Solution and solid state conformation of the human EGF receptor transmembrane region.
The epidermal growth factor receptor (EGFR) is a member of the tyrosine kinase family of signalling cell surface molecules. Signalling by this protein is mediated through binding of epidermal growth factor to its extracellular region ultimately leading to phosphorylation of several residues on the intracellular portion of the receptor. The only means of communication between the intracellular and extracellular domains is via the transmembrane region of the protein. In this work we describe the first structural studies of a 34-residue synthetic peptide (hEGFRp), representative of the human EGFR transmembrane region, using two-dimensional and 2H wideline NMR and CD spectroscopies. In water the peptide demonstrated a lack of regular secondary structure and existed as oligomers. Addition of the lipomimetic solvent, trifluoroethanol (TFE), led to the production of monomeric structured species. Analysis of NMR spectra of the hEGFRp indicated that an alpha-helix was present between residues M626 and R647. This observation was reinforced by solid state 2H NMR studies in lipid bilayers which showed typical 'Pake' spectra indicating axially symmetric motion. The helical region in hEGFRp commences four residues later than predicted via hydrophobicity profiles, and extends to include several charged arginine residues which would lie on the cytosolic side of the membrane. These observations provide the first evidence that the transmembrane alpha-helical region in EGFR may not only traverse the membrane but may continue to the cytosolic region near T654, an important phosphorylation site.
Rigby
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AC
Department of Biochemistry and McLaughlin Macromolecular Structure Facility, The University of Western Ontario, London, Ontario, Canada.
Grant
C W
CW
Shaw
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GS
D000595:Amino Acid Sequence; D002942:Circular Dichroism; D003593:Cytoplasm / Q000737:chemistry; D066246:ErbB Receptors / Q000737:chemistry*; D006801:Humans; D008051:Lipid Bilayers / Q000737:chemistry; D008565:Membrane Proteins / Q000737:chemistry*; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D012995:Solubility; D012996:Solutions / Q000737:chemistry
D008051:Lipid Bilayers; D008565:Membrane Proteins; D012996:Solutions; D066246:ErbB Receptors