Cell-free biosynthesis of cytokinins in cultured tobacco cells

Abstract

Cell-free biosynthesis of free cytokinins not related to the degradation of t-RNA was demonstrated by using enzyme preparation extracted from cultured tobacco (Nicotiana tabacum cv. Xanthi) cells. The reaction mixture for cytokinin biosynthesis contained 20 mM Tris-HCl buffer (pH 7.6), 5 mM magnesium acetate, 0.5 mM isopentenylpyrophosphate, 50 μM adenosine as a substrate and dialyzed enzyme extract. After 2.5 hours of reaction at 27°C, N6-(Δ2-isopentenyl)adenosine, N6-(Δ2-isopentenyl)adenine and trans-zeatin were biosynthesized. The radioactivity of [U-14C]adenosine was incorporated into these three cytokinins after 30 minutes of reaction in the cell-free system. From the time course of biosynthesis of cytokinins and the incorporation of [U-14C]adenosine into cytokinins, we concluded that adenosine accepts the isopentenyl group directly to form N6-(Δ2-isopentenyl) adenosine. This is converted to trans-zeatin via N6-(Δ2-isopentenyl)adenine by several enzymes.