Transmembrane domain of EphA1 receptor forms dimers in membrane-like environment

Abstract

Eph receptor tyrosine kinases (RTKs) are activated by a ligand-mediated dimerization in the plasma membrane and subjected to clusterization at a high local density of receptors and their membrane-anchored ligands. Interactions between transmembrane domains (TMDs) were recognized to assist to the ligand-binding extracellular domains in the dimerization of some RTKs, whereas a functional role of Eph-receptor TMDs remains unknown. We have studied a propensity of EphA1-receptor TMDs (TMA1) to self-association in membrane-mimetic environment. Dimerization of TMA1 in SDS environment was revealed by SDS-PAGE and confirmed by FRET analysis of the fluorescently labeled peptide (Kd=7.2+/-0.4 microM at 1.5 mM SDS). TMA1 dimerization was also found in 1,2-dimyristoyl-sn-glycero-3-phosphocholine liposomes (DeltaG=-15.4+/-0.5 kJ/mol). Stability of TMA1 dimers is comparable to the reported earlier stability of TMD dimers of fibroblast growth factor receptor 3 and tenfold weaker than the stability of TMD dimers of glycophorin A possessing high propensity to dimerization. Our results suggest that EphA1-receptor TMD contribute to the dimerization-mediated receptor activation. An assumed role of the TMD interactions is the efficient signal transduction due to TMD-driving mutual orientation of kinase domains in dimers, while a relatively low force of the TMD interactions does not prevent a ligand-controlled regulation of the receptor dimerization.

Receptor tyrosine kinase transmembrane domains

“…So far, three different TM domains from three different RTK subfamilies have been characterized in terms of their homodimerization energetics in lipid bilayers, the TM domains of ErbB1, FGFR3 and EphA1. [28][29][30] The strengths of interactions for these three RTK TM domains are very similar, about -3 kcal/mole.…”

Hydrophobic Matching Controls the Tilt and Stability of the Dimeric Platelet-derived Growth Factor Receptor (PDGFR) β Transmembrane Segment

“…Liquid-state NMR Spectroscopy-Triple-resonance experiments for assignment and 13 C NOESY experiments were recorded on 1 mM uniformly 15 N-labeled or 15 N 13 C-labeled PDGFR␤-TM peptide in 200 mM deuterated DPC micelles (Bruker Avance 600 spectrometer equipped with a TBI tripleresonance probe head). Intermonomer NOEs were acquired from a three-dimensional 13 C-filtered 13 C-edited NOESY measured on a 1:1 mixture of uniformly 15 N 13 C-labeled and unlabeled peptide dissolved in D 2 O (Bruker Avance 900 spectrometer).…”

“…Intermonomer NOEs were acquired from a three-dimensional 13 C-filtered 13 C-edited NOESY measured on a 1:1 mixture of uniformly 15 N 13 C-labeled and unlabeled peptide dissolved in D 2 O (Bruker Avance 900 spectrometer). ARIA1.2-CNS was used for NOE calibration and structure calculation (20).…”

“…The structure calculation of the PDGFR␤-TM dimer was performed in two steps. First, the structure of the monomer was solved from 13 C-and 15 N-edited NOE-derived distance restraints and -dihedral angles from TALOS analysis. The peptide forms a continuous slightly curved ␣-helix, extending from Phe-530 to Trp-556 (Fig.…”

“…Recent structural studies on the isolated TM helices of ErbB2, EphA1, EphA2, ErbB3, and the heterodimer ErbB1/ ErbB2 showed that the TM helices, at least in these systems, dimerize in two different ways: either right-handed parallel or left-handed coiled-coil ␣-helical dimers (12)(13)(14)(15)(16)(17). Ambiguity remains about what governs the arrangement in either of the two conformations or whether both conformations are relevant for activation and may in fact represent different activation states for a given RTK dimer.…”