Mutation in the leucine-rich repeat of platelet glycoprotein Ib alpha results in defects in its interaction with immobilized von Willebrand factor under flow

Objective To characterize effects of the GP Ib alpha mutation (A156V) on it s interaction with von Willebrand factor (vWf) under high fluid shear stres s. Methods The residue A156 of GP Ib alpha was converted to a valine and the m utant expressed in CHO cells expressing wild-type GP Ib beta and GPIX. The transfected cells were tested for their interaction with a panel of GP Ib a lpha: antibodies and for rolling on immobilized vWf under high shear. Results The mutation led to surface expression of a GP Ib alpha polypeptide that adopted a different conformation at its N-terminus because binding of the GP Ib alpha antibody AN51, which has a binding epitope in the N-termin al 35 residues, was eliminated, whereas binding of the others (AK2, MB45, a nd SZ2, all of which bind to regions C-terminal to the AN51 epitope) was no rmal. Mutant-expressing cells could adhere and roll on immobilized vWf unde r high fluid shear stress and rolled significantly faster than wild-type ce lls. Conclusion These studies demonstrate that the mutation A156V results in a c onformation change at the N-terminus of GP Ib alpha, which leads to an incr ease in the dissociation rate of the bond between the GP Ib alpha: mutant a nd vWf.