Enzymatic synthesis of L-5-Hydroxytryptophan from L-serine and 5-hydroxyindole catalyzed by tryptophan synthase from recombinant Escherichia coli were studied. The factors such as temperature, pH, molar ratio of L-serine to 5-hydroxyindole and substrate concentration were investigated. The optimal temperature and pH value were 35 篊 and 9, respectively. The optimal molar ratio of 5-hydroxyindole to L-serine was 1.1: 1. The optimal substrate concentration of L-serine was 200 mmol/L. Under the optimal conditions, bioconversion rate of L-serine reached 86% after 18 h.