PLANT PHYSIOLOGY , Vol 102, Issue 1 165-172, Copyright © 1993 by American Society of Plant Biologists
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CELL BIOLOGY AND SIGNAL TRANSDUCTION |
Purification and Characterization of Membrane-Bound Inositol Phospholipid-Specific Phospholipase C from Suspension-Cultured Rice (Oryza sativa L.) Cells (Identification of a Regulatory Factor)
K. Yotsushima, T. Mitsui, T. Takaoka, T. Hayakawa and I. Igaue
Department of Biosystem Science, Graduate School of Science and Technology (K.Y.), and Department of Applied Biological Chemistry, Faculty of Agriculture (T.M., T.T., T.H., I.I.), Niigata University, 2-Ikarashi, Niigata 950-21, Japan
A membrane-bound inositol phospholipid-specific phospholipase C was
solubilized from rice (Oryza sativa L.) microsomal membranes and purified
to apparent homogeneity using a series of chromatographic separations. The
apparent molecular mass of the enzyme was estimated by sodium dodecyl
sulfate-polyacrylamide gel electrophoresis to be 42,000 D, and the
isoelectric point was 5.1. The optimum pH for the enzyme activity was
approximately 6.5, and the enzyme was activated by both Ca2+ and Sr2+. The
chemical and catalytic properties of the purified membrane-bound
phospholipase C differed from those of the soluble enzyme reported
previously (K. Yotsushima, K. Nakamura, T. Mitsui, I. Igaue [1992] Biosci
Biotech Biochem 56: 1247-1251). In addition, we found a regulatory factor
for the phosphatidylinositol-4,5-bisphosphate (PIP2) hydrolyzing activity
of phospholipase C from rice cells. The regulatory factor was dissociated
from the catalytic subunit of phospholipase C during the purification. The
regulatory factor was necessary to induce PIP2-hydrolyzing activity of both
membrane-bound and -soluble phospholipase C; these purified enzymes had no
activity alone. Because the plasma membranes isolated from rice cells could
also act as a regulatory factor, the regulatory factor seems to be
localized in the plasma membranes. Regulation of inositol phospholipid
turnover in rice cells is discussed.
