PI 3-KINASE - STRUCTURAL AND FUNCTIONAL-ANALYSIS OF INTERSUBUNIT INTERACTIONS

Phosphatidylinositol (PI) 3-kinase has an 85 kDa subunit (p85alpha) wh ich mediates its association with activated protein tyrosine kinase re ceptors through SH2 domains, and an 110 kDa subunit (p110) which has i ntrinsic catalytic activity. Here p85alpha and a related protein p85be ta are shown to form stable complexes with recombinant p110 in vivo an d in vitro. Using a panel of glutathione S-transferase (GST) fusion pr oteins of the inter-SH2 region of p85, 104 amino acids were found to b ind directly the p110 protein, while deletion mutants within this regi on further defined the binding site to a sequence of 35 amino acids. T ransient expression of the mutant p85alpha protein in mouse L cells sh owed it was unable to bind PI 3-kinase activity in vivo. Mapping of th e complementary site of interaction on the p110 protein defined 88 ami no acids in the N-terminal region of p110 which mediate the binding of this subunit to either the p85alpha or the p85beta proteins. The inte r-SH2 region of p85 is predicted to be an independently folded module of a coiled-coil of two long anti-parallel alpha-helices. The predicte d structure of p85 suggests a basis for the intersubunit interaction a nd the relevance of this interaction with respect to the regulation of the PI 3-kinase complex is discussed.