Identification of a functional epitope of the Nogo receptor by a combinatorial approach using ribosome display.
The Nogo receptor (NgR) plays a central role in mediating growth-inhibitory activities of myelin-derived proteins, thereby severely limiting axonal regeneration after injury of the adult mammalian central nervous system (CNS). The inhibitory proteins Nogo, myelin-associated glycoprotein (MAG) and oligodendrocyte myelin glycoprotein (OMgp) all bind to the extracellular leucine-rich repeat (LRR) domain of NgR, which provides a large molecular surface for protein-protein interactions. However, epitopes within the LRR domain of NgR for binding Nogo, MAG and OMgp have not yet been revealed. Here, we report an evolutionary approach based on the ribosome display technology for detecting regions involved in ligand binding. By applying this method of "affinity fingerprinting" to the NgR ligand binding domain we were able to detect a distinct region important for binding to Nogo. Several residues defining the structural epitope of NgR involved in interaction with Nogo were subsequently confirmed by alanine scanning mutagenesis.
Schimmele
Bernhard
B
Biochemisches Institut der Universität Zürich, Winterthurer Strasse 190, CH-8057 Zürich, Switzerland.
Plückthun
Andreas
A
D000595:Amino Acid Sequence; D000915:Antibody Affinity; D001665:Binding Sites; D020650:Combinatorial Chemistry Techniques; D000939:Epitopes; D058851:GPI-Linked Proteins; D006801:Humans; D008958:Models, Molecular; D009185:Myelin Proteins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D000070798:Nogo Proteins; D000070816:Nogo Receptor 1; D017434:Protein Structure, Tertiary; D011863:Radioimmunoassay; D011956:Receptors, Cell Surface / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D012270:Ribosomes / Q000378:metabolism*; D017422:Sequence Analysis, DNA
D000939:Epitopes; D058851:GPI-Linked Proteins; D009185:Myelin Proteins; D000070798:Nogo Proteins; D000070816:Nogo Receptor 1; C000603199:RTN4 protein, human; C486580:RTN4R protein, human; D011956:Receptors, Cell Surface