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Measuring the Activity of 1-Deoxy-D-Xylulose 5-Phosphate Synthase, the First Enzyme in the MEP Pathway, in Plant Extracts

  • Louwrance P. WrightEmail author
  • Michael A. Phillips
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1153)

Abstract

The first enzyme in the methylerythritol phosphate (MEP) pathway is 1-deoxy-d-xylulose 5-phosphate (DXP) synthase (DXS). As such this enzyme is considered to be important in the control of plastidial isoprenoid production. Measuring the activity of DXS in plant extracts is therefore crucial to understanding the regulation of the MEP pathway. Due to the relatively low amounts of DXS, the activity of this enzyme can only be measured using highly sensitive analytical equipment. Here, a method is described to determine the DXS enzyme activity in a crude plant extract, by measuring DXP production directly using high performance liquid chromatography linked to a tandem triple quadrupole mass spectrometry detector (LC-MS/MS).

Keywords

DXS Enzyme assay Isoprenoid biosynthesis LC-MS/MS 

Notes

Acknowledgments

We thank Bettina Raguschke for technical assistance and Felix Rohdich, Adelbert Bacher and Wolfgang Eisenreich for the kind gift of 13C labelled DXP. The project was funded by the Max Planck Gesellschaft.

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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  1. 1.Department of BiochemistryMax Planck Institute for Chemical EcologyJenaGermany
  2. 2.Centre for Research in Agricultural Genomics (CRAG)CSIC-IRTA-UAB-UBBarcelonaSpain

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