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ErbB3 (HER3) interaction with the p85 regulatory subunit of phosphoinositide 3-kinase.

ABSTRACT: ErbB3 (HER3), a unique member of the ErbB receptor family, lacks intrinsic protein tyrosine kinase activity and contains six Tyr-Xaa-Xaa-Met (YXXM) consensus binding sites for the SH2 domains of the p85 regulatory subunit of phosphoinositide 3-kinase. ErbB3 also has a proline-rich sequence that forms a consensus binding site for the SH3 domain of p85. Here we have investigated the interacting domains of ErbB3 and p85 by a unique application of the yeast two-hybrid system. A chimaeric ErbB3 molecule containing the epidermal growth factor receptor protein tyrosine kinase domain was developed so that the C-terminal domain of ErbB3 could become phosphorylated in the yeast system. We also generated several ErbB3 deletion and Tyr-->Phe site-specific mutants, and observed that a single ErbB3 YXXM motif was necessary and sufficient for the association of ErbB3 with p85. The incorporation of multiple YXXM motifs into the ErbB3 C-terminus enabled a stronger ErbB3/p85 interaction. The proline-rich region of ErbB3 was not necessary for interaction with p85. However, either deletion or mutation of the p85 SH3 domain decreased the observed ErbB3/p85 association. Additionally an ErbB3/p85 SH3 domain interaction was detected by an assay in vitro. These results were consistent with a model in which pairs of phosphorylated ErbB3 YXXM motifs co-operate in binding to the tandem SH2 domains of p85. Although a contributing role for the p85 SH3 domain was suggested, the N- and C-terminal SH2 domains seemed to be primarily responsible for the high-affinity association of p85 and ErbB3.

SUBMITTER: Hellyer NJ

PROVIDER: S-EPMC1219642 | biostudies-other | 1998 Aug

REPOSITORIES: biostudies-other

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p85? phosphoinositide 3-kinase subunit regulates tumor progression.

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Project description:Src-homology 3 (SH3) domains are involved in extensive protein-protein interactions and constitute key elements of intracellular signal transduction. Three-dimensional structures have been reported for SH3 domains of various proteins, including the 85 kDa regulatory subunit (p85) of phosphoinositide 3-kinase. However, all of the latter structures are of p85 isoform ? and no crystal structure of the SH3 domain of the equally important isoform ? has been reported to date. In this structural communication, the recombinant production, crystallization and X-ray structure determination at 2.0 Å resolution of the SH3 domain of human p85? is described. The structure reveals a compact ?-barrel fold very similar to that of p85?. However, binding studies with two classes of proline-rich ligand peptides demonstrate that the ligand-binding specificity differs slightly between the SH3 domains of human p85? and p85?, despite their high structural similarity.

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Interaction of Calmodulin with the cSH2 Domain of the p85 Regulatory Subunit.

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Assembly and Molecular Architecture of the Phosphoinositide 3-Kinase p85? Homodimer.

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