ABSTRACT
The insulin receptor is a phylogenetically ancient tyrosine kinase receptor found in organisms as primitive as cnidarians and insects. In higher organisms it is essential for glucose homeostasis, whereas the closely related insulin-like growth factor receptor (IGF-1R) is involved in normal growth and development. The insulin receptor is expressed in two isoforms, IR-A and IR-B; the former also functions as a high-affinity receptor for IGF-II and is implicated, along with IGF-1R, in malignant transformation. Here we present the crystal structure at 3.8 A resolution of the IR-A ectodomain dimer, complexed with four Fabs from the monoclonal antibodies 83-7 and 83-14 (ref. 4), grown in the presence of a fragment of an insulin mimetic peptide. The structure reveals the domain arrangement in the disulphide-linked ectodomain dimer, showing that the insulin receptor adopts a folded-over conformation that places the ligand-binding regions in juxtaposition. This arrangement is very different from previous models. It shows that the two L1 domains are on opposite sides of the dimer, too far apart to allow insulin to bind both L1 domains simultaneously as previously proposed. Instead, the structure implicates the carboxy-terminal surface of the first fibronectin type III domain as the second binding site involved in high-affinity binding.
Subject(s)
Protein Folding , Receptor, Insulin/chemistry , Receptor, Insulin/metabolism , Crystallography, X-Ray , Dimerization , Immunoglobulin Fab Fragments/immunology , Microscopy, Electron , Models, Molecular , Protein Structure, Quaternary , Protein Structure, Tertiary , Receptor, Insulin/immunology , Receptor, Insulin/ultrastructureABSTRACT
ErbB2 does not bind ligand, yet appears to be the major signaling partner for other ErbB receptors by forming heteromeric complexes with ErbB1, ErbB3, or ErbB4. The crystal structure of residues 1-509 of ErbB2 at 2.5 A resolution reveals an activated conformation similar to that of the EGFR when complexed with ligand and very different from that seen in the unactivated forms of ErbB3 or EGFR. The structure explains the inability of ErbB2 to bind known ligands and suggests why ErbB2 fails to form homodimers. Together, the data suggest a model in which ErbB2 is already in the activated conformation and ready to interact with other ligand-activated ErbB receptors.
Subject(s)
Receptor, ErbB-2/chemistry , Receptor, ErbB-2/metabolism , Amino Acid Sequence , Animals , Base Sequence , Binding Sites , CHO Cells , Cricetinae , Crystallography, X-Ray , DNA, Complementary/genetics , ErbB Receptors/chemistry , ErbB Receptors/genetics , ErbB Receptors/metabolism , Humans , In Vitro Techniques , Ligands , Models, Molecular , Molecular Sequence Data , Peptide Fragments/chemistry , Peptide Fragments/genetics , Peptide Fragments/metabolism , Protein Conformation , Protein Structure, Tertiary , Receptor, ErbB-2/genetics , Receptor, ErbB-3/chemistry , Receptor, ErbB-3/genetics , Receptor, ErbB-3/metabolism , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Sequence Homology, Amino Acid , Static ElectricityABSTRACT
We report the crystal structure, at 2.5 A resolution, of a truncated human EGFR ectodomain bound to TGFalpha. TGFalpha interacts with both L1 and L2 domains of EGFR, making many main chain contacts with L1 and interacting with L2 via key conserved residues. The results indicate how EGFR family members can bind a family of highly variable ligands. In the 2:2 TGFalpha:sEGFR501 complex, each ligand interacts with only one receptor molecule. There are two types of dimers in the asymmetric unit: a head-to-head dimer involving contacts between the L1 and L2 domains and a back-to-back dimer dominated by interactions between the CR1 domains of each receptor. Based on sequence conservation, buried surface area, and mutagenesis experiments, the back-to-back dimer is favored to be biologically relevant.
