Abstract
To explore the structural basis of ligand binding to alphaIIbbeta3, we conducted a site-directed mutagenesis of Y178, which is located in the ligand-specificity region (C177-C184) of the beta3 subunit. Two mutant beta3 constructs, Y178A and Y178I, were transfected into CHO cells and co-expressed with human alphaIIb subunit on the cell surface. Our results showed that the Y178A mutation affected processing and cell surface exposure of recombinant alphaIIbbeta3 receptor, abrogated the binding of PAC-1, a ligand-mimetic antibody, to alphaIIbbeta3 pre-treated with the known activator DTT. The Y178A mutation also resulted in reduced adhesion of alphaIIbbeta3 on immobilized fibrinogen. In contrast, the interaction of alphaIIbbeta3 with the small molecular ligand RGDS was unaffected by Y178A mutation, as evidenced by the elevated LIBS-1 epitope expression following RGDS addition. Interestingly however, Y178I mutation did not affect the receptor synthesis and function at all. As for post-receptor occupancy, neither Y178A nor Y178I prevented alphaIIbbeta3 translocation to focal adhesion contacts. These results suggest that Y178 is involved in alphaIIb maturation and alphaIIbbeta3 complex expression. This residue is also critical for alp...Continue Reading
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