title: Reconstitution of Functionalized Transmembrane Domains of Receptor Proteins into Biomimetic Membranes.
abstract: For integral membrane proteins, an assessment of their structures and interactions within a biomimetic lipid bilayer environment is critical for evaluating their cellular function. Hydrophobic sequences prevalent within transmembrane domains, however, make these proteins susceptible to aggregation and, thus, create difficulties in examining their structural and functional properties via canonical techniques. Working exclusively with single-pass transmembrane (TM) segments of bitopic membrane proteins, in the form of soluble peptides, bypasses many of the pitfalls of full-length protein preparations while allowing for the opportunity to examine the properties of TM domains within biomimetic membrane environments. In this study, peptides mimicking the TM domains of the epidermal growth factor receptor (EGFR) and CD4 co-receptor, both cell-signaling surface receptors, have been reconstituted into 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) lipid bilayers. The formation of their native α-helical structures within vesicle membranes was observed from circular dichroism, and full partition of the peptides into the membrane was demonstrated by tryptophan fluorescence and neutron reflectivity (NR). Using an engineered planar lipid bilayer system ideal for surface characterization methods, such as surface plasmon resonance (SPR) and NR, the TM peptides, functionalized with a N-terminal biotin tag, proved capable of "activating" a membrane surface, as evidenced by the capture of streptavidin. On the basis of these initial assessments, we anticipate these membrane-bound peptides will provide a versatile platform for understanding the intricate roles of receptor TM domains in cell signaling.
authors.affiliation: Center for Neutron Research, National Institute of Standards and Technology (NIST) , Gaithersburg, Maryland 20899, United States.|Institute of Bioscience and Biotechnology Research, University of Maryland , Rockville, Maryland 20850, United States.
authors.forename: Daniel R
authors.initials: DR
authors.lastname: Scott
authors.affiliation: Institute of Bioscience and Biotechnology Research, University of Maryland , Rockville, Maryland 20850, United States.
authors.forename: Vitalii
authors.initials: V
authors.lastname: Silin
authors.affiliation: Center for Neutron Research, National Institute of Standards and Technology (NIST) , Gaithersburg, Maryland 20899, United States.
authors.forename: Hirsh
authors.initials: H
authors.lastname: Nanda
mesh_terms: D015704:CD4 Antigens / Q000737:chemistry*; D066246:ErbB Receptors / Q000737:chemistry*; D006801:Humans; D008051:Lipid Bilayers / Q000737:chemistry*; D010713:Phosphatidylcholines / Q000737:chemistry*; D017434:Protein Structure, Tertiary
chemical_list: D015704:CD4 Antigens; D008051:Lipid Bilayers; D010713:Phosphatidylcholines; C512478:EGFR protein, human; D066246:ErbB Receptors; C028694:1-palmitoyl-2-oleoylphosphatidylcholine