Molecular cloning, expression, and induction of berberine bridge enzyme, an enzyme essential to the formation of benzophenanthridine alkaloids in the response of plants to pathogenic attack

H. Dittrich, T. M. Kutchan

Research output: Contribution to journalArticlepeer-review

244 Scopus citations

Abstract

The berberine bridge enzyme [(S)-reticuline: oxygen oxidoreductase (methylene-bridge-forming), EC 1.5.3.9] is a vesicular plant enzyme that catalyzes the formation if the berberine bridgehead carbon of (S)-scoulerine from the N-methyl carbon of (S)-reticuline in a specific, unparalleled reaction along the biosynthetic pathway that leads to bensophenanthridine alkaloids. Cytotoxic benzophenanthridine alkaloids are accumulated in certain species of Papaveraceae and Fumorioceoe in response to pathogenic attack and, therefore, function as phytoalexins. The berberine bridge enzyme has been purified to homogeneity from elicited cell-suspension culture of Eschscholtzio californien, and partial amino acid foences have been determined. A cDNA, isolated from a Agt11 cDNA bank of elicited E. californica cell-suspension cultures, coded for an open reading frame of 538 amino acids. The first 22 amino acids constitute the putative signal peptide. The mature protein has a Mr of 57,352, excluding carbohydrate. The berberine bridge enzyme was heterologously expressed in a catalytically active form in Saccharomyces cerevilisiae. Southern hybridization with genomic DNA suggests that there is only one gene for the enzyme in the E. californica genome. Hybridized RNA blots from elicited E. californica cell-suspension cultures revealed a rapid and transient increase in poly(A)+ RNA levels that preceded both the increase in enzyme activity and the accumulation of benzophenanthridine alkaloids, emphasizing the integral role of the berberine bridge enzyme in the plant response to pathogens.

Original languageEnglish
Pages (from-to)9969-9973
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume88
Issue number22
StatePublished - 1991

Keywords

  • Elicitation
  • Eschscholtzia californica
  • Heterologous expression
  • Phytoalexin biosynthesis

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