Molecular cloning and characterization of the Strictosidine synthase Gene from Rauwolfia verticillata

Abstract

The strictosidine synthase (STR, EC 4.3.3.2) catalyzes the condensation of tryptamine and secologanin to form strictosidine, which is the universal precursor for a wide range of pharmaceutical terpenoid indole alkaloids (TIAs). The full-length cDNA encoding STR was cloned and characterized from Rauwolfia verticillata a Chinese native plant producing TIAs, such as reserpine and ajmalicine. The new cDNA was designed as RvSTR and submitted to GenBank to get an accession number DQ017054. The full-length cDNA of RvSTR was 1211 bp containing a 1035-bp open reading frame encoding a deduced 344-amino-acid polypeptide with a calculated mol wt of 38.2 kD and an isoelectric point of 5.19. Comparative and bioinformatic analysis revealed that RvSTR showed a higher similarity to STRs from Apocynaceae species, including Catharanthus roseus and Ophiorrhiza pumila, but a relatively lower similarity to other plant STRs. The unique essential catalytic residue Glu-309 was conserved in all alignment plant species. The phylogenetic analysis revealed that STRs were divided into two groups, including plant and bacterial enzymes. The tissue expression pattern analysis indicated that RvSTR expression could be detected in all tested organs of R. verticillata, including roots, stems, leaves, fruits, and flowers. The lowest transcription level was observed in flowers and the highest was found in fruits; subsequently, the order of transcription level decrease was stems > roots > leaves. The cloning and characterization of RvSTR give a new STR sequence involved in TIA biosynthesis of R. verticillata, and provide a candidate gene for metabolic engineering of the TIA pathway in R. verticillata.