Coiled-coil molecular recognition: directed solution assembly of receptor ectodomains.
The high affinity interleukin-2 (IL-2) receptor is composed of at least three cell surface proteins (alpha, beta and gamma subunits), each of which is independently capable of ligand binding. Physiologically, these subunits cooperate to form dimeric and trimeric complexes that efficiently capture IL-2 and transmit the signal across the membrane. The knowledge of how each subunit functions with respect to ligand capture, signal transmission and internalization is essential for the development of ligand-based IL-2 agonists and antagonists, as well as receptor-related therapeutic and diagnostic reagents. Only one of the subunits (p55 or alpha chain) is capable of interacting with ligand in solution in a manner that resembles cell surface binding. To generate soluble multimeric complexes of the IL-2 receptor subunits that may bind ligand in solution in a fashion that mimics the same receptor complexes on the cell surface, we have added recognition sequences (coiled-coil heptad repeats) to the ectodomains of the individual receptors. Here we describe the expression and characterization of a prototype IL-2 beta receptor ectodomain-coiled-coil fusion protein and demonstrate that this is a feasible approach to the preparation of cytokine receptor solution complexes.
Wu
Z
Z
Department of Pharmacology and Toxicology, Dartmouth Medical School, Hanover, NH 03755.
Eaton
S F
SF
Laue
T M
TM
Johnson
K W
KW
Sana
T R
TR
Ciardelli
T L
TL
D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D001665:Binding Sites; D001667:Binding, Competitive; D003001:Cloning, Molecular; D006801:Humans; D007376:Interleukin-2 / Q000378:metabolism; D008024:Ligands; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D011487:Protein Conformation; D015202:Protein Engineering; D015375:Receptors, Interleukin-2 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D018411:Spodoptera
D007376:Interleukin-2; D008024:Ligands; D046911:Macromolecular Substances; D015375:Receptors, Interleukin-2; D011993:Recombinant Fusion Proteins