Cloning and functional analysis of geraniol 10-hydroxylase, a cytochrome P450 from Swertia mussotii Franch

Biosci Biotechnol Biochem. 2010;74(8):1583-90. doi: 10.1271/bbb.100175. Epub 2010 Aug 7.

Abstract

Swertia mussotii Franch has anti-hepatitis activity and contains a high level of iridoid monoterpenoids. The cytochrome P450 monooxygenase (CYP) geraniol 10-hydroxylase (G10H) is thought to play an important role in iridoid monoterpenoid and indole alkaloid biosynthesis. Here we report the isolation of a full-length cDNA clone of S. mussotii G10H (SmG10H). The predicted gene product was a 496 residue protein designated CYP76B10, the sequence of which was highly similar to that of the CYP76 family, particularly to Catharanthus roseus G10H (80.2% homology). SmG10H transcripts were much more abundant in the leaves than in either the root or the stem, and were derived from a single copy gene. SmG10H expression was upregulated by treatment with methyljasmonate (MeJA) over a period from 6 h to 36 h after treatment. Accumulation of swertiamarin increased after elicitation by MeJA. SmG10H was heterologously expressed in both Escherichia coli and Pichia pastoris (yeast), forming a 55.5-kDa protein. Based on analysis in vitro, SmG10H was found to have catalytic activity hydroxylating geraniol. In the SmG10H overexpression plants, the level of SmG10H transcript and the contents of 10-hydroxygeraniol and swertiamarin increased simultaneously.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acetates / pharmacology
  • Amino Acid Sequence
  • Base Sequence
  • Cloning, Molecular
  • Cyclopentanes / pharmacology
  • Cytochrome P-450 Enzyme System / chemistry
  • Cytochrome P-450 Enzyme System / genetics*
  • Cytochrome P-450 Enzyme System / metabolism*
  • DNA, Complementary / genetics
  • Gene Expression Regulation, Plant / drug effects
  • Iridoid Glucosides / metabolism
  • Molecular Sequence Data
  • Oxylipins / pharmacology
  • Pyrones / metabolism
  • Swertia / drug effects
  • Swertia / enzymology*
  • Swertia / genetics*
  • Swertia / metabolism

Substances

  • Acetates
  • Cyclopentanes
  • DNA, Complementary
  • Iridoid Glucosides
  • Oxylipins
  • Pyrones
  • swertiamarin
  • methyl jasmonate
  • Cytochrome P-450 Enzyme System
  • geraniol 10-hydroxylase

Associated data

  • GENBANK/GU168041