Spatial Structure of the Dimeric Transmembrane Domain of the Growth Factor Receptor ErbB2 Presumably Corresponding to the Receptor Active State

Abstract

Proper lateral dimerization of the transmembrane domains of receptor tyrosine kinases is required for biochemical signal transduction across the plasma membrane. The spatial structure of the dimeric transmembrane domain of the growth factor receptor ErbB2 embedded into lipid bicelles was obtained by solution NMR, followed by molecular dynamics relaxation in an explicit lipid bilayer. ErbB2 transmembrane segments associate in a right-handed ␣-helical bundle through the N-terminal tandem GG4-like motif Thr 652 -X 3 -Ser 656 -X 3 -Gly 660 , providing an explanation for the pathogenic power of some oncogenic mutations.The epidermal growth factor receptor (or ErbB) family is an important class of receptor tyrosine kinases involved in transmission of biochemical signals governing cell fate (1). Four human ErbB family members form numerous homo-and heterodimer combinations and bind different epidermal growth factor-related ligands, thus performing diverse functions in a complex signaling network (2). The binding of peptide growth factors to the extracellular domain of the receptor triggers the dimerization of receptor monomers or a change in the relative orientation of monomers in preformed receptor dimers, leading to autophosphorylation of tyrosine residues in the cytoplasmic kinase domain (3, 4). Biochemical and genetic studies have revealed that the single-helix transmembrane (TM) 3 domains of ErbB play an active role in the dimerization process and associate strongly in the absence of extracellular ligand-binding and cytoplasmic kinase domains (5, 6). Mutational analysis assumed that the dimerization involves consensus small-X 3 -small (so-called GG4-like) motifs, formed by residues with small side chains allowing tight helix packing (7-9). Receptor tyrosine kinase TM sequences often contain several remote GG4-like motifs, suggesting the ability of their TM domains to adopt more than one conformation, e.g. upon so-called rotation-coupled activation of the receptor (4, 10, 11). Recent molecular modeling and solid-state NMR studies performed to predict the spatial structures of the dimeric TM domains of the human ErbB2 receptor and its rat homolog have disclosed two possible dimer conformations with interfaces located at either the N or C terminus of the ␣-helical TM segment, employing different GG4-like motifs for dimerization (7,(11)(12)(13). Nevertheless, an experimental spatial structure of the dimeric TM domain for ErbB2 as well as for any other receptor tyrosine kinase family members has not been reported so far.Here, we present the high resolution structure of the homodimeric ErbB2 TM domain in a membrane-mimicking lipid environment solved by a heteronuclear NMR technique combined with molecular dynamics (MD) relaxation in an explicit membrane. Our results distinguish one of the potential conformations of the homodimer, which can be ascribed to the active state of the tyrosine kinase. On the basis of the analysis of the local conformation of the dimerization interface, we propose a molecular mechanism of actio...

Receptor tyrosine kinase transmembrane domains

“…41 In addition, computational and experimental studies of the structure of the isolated Neu/ErbB2 and FGFR3 TM domain dimers demonstrate dimer stabilization via Glu-mediated hydrogen bonding. 9,12,14,[41][42][43] Taken together, these results strongly suggest that hydrogen bonds stabilize the mutant Neu/Val664Glu and Neu_FGFR3/Ala391Glu receptors in mammalian cells, a mechanism that was originally proposed 18 years ago, 42 but has been debated ever since. Furthermore, the results suggest that an increase of the order of -1 kcal/mole may be sufficient to transform normal RTK signaling processes into pathogenic processes.…”

“…12,19 We anticipate more such homodimer structures and we hope to see the first heterodimer structure in the near future. The role of heterodimerization within RTK subfamilies is now well established, and yet biophysical studies of heterodimers in membranes are rare.…”

“…12 The structure was solved in bicelles composed of dimyristoylphosphatidylcholine (DMPC) and dihexanoylphosphatidylcholine (DHPC). To address the effect of the bicelle environment, the authors performed circular dichroism measurements in the bicelles and in lipid vesicles and demonstrated that the secondary structure is the same.…”

A frequent, GxxxG-mediated, transmembrane association motif is optimized for the formation of interhelical Cα–H hydrogen bonds

“…The TM region of another member of the same family, ErbB2, has also been solved by NMR in dimeric form (31). The NMR model has a crossing angle of −41°and an interhelical distance of 7.6 Å; however, this structure is not mediated by Cα-H hydrogen bonds.…”

Conformational flexibility of the ErbB2 ectodomain and trastuzumab antibody complex as revealed by molecular dynamics and principal component analysis

“…Several computational studies based on computer simulations have tackled the structure and interactions between transmembrane ErbB2 domains in lipidic bilayer models [19][20][21][22][23][24] and in tyrosine kinase domain activation [25][26][27]. However, computational studies of the interaction of ErbB2 ectodomain (ErbB2 ECD) with antibodies, such as trastuzumab, are less considered.…”