The 'ligand-induced conformational change' of alpha 5 beta 1 integrin. Relocation of alpha 5 subunit to uncover the beta 1 stalk region.
Integrin heterodimers undergo a conformational change upon the binding of ligand to their extracellular domains. An anti-beta1 integrin monoclonal antibody AG89 can detect such a conformational change since it recognizes a ligand-inducible epitope in the stalk-like region of beta1 subunits. The binding of a 125I-labeled AG89 Fab fragment to alpha5 beta1 integrins on K562 cells was assessed and analyzed by the Scatchard method. High affinity binding sites for AG89 are present on cells treated with ligand peptide. In addition, results revealed that cells treated with EDTA also express AG89 binding sites with the same affinity although the number of binding sites is 4-fold lower. AG89 immunoprecipitated alpha5 beta1 complexes from surface-labeled K562 cells treated with ligand peptide. By contrast, it immunoprecipitated only beta1 chains when the ligand peptide was absent, suggesting that high affinity binding sites on EDTA-treated cells are associated with non-functional beta1 monomer. Additional studies show that the epitope for AG89 is constitutively exposed on mutant beta1 that cannot complex with alpha5. These data suggest that the AG89 epitope is masked by the alpha5 subunit. Ligand binding and integrin activation may uncover the beta1 stalk region by triggering a conformational shift of alpha5 relative to beta1.
Tsuchida
J
J
Department of Biological Sciences, Tokyo Institute of Technology, Midori-ku, Yokohama 226, Japan.
Ueki
S
S
Takada
Y
Y
Saito
Y
Y
Takagi
J
J
D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D001666:Binding Sites, Antibody; D001667:Binding, Competitive; D016466:CHO Cells; D006224:Cricetinae; D000939:Epitopes / Q000378:metabolism; D006801:Humans; D020014:K562 Cells; D008024:Ligands; D011233:Precipitin Tests; D011487:Protein Conformation; D017469:Receptors, Fibronectin / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D008024:Ligands; D017469:Receptors, Fibronectin