Conformational change in the integrin extracellular domain is required for high affinity ligand binding and is also involved in post-ligand binding cellular signaling. Although there is evidence to the contrary, electron microscopic studies showing that ligand binding triggers α- and β-subunit dissociation in the integrin head-piece have gained popularity and support the hypothesis that head separation activates integrins. To test directly the head separation hypothesis, we enforced head association by introducing disulfide bonds across the interface between the α-subunit β-propeller domain and the β-subunit I-like domain. Basal and activation-dependent ligand binding by αIIbβ3 and αVβ3 was unaffected. The covalent linkage prevented dissociation of αIIbβ3 into its subunits on EDTA-treated cells. Whereas EDTA dissociated wild type αIIbβ3 on the cell surface, a ligand-mimetic Arg-Gly-Asp peptide did not, as judged by binding of complex-specific antibodies. Finally, a high affinity ligand-mimetic compound stabilized noncovalent association between αIIb and β3 headpiece fragments in the presence of SDS, indicating that ligand binding actually stabilized subunit association at the head, as opposed to the suggested subunit separation. The mechanisms of conformational regulation of integrin function should therefore be considered in the context of the associated αβ headpiece.
Publication Source (Journal or Book title)
Journal of Biological Chemistry
Luo, B., Springer, T., & Takagi, J. (2003). High affinity ligand binding by integrins does not involve head separation. Journal of Biological Chemistry, 278 (19), 17185-17189. https://doi.org/10.1074/jbc.M301516200