Limits...
Membrane interaction of bound ligands contributes to the negative binding cooperativity of the EGF receptor.

Arkhipov A, Shan Y, Kim ET, Shaw DE - PLoS Comput. Biol. (2014)

Bottom Line: This cooperativity is widely believed to be central to the effects of ligand concentration on EGFR-mediated intracellular signaling.Although the extracellular portion of the human EGFR dimer has been resolved crystallographically, the crystal structures do not reveal the structural origin of this negative cooperativity, which has remained unclear.Here we report the results of molecular dynamics simulations suggesting that asymmetrical interactions of the two binding sites with the membrane may be responsible (perhaps along with other factors) for this negative cooperativity.

View Article: PubMed Central - PubMed

Affiliation: D. E. Shaw Research, New York, New York, United States of America.

ABSTRACT
The epidermal growth factor receptor (EGFR) plays a key role in regulating cell proliferation, migration, and differentiation, and aberrant EGFR signaling is implicated in a variety of cancers. EGFR signaling is triggered by extracellular ligand binding, which promotes EGFR dimerization and activation. Ligand-binding measurements are consistent with a negatively cooperative model in which the ligand-binding affinity at either binding site in an EGFR dimer is weaker when the other site is occupied by a ligand. This cooperativity is widely believed to be central to the effects of ligand concentration on EGFR-mediated intracellular signaling. Although the extracellular portion of the human EGFR dimer has been resolved crystallographically, the crystal structures do not reveal the structural origin of this negative cooperativity, which has remained unclear. Here we report the results of molecular dynamics simulations suggesting that asymmetrical interactions of the two binding sites with the membrane may be responsible (perhaps along with other factors) for this negative cooperativity. In particular, in our simulations the extracellular domains of an EGFR dimer spontaneously lay down on the membrane in an orientation in which favorable membrane contacts were made with one of the bound ligands, but could not be made with the other. Similar interactions were observed when EGFR was glycosylated, as it is in vivo.

Show MeSH

Related in: MedlinePlus

The ectodomain portion of the EGFR dimer lying on the membrane.(A) A schematic description of EGFR ligand binding. The basic scheme is taken from ref. [15], but we have color coded the ligands to distinguish the free, the membrane-facing bound, and the solvent-facing bound ligands according to the simulation findings. “L” and “K” are used to denote the association constants of EGFR dimerization and ligand binding, respectively. The negative cooperativity is reflected in K21≫K22. The ligands bound to monomers are assumed to face the membrane as found in previous simulations of EGFR monomers (ref. [26]; see the Discussion). (B–D) Simulation of an EGFR dimer construct consisting of the ectodomains and TM helices. One EGFR subunit is colored in shades of blue, the other in shades of red, and the two bound EGF ligands in yellow. In the initial state (B), the ectodomain dimer is standing upright, perpendicular to the membrane. In the course of the simulation, the ectodomain dimer approaches the membrane (C), permitting the formation of extensive interactions between one of the ligands and the membrane, resulting in the partial-resting orientation. The EGF side chains Pro7 and Leu8 (orange) penetrate deep into the membrane, reaching the lipid tails (inset). Later in the course of the simulation (D), the ectodomain dimer approaches closer to the membrane and lies flat on its surface, in the full-resting orientation. (E) The free energy of each ligand's interaction with its host receptor (EGF1, blue; EGF2, red) in a two-ligand EGFR dimer (left panel) and of its interaction with the membrane bilayer (right panel); calculations used the MM/GBVI method. (F) The N-terminal sequences of EGF in various vertebrate species and from a set of members of the human EGF family. The amino acids shown correspond to residues 1–10 of human EGF. The conserved Cys6 is marked in gray and hydrophobic residues in positions 1–8 are marked in orange. Hydrophobic and aromatic residues beyond position 8 are expected to be buried in the protein interior.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4109842&req=5

