Characterization of a protein which binds phosphatidylinositol 3,4,5-trisphosphate and 4,5-bisphosphate.
Protein(s) which bind polyphosphatidylinositol phosphates (PI 3,4,5-P3 and PI 4,5-P2) were identified in the wheat-germ agglutinin bound fraction of cells and tissues. The binding of this protein(s) to the phospholipid could be demonstrated in two ways, either by a shift in the migration of the lipid by size exclusion column chromatography or directly by binding to the protein after capture on wheat-germ agglutinin-coupled beads. Of the rat tissues tested (muscle, spleen, brain, heart, kidney and liver), the activity was highest in liver. The protein(s) was purified more than 5000-fold by sequential chromatography on columns of wheat-germ agglutinin, phosphocellulose, Blue-Sepharose, Mono Q and Superose 6. The peak of activity appeared to have a molecular weight on this latter column of approx. 240,000. The protein(s) bound PI 3,4,5-P3, PI 3,4-P2, and PI 3-P in the ratio of 4:2:1. The binding of 3-phosphorylated PI phosphates to the protein(s) was not significantly inhibited by 36 micrograms/ml of either phosphatidylinositol or phosphatidylcholine, but was inhibited 10% and 65% by 36 micrograms/ml of PI 4-P and PI 4,5-P2, respectively. Since these results suggested that the binding protein(s) could also bind PI 4,5-P2, binding of this lipid was directly tested and found to be comparable to that of PI 3,4,5-P3. These results suggest that this protein(s) could be involved in the signaling mechanism elicited by these polyphosphoinositides.
Ogawa
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Department of Molecular Pharmacology, Stanford University School of Medicine, CA 94305.
Roth
R A
RA
D000818:Animals; D001921:Brain / Q000378:metabolism; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D007668:Kidney / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D009132:Muscles / Q000378:metabolism; D009206:Myocardium / Q000378:metabolism; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism*; D011485:Protein Binding; D011506:Proteins / Q000302:isolation & purification / Q000378:metabolism*; D051381:Rats; D013154:Spleen / Q000378:metabolism
D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates; D011506:Proteins; C060974:phosphatidylinositol 3,4,5-triphosphate