Biosynthesis of terpenoids: 4-Diphosphocytidyl-2C-methyl-d-erythritol synthase of Arabidopsis thaliana
- Felix Rohdich*,
- Juraithip Wungsintaweekul†,
- Wolfgang Eisenreich*,
- Gerald Richter*,
- Christoph A. Schuhr*,
- Stefan Hecht*,
- Meinhart H. Zenk†, and
- Adelbert Bacher*,‡
- *Lehrstuhl für Organische Chemie und Biochemie, Technische Universität München, Lichtenbergstrasse 4, D-85747 Garching, Germany; and †Biozentrum–Pharmazie, Universität Halle, Weinbergweg 22, D-06120 Halle, Germany
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Contributed by Meinhart H. Zenk
Abstract
A hypothetical gene with similarity to the ispD gene of Escherichia coli was cloned from Arabidopsis thaliana cDNA. The ORF of 909 bp specifies a protein of 302 amino acid residues. The cognate chromosomal gene consists of 2,071 bp and comprises 11 introns with a size range of 78–202 bp. A fragment comprising amino acid residues 76–302 was expressed in a recombinant E. coli strain. The protein was purified to homogeneity and was shown to catalyze the formation of 4-diphosphocytidyl-2C-methyl-d-erythritol from 2C-methyl-d-erythritol 4-phosphate with a specific activity of 67 μmol⋅min−1 mg−1. The Michaelis constants for 4-diphosphocytidyl-2C-methyl-d-erythritol and CTP were 500 μM and 114 μM, respectively.
