An antagonist peptide–EPO receptor complex suggests that receptor dimerization is not sufficient for activation

Dimerization of the erythropoietin (EPO) receptor (EPOR), in the presence of either natural (EPO) or synthetic (EPO-mimetic peptides, EMPs) ligands is the principal extracellular event that leads to receptor activation. The crystal structure of the extracellular domain of EPOR bound to an inactive (antagonist) peptide at 2.7 Å resolution has unexpectedly revealed that dimerization still occurs, but the orientation between receptor molecules is altered relative to active (agonist) peptide complexes. Comparison of the biological properties of agonist and antagonist EMPs with EPO suggests that the extracellular domain orientation is tightly coupled to the cytoplasmic signaling events and, hence, provides valuable new insights into the design of synthetic ligands for EPOR and other cytokine receptors.

[1]  J. Thornton,et al.  PROCHECK: a program to check the stereochemical quality of protein structures , 1993 .

[2]  D. Chang,et al.  Activation of the Erythropoietin (EPO) Receptor by Bivalent Anti-EPO Receptor Antibodies* , 1996, The Journal of Biological Chemistry.

[3]  Alexander McPherson,et al.  The three-dimensional structure of an intact monoclonal antibody for canine lymphoma , 1992, Nature.

[4]  Z. Otwinowski,et al.  Processing of X-ray diffraction data collected in oscillation mode. , 1997, Methods in enzymology.

[5]  B. Matthews.  Solvent content of protein crystals. , 1968, Journal of molecular biology.

[6]  L S Mulcahy,et al.  Identification of a 13 amino acid peptide mimetic of erythropoietin and description of amino acids critical for the mimetic activity of EMP1. , 1998, Biochemistry.

[7]  S. Pestka,et al.  Identification and sequence of an accessory factor required for activation of the human interferon γ receptor , 1994, Cell.

[8]  Scott R. Presnell,et al.  Erythropoietin structure-function relationships. Mutant proteins that test a model of tertiary structure. , 1993, The Journal of biological chemistry.

[9]  S. Shoelson,et al.  Insulin receptor autophosphorylation occurs asymmetrically. , 1993, The Journal of biological chemistry.

[10]  P. Pilch,et al.  The insulin receptor. Structural basis for high affinity ligand binding. , 1987, The Journal of biological chemistry.

[11]  J. Zou,et al.  Improved methods for building protein models in electron density maps and the location of errors in these models. , 1991, Acta crystallographica. Section A, Foundations of crystallography.

[12]  S. Pestka,et al.  Interaction between the Components of the Interferon γ Receptor Complex (*) , 1995, The Journal of Biological Chemistry.

[13]  S A Kaplan,et al.  The insulin receptor. , 1984, The Journal of pediatrics.

[14]  P Bork,et al.  The immunoglobulin fold. Structural classification, sequence patterns and common core. , 1994, Journal of molecular biology.

[15]  L. Jolliffe,et al.  Refolding, purification, and characterization of human erythropoietin binding protein produced in Escherichia coli. , 1996, Protein expression and purification.

[16]  A. Yoshimura,et al.  Ligand-induced activation of chimeric receptors between the erythropoietin receptor and receptor tyrosine kinases. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[17]  H. Lodish,et al.  Activation and inhibition of erythropoietin receptor function: role of receptor dimerization , 1994, Molecular and cellular biology.

[18]  T. Harvey,et al.  NMR structure of human erythropoietin and a comparison with its receptor bound conformation , 1998, Nature Structural Biology.

[19]  R. Gregory,et al.  Signal transduction in the erythropoietin receptor system. , 1999, Experimental cell research.

[20]  Enrico A. Stura,et al.  Functional Mimicry of a Protein Hormone by a Peptide Agonist: The EPO Receptor Complex at 2.8 Å , 1996, Science.

[21]  M. Ultsch,et al.  Human growth hormone and extracellular domain of its receptor: crystal structure of the complex. , 1992, Science.

[22]  M. Goldsmith,et al.  The cytoplasmic domain of the interleukin-2 receptor beta chain contains both unique and functionally redundant signal transduction elements. , 1994, The Journal of biological chemistry.

[23]  E. Goldwasser,et al.  Evidence for an accessory component that increases the affinity of the erythropoietin receptor. , 1993, Experimental hematology.

[24]  G. Barton,et al.  A structural analysis of phosphate and sulphate binding sites in proteins. Estimation of propensities for binding and conservation of phosphate binding sites. , 1994, Journal of molecular biology.

[25]  R. Huber,et al.  Crystallographic structure studies of an IgG molecule and an Fc fragment , 1976, Nature.

[26]  S. Pestka,et al.  Chimeric Erythropoietin-Interferon γ Receptors Reveal Differences in Functional Architecture of Intracellular Domains for Signal Transduction* , 1997, The Journal of Biological Chemistry.

[27]  M. Boyle,et al.  Homodimerization of erythropoietin receptor by a bivalent monoclonal antibody triggers cell proliferation and differentiation of erythroid precursors. , 1997, Blood.

[28]  A T Brünger,et al.  Protein hydration observed by X-ray diffraction. Solvation properties of penicillopepsin and neuraminidase crystal structures. , 1994, Journal of molecular biology.

[29]  E. Scolnick,et al.  Growth of factor-dependent hemopoietic precursor cell lines , 1980, The Journal of experimental medicine.

[30]  Ronald W. Barrett,et al.  Small Peptides as Potent Mimetics of the Protein Hormone Erythropoietin , 1996, Science.

[31]  R. Zivin,et al.  Amino-terminal dimerization of an erythropoietin mimetic peptide results in increased erythropoietic activity. , 1997, Chemistry & biology.

[32]  S. Pestka,et al.  Identification and sequence of an accessory factor required for activation of the human interferon gamma receptor. , 1994, Cell.

[33]  I. Wilson,et al.  Preliminary crystallographic investigations of glycinamide ribonucleotide transformylase. , 1989, The Journal of biological chemistry.

[34]  J. Navaza,et al.  AMoRe: an automated package for molecular replacement , 1994 .

[35]  O. Silvennoinen,et al.  Signaling through the hematopoietic cytokine receptors. , 1995, Annual review of immunology.

[36]  M. Goldsmith,et al.  Shared gamma(c) subunit within the human interleukin-7 receptor complex. A molecular basis for the pathogenesis of X-linked severe combined immunodeficiency. , 1997, The Journal of clinical investigation.

[37]  B. Groner,et al.  Signaling through the interleukin 2 receptor beta chain activates a STAT-5-like DNA-binding activity. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[38]  M. McMahon,et al.  Erythropoietin induces Raf-1 activation and Raf-1 is required for erythropoietin-mediated proliferation. , 1991, The Journal of biological chemistry.

[39]  M. Goldsmith,et al.  Interleukin‐4‐specific signal transduction events are driven by homotypic interactions of the interleukin‐4 receptor alpha subunit. , 1996, The EMBO journal.

[40]  Robert M. Stroud,et al.  Efficiency of signalling through cytokine receptors depends critically on receptor orientation , 1998, Nature.

[41]  M. Goldsmith,et al.  The molecular role of the common gamma c subunit in signal transduction reveals functional asymmetry within multimeric cytokine receptor complexes. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[42]  I. Wilson,et al.  Applications of the streak seeding technique in protein crystallization , 1991 .

[43]  C. Heldin,et al.  Dimerization of cell surface receptors in signal transduction , 1995, Cell.

[44]  R. Barrett,et al.  Increased potency of an erythropoietin peptide mimetic through covalent dimerization , 1997, Nature Biotechnology.