Evidence for Enzymatic 5-Hydroxylation of Indole-3-acetic Acid in vitro by Extracts of Sedum morganianum

Summary The kinetic properties of a soluble enzyme preparation from Sedum morganianum have been studied using both tryptophan and indole-3-acetic acid (IAA) as substrates. This soluble enzyme preparation utilized tryptophan and IAA as substrates forming 5-hydroxytryptophan (5-HTP) and 5-hydroxyindole-3-acetic acid (5-HIAA) respectively although the affinity for IAA was much lower (K m = 208.8 ± 8.5 μM IAA) as opposed to tryptophan (K m = 102.5 ± 8.7 μM L-tryptophan) Furthermore, endogenous serotonin (5-hydroxytryptamine; 5-HT) and 5-HIAA were identified in the leaves of this species. Both spectrophotofluorometric and thin layer chromatographic identification of enzyme reaction products indicate that tryptophan is converted to 5-HTP and IAA is converted to 5-HIAA. Tryptophan hydroxylating activity and IAA hydroxylating activity show a maximum at pH 7.5 and pH 8.0 respectively. These results demonstrate that an extract from Sedum morganianum leaves is capable of hydroxylating both tryptophan and IAA in vitro and that these reaction products are present in the plant tissue.