Ligand binding to integrin alphaIIbbeta3 is dependent on a MIDAS-like domain in the beta3 subunit.
Substitution of beta3 residue Asp119, Ser121, or Ser123 results in a loss of the ligand binding function of integrin alphaIIbbeta3. 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 alpha 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 alphaM I domain to produce a three-dimensional model of the homologous region in the integrin beta3 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 Asp217 and Glu220 as residues essential for the ligand binding function of alphaIIbbeta3. 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, beta3 Asp217 is analogous to a metal-coordinating residue in the alphaM MIDAS domain, while Glu220 does not correspond to a functional MIDAS domain residue. Substitution of the highly conserved beta3 residue Thr197 corresponding to a critical MIDAS metal-coordinating Thr residue did not affect ligand binding function, suggesting that this region of beta3 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.
Tozer
E C
EC
Department of Vascular Biology, The Scripps Research Institute, La Jolla, California 92037, USA.
Liddington
R C
RC
Sutcliffe
M J
MJ
Smeeton
A H
AH
Loftus
J C
JC
D000595:Amino Acid Sequence; D000818:Animals; D001224:Aspartic Acid; D001665:Binding Sites; D018698:Glutamic Acid; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000378:metabolism*; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D012694:Serine
D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D001224:Aspartic Acid; D018698:Glutamic Acid; D012694:Serine