermediate on the reaction pathway. The intermediate can only reasonably be a tetrahedral addition intermediate. Kinetic evidence is now available for all the major steps on the reaction pathway, and the requirements for an enzyme catalyst carrying out the reaction can be specified in detail. The full mechanism specifically implicates the O-protonated amide as the reactive species in dilute acid.

Intramolecular catalysis of amide hydrolysis by two carboxy-groups

In di-isopropylmaleamic acids derived from β-amino-acids (i) the rate constant for hydrolysis of the unactivated amide group is comparable with Kcat for pepsin hydrolysing good syntheic substrates; (ii) a neighbouring carboxy-group participates as a nucleophilic catalyst, but requires a specific structural relationship with the amide group to reach high efficiency; (iii) there is a requirement for a second carboxy-group, also in a special structural relationship with the amide group, but this time in the ionised form, for full catalytic efficiency to be maintained. Remarkably, the ionised carboxy-group acts as a general base to catalyse the interconversion of neutral and zwitterionic forms of the tetrahedral carbinolamine intermediate, a reaction which is catalysed by external general acids. As a result we have developed a system showing a bell-shaped pH–rate profile for hydrolysis, of the sort shown by many enzyme-catalysed reactions, and for the same reasons.