Figure 1.

Sequence and function of the integrin α_IIb-cytoplasmic domain. (A) Sequence alignment of various human integrin α-cytoplasmic domains with both wild-type and mutant α_IIb. The sequence identity and similarity are highlighted in decreased darkness. Residue numbers correspond to α_IIb. (B) Effect of α_IIb-cytoplasmic peptides on the binding of fibrinogen (Fg) to stimulated platelets. Human platelets in Tyrode's buffer alone (buffer), with or without 50 μM (final concentration) of nonmyristoylated α_IIb K989–E1008 (α_IIb), m-α_IIb-wt (m-α_IIb), and myristic acid (Myr) were preincubated for 30 min with 300 nM ¹²⁵I-fibrinogen. The platelets were then stimulated with either 10 μM ADP/20 μM epinephrine (shaded bars) or 50 μM thrombin receptor agonist peptide (hatched bars). After 15 min, ¹²⁵I-fibrinogen binding was measured. Incubation of nonstimulated platelets with cytoplasmic peptides or with myristic acid had no effect on fibrinogen binding (data not shown). (C) Effects of α_IIb-cytoplasmic peptides on the binding of fibrinogen to immunocaptured forms of α_IIbβ₃. Both forms of the receptor, platelet α_IIbβ₃ (shaded bars) or sr-α_IIbβ₃ (hatched bars), were immunocaptured onto immobilized monoclonal antibody AP3. The captured receptors were preincubated for 30 min with either buffer alone (buffer) or 50 μM nonmyristoylated α_IIb K989–E1008 (α_IIb). Peptide specificity was confirmed with a nonrelated acidic peptide (control peptide; GDKNADGWIEFEEL). Results are expressed as a percentage of maximal fibrinogen binding in the absence of peptides. (D) Divergent effects of myristoylated wild-type and mutant α_IIb-cytoplasmic peptides on the binding of fibrinogen to stimulated platelets. Platelets were preincubated with varying concentrations of either m-α_IIb-wt (closed circles) or m-α_IIb-mut peptides (open circles), and then ¹²⁵I-fibrinogen binding to ADP/epinephrine-stimulated platelets was measured.