title: Structure and Function of the Golgi Complex in Rice Cells (II. Purification and Characterization of Golgi Membrane-Bound Nucleoside Diphosphatase).
abstract: Inosine diphosphatase bound to Golgi membranes was studied in rice (Oryza sativa L. cv Nipponkai) cells. The enzyme was solubilized with Triton X-100 from isolated rice Golgi membranes and was highly purified employing a series of chromatography steps in the presence of 20% glycerol and 0.1% Triton X-100. The apparent molecular mass of the enzyme was estimated by gel filtration column chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis to be 200 and 55 kD, respectively. The isoelectric point of the enzyme was determined to be 7.5. The optimal pH for the enzyme activity was around 7 and the enzyme required Mg2+ for hydrolyzing activity. IDP, UDP, and GDP were effective substrate for the purified rice Golgi membrane-bound inosine diphosphatase, whereas activity with ADP, CDP, and thymidine 5[prime]-diphosphate was 10 to 20% of IDP. The Km values for IDP, UDP, and GDP were 0.48, 0.50, and 0.67 mM, respectively, and Vmax values were 1.85, 1.54, and 1.67 [mu]mol min-1 mg-1, respectively. These results indicate that the rice Golgi enzyme is a nucleoside diphosphatase that is specific for IDP, UDP, and GDP. Furthermore, this rice Golgi nucleoside diphosphatase stimulated the activity of glucan synthase I also localized in rice Golgi membranes. The results strongly support the view that this nucleoside diphosphatase is involved in regulation of [beta]-glucan synthesis in the plant Golgi complex.
authors.affiliation: Department of Applied Biological Chemistry, Faculty of Agriculture, Niigata University, Niigata 950-21, Japan.
authors.forename: T.
authors.initials: T
authors.lastname: Mitsui
authors.forename: M.
authors.initials: M
authors.lastname: Honma
authors.forename: T.
authors.initials: T
authors.lastname: Kondo
authors.forename: N.
authors.initials: N
authors.lastname: Hashimoto
authors.forename: S.
authors.initials: S
authors.lastname: Kimura
authors.forename: I.
authors.initials: I
authors.lastname: Igaue