The extracellular region of the receptor for advanced glycation end products is composed of two independent structural units.
The receptor for advanced glycation end products (RAGE) is an important cell surface receptor being pursued as a therapeutic target because it has been implicated in complications arising from diabetes and chronic inflammatory conditions. RAGE is a single membrane spanning receptor containing a very small approximately 40 residue cytosolic domain and a large extracellular region composed of 3 Ig-like domains. In this study, high level bacterial expression systems and purification protocols were generated for the extracellular region of RAGE (sRAGE) and the five permutations of single and tandem domain constructs to enable biophysical and structural characterization of its tertiary and quaternary structure. The structure and stability of each of these six protein constructs was assayed by biochemical methods including limited proteolysis, dynamic light scattering, CD, and NMR. A homology model of sRAGE was constructed to aid in the interpretation of the experimental data. Our results show that the V and C1 domains are not independent domains, but rather form an integrated structural unit. In contrast, C2 is attached to VC1 by a flexible linker and is fully independent. The interaction with a known RAGE ligand, Ca2+-S100B, was mapped to VC1, with the major contribution from the V domain but clearly defined secondary effects from the C1 domain. The implications of these results are discussed with respect to models for RAGE signaling.
Dattilo
Brian M
BM
Department of Biochemistry, Center for Structural Biology, 465 21st Avenue South, 5140 BIOSCI/MRBIII, Vanderbilt University, Nashville, Tennessee 37232-8725, USA.
Fritz
Günter
G
Leclerc
Estelle
E
Kooi
Craig W Vander
CW
Heizmann
Claus W
CW
Chazin
Walter J
WJ
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA / Q000737:chemistry / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D017127:Glycation End Products, Advanced / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D011487:Protein Conformation; D000067759:Receptor for Advanced Glycation End Products; D011971:Receptors, Immunologic / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
D017127:Glycation End Products, Advanced; D010446:Peptide Fragments; D000067759:Receptor for Advanced Glycation End Products; D011971:Receptors, Immunologic; D011994:Recombinant Proteins; D004247:DNA