Dimeric Structure of the Transmembrane Domain of Glycophorin A in Lipidic and Detergent Environments

Abstract

Specific interactions between transmembrane α-helices, to a large extent, determine the biological function of integral membrane proteins upon normal development and in pathological states of an organism. Various membrane-like media, partially those mimicking the conditions of multicomponent biological membranes, are used to study the structural and thermodynamic features that define the character of oligomerization of transmembrane helical segments. The choice of the composition of the membrane-mimicking medium is conducted in an effort to obtain a biologically relevant conformation of the protein complex and a sample that would be stable enough to allow to perform a series of long-term experiments with its use. In the present work, heteronuclear NMR spectroscopy and molecular dynamics simulations were used to demonstrate that the two most widely used media (detergent DPC micelles and lipid DMPC/DHPC bicelles) enable to perform structural studies of the specific interactions between transmembrane α-helices by the example of dimerizing the transmembrane domain of the bitopic protein glycophorin A. However, a number of peculiarities place lipid bicelles closer to natural lipid bilayers in terms of their physical properties.

Molecular dynamics simulations of biological membranes and membrane proteins using enhanced conformational sampling algorithms

“…Its transmembrane (TM) dimer structure was determined by solution nuclear magnetic resonance (NMR) spectroscopy in detergent micelles (PDB ID: 1AFO) [194] and in lipid bicelles (PDB ID: 2KPE, 2KPF) [195]. GpA has long been a model protein to investigate structure and stability of membrane protein dimers in different membrane environments [196,197] as well as a popular target of membrane simulations [72,74,75,119,123,187,198].…”

Understanding single‐pass transmembrane receptor signaling from a structural viewpoint—what are we missing?

“…). Nine of these are RTKs (ErbB1‐4 , FGFR3 , PDGFR , VEGFR2 , EphA1‐2 ), three are immunoreceptors (lymphoid/myeloid receptor signaling module (DAP12) , the ζ chain (ζζ) , Toll‐like receptor 3 (TLR3) ), one belongs to the APLPs (APP ), and one belongs to the GP family (GpA , ). Two similar homodimer structures exist of the GpA‐TMD and the APP‐TMD (http://www.rcsb.org/pdb/search/structidSearch.do?structureId=1AFO and http://www.rcsb.org/pdb/search/structidSearch.do?structureId=2KPF/http://www.rcsb.org/pdb/search/structidSearch.do?structureId=2KPE for the GpA and http://www.rcsb.org/pdb/search/structidSearch.do?structureId=2LOH and http://www.rcsb.org/pdb/search/structidSearch.do?structureId=2LZ3 for the APP), which in each case have been solved by different groups, while two different homodimer structures have been solved of the ErbB1‐TMD (http://www.rcsb.org/pdb/search/structidSearch.do?structureId=2M20 and http://www.rcsb.org/pdb/search/structidSearch.do?structureId=2M0B ) and the ErbB2‐TMD (http://www.rcsb.org/pdb/search/structidSearch.do?structureId=2JWA and http://www.rcsb.org/pdb/search/structidSearch.do?structureId=2N2A ).…”

“…The GpA‐TMD homodimers, solved in DPC micelles and DMPC bicelles by two different groups (http://www.rcsb.org/pdb/search/structidSearch.do?structureId=1AFO in DPC micelles , and http://www.rcsb.org/pdb/search/structidSearch.do?structureId=2KPE and http://www.rcsb.org/pdb/search/structidSearch.do?structureId=2KPF in DPC micelles and DMPC bicelles, respectively ) have very similar backbone structures. Only a small deviation in the last helical turn is immediately apparent, giving a variation in the helical C‐terminal distances of 2.5 Å between the structures solved in DPC.…”

“…As discussed, the overall quality of the available structures appears limited, and, in addition, the oligomeric state of the protein is not always clearly defined. The homodimeric state of most of the TMDs has mainly been confirmed under the applied conditions by acquisition of interhelical NOEs , sometimes in combination with cross‐linking experiments or with the detection of size changes as estimated from backbone relaxation rates or ultracentrifugation . Interhelical NOEs obtained in filter‐based experiments should preferably be compared to background reference spectra of the protein in the absence of unlabeled protein to avoid misinterpretation of strong intramolecular peaks leaking through the filter due to imperfect isotope labeling , but it is rarely clear if these have been acquired.…”

“…Nine of these are RTKs (ErbB1-4 [15,[58][59][60][61][62], FGFR3 [63], PDGFR [64], VEGFR2 [23], EphA1-2 [65,66]), three are immunoreceptors (lymphoid/myeloid receptor signaling module (DAP12) [67], the f chain (ff) [68], Toll-like receptor 3 (TLR3) [69]), one belongs to the APLPs (APP [70,71]), and one belongs to the GP family (GpA [33], [72]). Two similar homodimer structures exist of the GpA-TMD and the APP-TMD (1AFO [33] and 2KPF/2KPE [72] for the GpA and 2LOH [71] and 2LZ3 [70] for the APP), which in each case have been solved by different groups, while two different homodimer structures have been solved of the ErbB1-TMD (2M20 [15] and 2M0B [62]) and the ErbB2-TMD (2JWA [58] and 2N2A [61]). The TMD homodimer structures all consist of two main a-helices harboring 23-38 residues each, and include the regions bioinformatically predicted to be the TMD [57].…”