pcbi-1003742-g001: The ectodomain portion of the EGFR dimer lying on the membrane.(A) A schematic description of EGFR ligand binding. The basic scheme is taken from ref. [15], but we have color coded the ligands to distinguish the free, the membrane-facing bound, and the solvent-facing bound ligands according to the simulation findings. “L” and “K” are used to denote the association constants of EGFR dimerization and ligand binding, respectively. The negative cooperativity is reflected in K21≫K22. The ligands bound to monomers are assumed to face the membrane as found in previous simulations of EGFR monomers (ref. [26]; see the Discussion). (B–D) Simulation of an EGFR dimer construct consisting of the ectodomains and TM helices. One EGFR subunit is colored in shades of blue, the other in shades of red, and the two bound EGF ligands in yellow. In the initial state (B), the ectodomain dimer is standing upright, perpendicular to the membrane. In the course of the simulation, the ectodomain dimer approaches the membrane (C), permitting the formation of extensive interactions between one of the ligands and the membrane, resulting in the partial-resting orientation. The EGF side chains Pro7 and Leu8 (orange) penetrate deep into the membrane, reaching the lipid tails (inset). Later in the course of the simulation (D), the ectodomain dimer approaches closer to the membrane and lies flat on its surface, in the full-resting orientation. (E) The free energy of each ligand's interaction with its host receptor (EGF1, blue; EGF2, red) in a two-ligand EGFR dimer (left panel) and of its interaction with the membrane bilayer (right panel); calculations used the MM/GBVI method. (F) The N-terminal sequences of EGF in various vertebrate species and from a set of members of the human EGF family. The amino acids shown correspond to residues 1–10 of human EGF. The conserved Cys6 is marked in gray and hydrophobic residues in positions 1–8 are marked in orange. Hydrophobic and aromatic residues beyond position 8 are expected to be buried in the protein interior.

Mentions: More recently, in a study conducted by Pike and colleagues [15], a characterization of EGFR ligand binding based on a simultaneous fitting of binding isotherms from cells with different levels of EGFR expression showed that EGFR ligand binding may be described by a simple model (shown in Fig. 1A). In this model, which is consistent with earlier results [16], negative cooperativity underlies the heterogeneity of EGFR ligand binding [15], [16]: The binding affinity of a ligand at one EGFR binding site is smaller when the other site is occupied).


Membrane interaction of bound ligands contributes to the negative binding cooperativity of the EGF receptor.

Arkhipov A, Shan Y, Kim ET, Shaw DE - PLoS Comput. Biol. (2014)

The ectodomain portion of the EGFR dimer lying on the membrane.(A) A schematic description of EGFR ligand binding. The basic scheme is taken from ref. [15], but we have color coded the ligands to distinguish the free, the membrane-facing bound, and the solvent-facing bound ligands according to the simulation findings. “L” and “K” are used to denote the association constants of EGFR dimerization and ligand binding, respectively. The negative cooperativity is reflected in K21≫K22. The ligands bound to monomers are assumed to face the membrane as found in previous simulations of EGFR monomers (ref. [26]; see the Discussion). (B–D) Simulation of an EGFR dimer construct consisting of the ectodomains and TM helices. One EGFR subunit is colored in shades of blue, the other in shades of red, and the two bound EGF ligands in yellow. In the initial state (B), the ectodomain dimer is standing upright, perpendicular to the membrane. In the course of the simulation, the ectodomain dimer approaches the membrane (C), permitting the formation of extensive interactions between one of the ligands and the membrane, resulting in the partial-resting orientation. The EGF side chains Pro7 and Leu8 (orange) penetrate deep into the membrane, reaching the lipid tails (inset). Later in the course of the simulation (D), the ectodomain dimer approaches closer to the membrane and lies flat on its surface, in the full-resting orientation. (E) The free energy of each ligand's interaction with its host receptor (EGF1, blue; EGF2, red) in a two-ligand EGFR dimer (left panel) and of its interaction with the membrane bilayer (right panel); calculations used the MM/GBVI method. (F) The N-terminal sequences of EGF in various vertebrate species and from a set of members of the human EGF family. The amino acids shown correspond to residues 1–10 of human EGF. The conserved Cys6 is marked in gray and hydrophobic residues in positions 1–8 are marked in orange. Hydrophobic and aromatic residues beyond position 8 are expected to be buried in the protein interior.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4109842&req=5

