Radiolabeling of catalytic subunits of PI 3-kinases with 17 beta-hydroxy-16 alpha(-)[125I]iodowortmannin: identification of the G beta gamma-sensitive isoform as a complex composed of 46-kDa and 100-kDa subunits.
A fungal metabolite, wortmannin, is a potent inhibitor of phosphatidylinositol (PI) 3-kinases. In the present study, we prepared a radiolabeled derivative of wortmannin, 17 beta-hydroxy-16 alpha(-)[125I]iodowortmannin. The compound bound tightly to a 110-kDa subunit in the previously identified isoform of PI 3-kinase (p85/p110), and also to a 100-kDa peptide in a partially purified preparation of another isoform of PI 3-kinase whose activity was markedly stimulated by the beta gamma subunits of GTP-binding proteins (G beta gamma). The binding to both peptides was inhibited by non-radiolabeled wortmannin and also by LY294002, another inhibitor of PI 3-kinases. An antibody against p85 recognized a 46-kDa peptide in the G beta gamma-sensitive isozyme and precipitated the 100-kDa peptide specifically labeled with 17 beta-hydroxy-16 alpha(-)[125I]iodowortmannin. These results suggested that the newly found isozyme was a complex composed of 46-kDa and 100-kDa peptides.
D000730:Androstadienes / Q000138:chemical synthesis; D000818:Animals; D001345:Autoradiography; D002852:Chromatography, Ion Exchange; D003600:Cytosol / Q000201:enzymology; D019204:GTP-Binding Proteins / Q000378:metabolism*; D015151:Immunoblotting; D007457:Iodine Radioisotopes; D007527:Isoenzymes / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D046911:Macromolecular Substances; D008297:Male; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D011485:Protein Binding; D051381:Rats; D017208:Rats, Wistar
C096790:17-hydroxy-16-iodowortmannin; D000730:Androstadienes; D007457:Iodine Radioisotopes; D007527:Isoenzymes; D046911:Macromolecular Substances; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D019204:GTP-Binding Proteins