A new method for enzymatic preparation of isopentenyladenine-type and trans-zeatin-type cytokinins with radioisotope-labeling

Byline: Kentaro Takei (1), Yasumasa Dekishima (2), Tadashi Eguchi (2), Tomoyuki Yamaya (1), Hitoshi Sakakibara (1) Keywords: Adenylate isopentenyltransferase; Cytokinin; Isopentenyladenine; Purine-nucleoside phosphorylase; Radioisotope labeling; trans-Zeatin Abstract: We describe a new enzymatic reaction method for the preparation of the radioisotope-labeled cytokinins isopentenyladenine (iP), trans-zeatin (tZ), and their ribosides. The method is based on the three enzyme activities of an adenylate isopentenyltransferase (IPT EC 2.5.1.27) from Arabidopsis thaliana, an alkaline phosphatase (EC 3.1.3.1) from calf intestine, and a purine-nucleoside phosphorylase (EC 2.4.2.1) from Escherichia coli. The A. thaliana IPT, AtIPT7, utilized both dimethylallyldiphosphate and 4-hydroxy-3-methyl-2-(E)-butenyl diphosphate as isoprenoid donors. The dual specificity of the substrates enabled us to produce iP-type and tZ-type cytokinins separately in the same system simply by switching the substrates. Our method affords a much higher yield of the labeled products than the chemical reaction methods previously used. These labeled compounds will be useful tools for cytokinin research, such as receptor--ligand assays and cell metabolism studies. Author Affiliation: (1) Plant Science Center, RIKEN (The Institute of Physical and Chemical Research), 1-7-22 Suehiro, Tsurumi, Yokohama 230-0045, Japan (2) Department of Chemistry and Materials Science, Tokyo Institute of Technology, O-okayama, Meguro, Tokyo 152-8551, Japan Article History: Received Date: 29/12/2002 Accepted Date: 06/03/2003 Online Date: 01/05/2003