pcbi-1003742-g001: The ectodomain portion of the EGFR dimer lying on the membrane.(A) A schematic description of EGFR ligand binding. The basic scheme is taken from ref. [15], but we have color coded the ligands to distinguish the free, the membrane-facing bound, and the solvent-facing bound ligands according to the simulation findings. “L” and “K” are used to denote the association constants of EGFR dimerization and ligand binding, respectively. The negative cooperativity is reflected in K21≫K22. The ligands bound to monomers are assumed to face the membrane as found in previous simulations of EGFR monomers (ref. [26]; see the Discussion). (B–D) Simulation of an EGFR dimer construct consisting of the ectodomains and TM helices. One EGFR subunit is colored in shades of blue, the other in shades of red, and the two bound EGF ligands in yellow. In the initial state (B), the ectodomain dimer is standing upright, perpendicular to the membrane. In the course of the simulation, the ectodomain dimer approaches the membrane (C), permitting the formation of extensive interactions between one of the ligands and the membrane, resulting in the partial-resting orientation. The EGF side chains Pro7 and Leu8 (orange) penetrate deep into the membrane, reaching the lipid tails (inset). Later in the course of the simulation (D), the ectodomain dimer approaches closer to the membrane and lies flat on its surface, in the full-resting orientation. (E) The free energy of each ligand's interaction with its host receptor (EGF1, blue; EGF2, red) in a two-ligand EGFR dimer (left panel) and of its interaction with the membrane bilayer (right panel); calculations used the MM/GBVI method. (F) The N-terminal sequences of EGF in various vertebrate species and from a set of members of the human EGF family. The amino acids shown correspond to residues 1–10 of human EGF. The conserved Cys6 is marked in gray and hydrophobic residues in positions 1–8 are marked in orange. Hydrophobic and aromatic residues beyond position 8 are expected to be buried in the protein interior.
Mentions: More recently, in a study conducted by Pike and colleagues [15], a characterization of EGFR ligand binding based on a simultaneous fitting of binding isotherms from cells with different levels of EGFR expression showed that EGFR ligand binding may be described by a simple model (shown in Fig. 1A). In this model, which is consistent with earlier results [16], negative cooperativity underlies the heterogeneity of EGFR ligand binding [15], [16]: The binding affinity of a ligand at one EGFR binding site is smaller when the other site is occupied).

Bottom Line: This cooperativity is widely believed to be central to the effects of ligand concentration on EGFR-mediated intracellular signaling.Although the extracellular portion of the human EGFR dimer has been resolved crystallographically, the crystal structures do not reveal the structural origin of this negative cooperativity, which has remained unclear.Here we report the results of molecular dynamics simulations suggesting that asymmetrical interactions of the two binding sites with the membrane may be responsible (perhaps along with other factors) for this negative cooperativity.

View Article: PubMed Central - PubMed

Affiliation: D. E. Shaw Research, New York, New York, United States of America.

ABSTRACT
The epidermal growth factor receptor (EGFR) plays a key role in regulating cell proliferation, migration, and differentiation, and aberrant EGFR signaling is implicated in a variety of cancers. EGFR signaling is triggered by extracellular ligand binding, which promotes EGFR dimerization and activation. Ligand-binding measurements are consistent with a negatively cooperative model in which the ligand-binding affinity at either binding site in an EGFR dimer is weaker when the other site is occupied by a ligand. This cooperativity is widely believed to be central to the effects of ligand concentration on EGFR-mediated intracellular signaling. Although the extracellular portion of the human EGFR dimer has been resolved crystallographically, the crystal structures do not reveal the structural origin of this negative cooperativity, which has remained unclear. Here we report the results of molecular dynamics simulations suggesting that asymmetrical interactions of the two binding sites with the membrane may be responsible (perhaps along with other factors) for this negative cooperativity. In particular, in our simulations the extracellular domains of an EGFR dimer spontaneously lay down on the membrane in an orientation in which favorable membrane contacts were made with one of the bound ligands, but could not be made with the other. Similar interactions were observed when EGFR was glycosylated, as it is in vivo.

Show MeSH
Related in: MedlinePlus