@article{oai:soar-ir.repo.nii.ac.jp:00013455, author = {Taguchi, G and Imura, H and Maeda, Y and Kodaira, R and Hayashida, N and Shimosaka, M and Okazaki, M}, issue = {1}, journal = {Plant Science}, month = {}, note = {The enzyme UDP-glucose: hydroxycoumarin 7-O-glucosyltransferase (CGTase), which catalyzes the formation of scopolin from scopoletin, was purified approximately 1200-fold from a culture of 2,4-D-treated tobacco cells (Nicotiana tabacum L. cv. Bright Yellow T-13) with a yield of 7%. Purification to apparent homogeneity, as judged by SDS-PAGE, was achieved by sequential anion-exchange chromatography, hydroxyapatite chromatography, gel filtration, a second round of anion-exchange chromatography, and affinity chromatography on UDP-glucuronic acid agarose. The purified enzyme had a pH optimum of 7.5, an isoelectric point (pI) of 5.0, and a molecular mass of 49 kDa. The enzyme did not require metal cofactors for activity. Its activity was inhibited by Zn2+, Co2+ and Cu2+ ions, as well as by SH-blocking reagents. The K-m values for UDP-glucose, scopoletin and esculetin were 43, 150 and 25 mu M. respectively. A study of the initial rate of the reaction suggested that the reaction proceeded via a sequential mechanism. The purified enzyme preferred hydroxycoumarins as substrates but also exhibited significant activity with flavonoids. A database search using the amino terminus amino acid sequence of CGTase revealed strong homology to the amino acid sequences of other glucosyltransferases in plants., Article, Plant Science. 157(1):105-112 (2000)}, pages = {105--112}, title = {Purification and characterization of UDP-glucose: hydroxycoumarin 7-O-glucosyltransferase, with broad substrate specificity from tobacco cultured cells}, volume = {157}, year = {2000} }