Mandelate racemase enolization

If Ke signifies the concentrations of enol and keto tautomers in a carboxylic acid, then pfCE is the difference between the pKa values of the a-protons of the keto tautomer and the hydroxyl group of the enol tautomer. pfCE, however, is also the difference in pfCa values between the a-protons and the proton of the carbonyl group of the carbonyl-protonated acid, that is deemed to be decisive (Gerlt, 1991) for the kinetics of abstraction of a proton, rather than the pfQ value of the substrate in solution. The pki of mandelic acid (15.4) links the pka of the a-proton of the keto tautomer (22.0) with the pK.d value of the enol tautomer (6.6). The pk, value also links the pka value of the a-proton with that of the carbonyl-bound proton of the protonated mandelic acid. If the pka value of the carbonyl-bound proton of the protonated mandelic acid is assumed to be about -8.0 then the pfQ value of the a-proton is about 7.4. This value matches well with the pka -values of Lys and His residues which have been assigned recently in the active center of mandelate race-mase, so electrophilic catalysis alone is able to explain the catalytic power of mandelate racemase. 

X 10 if the enolate anion intermediate were not stabilized in the active site this value is 10 -fold less than the observed value for the kcat, 500 s. Recall that an enolate anion is necessarily on the reaction coordinate, so the value of AG° must be reduced for the enolate anion to be kinetically competent irrespective of whether AG int can be reduced. Thus, the active site of mandelate racemase must decrease AG° from the value predicted from the values of the substrate carbon acid and the active site base in solution. The obvious strategy to accomplish this reduction is preferential stabilization of the enolate anion intermediate relative to the carbon acid substrate, the increased negative charge on (or proton affinity of) the carbonyl/carboxylate oxygen of the enolate anion intermediate provides a convenient handle for enhanced electrostatic or hydrogen bonding interactions with the active site. 

Figure 17-20. Mechanism of the reaction catalyzed by mandelate racemase with concerted general acid-general base through an enolic intermediate. Reprinted from Mitra et al. 1981.

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