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-beta 1 integrin mon oclonal antibody AG89 can detect such a conformational change since it recognizes a ligand-inducible epitope in the stalk-like region of bet a 1 subunits, The binding of a I-125-labeled AG89 Fab fragment to alph a 5 beta 1 integrins on K562 cells was assessed and analyzed by the Sc atchard method. High affinity binding sites for AG89 are present on ce lls treated with ligand peptide. In addition, results revealed that ce lls treated with EDTA also express AG89 binding sites with the same af finity although the number of binding sites is 4-fold lower. AG89 immu noprecipitated alpha 5 beta 1 complexes from surface-labeled K562 cell s treated with ligand peptide. By contrast, it immunoprecipitated only beta 1 chains when the ligand peptide was absent, suggesting that hig h affinity binding sites on EDTA-treated cells are associated with non functional beta 1 monomer. Additional studies show that the epitope fo r AG89 is constitutively exposed on mutant pi that cannot complex with alpha 5. These data suggest that the AG89 epitope is masked by the al pha 5 subunit. Ligand binding and integrin activation may uncover the beta 1 stalk region by triggering a conformational shift of alpha 5 re lative to beta 1.