Abstract
For further clarification of the role of L-ascorbic acid (AsA) in the proline hydroxylation reaction, the specificity of AsA for the decarboxylation of α-keto-glutarate (KGA) was studied using various reductants AsA and its structural analogs. Decarboxylation of KGA was not observed in the absence of AsA. Erythorbic acid (ErA) was found to be as effective as AsA, and D-ascorbic acid was almost as effective as AsA in the reaction, whereas, thiol compounds showed a very slight accelerating effect on the decarboxylation of KGA. L-scorbamic acid (SCA) or erythroscorbamic acid (ErS), at a concentration 10 folds greater than AsA, showed a decarboxylation level of 40-45% that of AsA. Furthermore, in the presence of AsA, the pH-dependence and concentration effect on the decarboxylation of KGA were different from those in the presence of SCA. Moreover, the Lineweaver-Burk plot of the inhibition by SCA of AsA showed that the mode of interac-tion of SCA with AsA may be apparently noncompetitive. From these results, it is suggested that, due to its plane γ-lactone ring system with an endiol group, AsA is a specifically suitable reducing compound for the proline hydroxylation. AsA is considered to be most effective in its approaching and binding to the enzyme active site and reducing the enzyme bound Fe^<3+>. In this study, it was also found that the uncoupled reaction inevitable occurred during proline hydroxylation and this reaction was accompanied by the oxidation of AsA, thus leading to its consumption.
