Klonierung von O-Methyltransferasen zur Furanocumarinbiosynthese in Ammi majus L.

Vertreter der Apiaceae oder Rutaceae akkumulieren methoxylierte Psoralene wie Bergapten oder Xanthotoxin als Endprodukte der Furanocumarinbiosynthese. O-Methyltransferase-Aktivitäten mit Umsetzungen von Bergaptol zu Bergapten bzw. Xanthotoxol zu Xanthotoxin wurden bereits aus induzierten Zellkulturs...

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Bibliographic Details
Main Author: Hehmann, Marc
Contributors: Matern, Ulrich (Prof. Dr.) (Thesis advisor)
Format: Doctoral Thesis
Published: Philipps-Universität Marburg 2004
Online Access:PDF Full Text
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Table of Contents: Plants belonging to the Apiaceae or Rutaceae accumulate methoxylated psoralens, such as bergapten or xanthotoxin, as the final products of their furanocumarin biosynthesis, and the rate of accumulation depends on environmental and other cues. Distinct O-methyltransferase activities had been reported to methylate bergaptol to bergapten and xanthotoxol to xanthotoxin, from induced cell cultures of Ruta graveolens, Petroselinum crispum and Ammi majus. Bergaptol 5-O-methyltransferase (BMT) cDNA was cloned from dark-grown Ammi majus L. cells treated with a crude fungal elicitor. The translated polypeptide revealed considerable sequence similarity to heterologous caffeic acid 3-O-methyltransferases (COMTs). For homologous comparison, COMT was cloned from A. majus plants and shown a 64% identity with the BMT sequence at the polypeptide level. Functional expression of both enzymes in Escherichia coli revealed that the BMT activity in the bacterial extracts was labile and rapidly lost on purification, whereas theCOMT activity remained stable. Furthermore, the recombinant AmBMT showed narrow substrate specificity for bergaptol, while the AMCOMT accepted 5-hydroxyferulic acid, esculetin and other substrates. The AmBMT sequence thus represents a novel database accession specific for the biosynthesis of psoralens.