Ligand binding to integrin alpha(IIb)beta(3) is dependent on a MIDAS-like domain in the beta(3) subunit

Substitution of beta(3) residue Asp(119), Ser(121), or Ser(123) results in a loss of the ligand binding function of integrin alpha(IIb)beta(3). Homologous residues in other integrin beta subunits are similarly critical for ligand binding function. This DXSXS motif is also present in the I domain of certain integrin iv subunits, where it constitutes a portion of the unique metal ion-dependent adhesion site (MIDAS). In this report, we have utilized the crystal structure of the recombinant alpha(M) I domain to produce a three-dimensional model of the homologous region in the integrin beta(3) subunit. We performed mutagenesis of candidate amino acid residues predicted from this model to be involved in cation coordination and ligand binding. We report the identification of Asp(217) and Glu(220) as residues essential for the ligand binding function of alpha(IIb)beta(3). Alanine substitution of these residues did not affect receptor expression but abolished the binding of activation-dependent (PAC1) and -independent (OPG2) ligand mimetic antibodies. In our proposed model, beta(3) Asp(217) is analogous to a metal-coordinating residue in the alpha(M) MIDAS domain, while Glu(220) does not correspond to a functional MIDAS domain residue. Substitution of the highly conserved beta(3) residue Thr(197) corresponding to a critical MIDAS metal-coordinating Thr residue did not affect ligand binding function, suggesting that this region of beta(3) adopts a structure that is very similar to but not identical to that of the MIDAS domain. These data support a functional linkage between these two sequences and further define a common feature of ligand binding to integrins.