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Updated: Jul 20, 2026

Extraction and Quantification of Soluble, Radiolabeled Inositol Polyphosphates from Different Plant Species using SAX-HPLC
Published on: June 26, 2020
Isoprenoid biosynthesis in plants - 2C-methyl-D-erythritol-4-phosphate synthase (IspC protein) of Arabidopsis
Felix Rohdich1, Susan Lauw, Johannes Kaiser
1Lehrstuhl für Organische Chemie und Biochemie, Technische Universität München, Garching, Germany. felix.rohdich@ch.tum.de
Abstract:
The ispC gene of Arabidopsis thaliana was expressed in pseudomature form without the putative plastid-targeting sequence in a recombinant Escherichia coli strain. The recombinant protein was purified by affinity chromatography and was shown to catalyze the formation of 2C-methyl-D-erythritol 4-phosphate from 1-deoxy-D-xylulose 5-phosphate at a rate of 5.6 micromol x min(-1) x mg(-1) (k(cat) 4.4 s(-1)). The Michaelis constants for 1-deoxy-D-xylulose 5-phosphate and the cosubstrate NADPH are 132 and 30 microm, respectively. The enzyme has an absolute requirement for divalent metal ions, preferably Mn2+ and Mg2+, and is inhibited by fosmidomycin with a Ki of 85 nm. The pH optimum is 8.0. NADH can substitute for NADPH, albeit at a low rate (14% as compared to NADPH). The enzyme catalyzes the reverse reaction at a rate of 2.1 micromol x min(-1) x mg(-1).
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