title: The novel S527F mutation in the integrin beta3 chain induces a high affinity alphaIIbbeta3 receptor by hindering adoption of the bent conformation.
abstract: Three heterozygous mutations were identified in the genes encoding platelet integrin receptor alphaIIbbeta3 in a patient with an ill defined platelet disorder: one in the beta3 gene (S527F) and two in the alphaIIb gene (R512W and L841M). Five stable Chinese hamster ovary cell lines were constructed expressing recombinant alphaIIbbeta3 receptors bearing the individual R512W, L841M, or S527F mutation; both the R512W and L841M mutations; or all three mutations. All receptors were expressed on the cell surface, and mutations R512W and L841M had no effect on integrin function. Interestingly, the beta3 S527F mutation produced a constitutively active receptor. Indeed, both fibrinogen and the ligand-mimetic antibody PAC-1 bound to non-activated alphaIIbbeta3 receptors carrying the S527F mutation, indicating that the conformation of this receptor was altered and corresponded to the high affinity ligand binding state. In addition, the conformational change induced by S527F was evident from basal anti-ligand-induced binding site antibody binding to the receptor. A molecular model bearing this mutation was constructed based on the crystal structure of alphaIIbbeta3 and revealed that the S527F mutation, situated in the third integrin epidermal growth factor-like (I-EGF3) domain, hindered the alphaIIbbeta3 receptor from adopting a wild type-like bent conformation. Movement of I-EGF3 into a cleft in the bent conformation may be hampered both by steric hindrance between Phe(527) in beta3 and the calf-1 domain in alphaIIb and by decreased flexibility between I-EGF2 and I-EGF3.
authors.affiliation: Laboratory for Thrombosis Research, Interdisciplinary Research Center, Katholieke Universiteit Leuven Campus Kortrijk, 8500 Kortrijk, Belgium. Karen.Vanhoorelbeke@kuleuven-kortrijk.be
authors.forename: Karen
authors.initials: K
authors.lastname: Vanhoorelbeke
authors.forename: Simon F
authors.initials: SF
authors.lastname: De Meyer
authors.forename: Inge
authors.initials: I
authors.lastname: Pareyn
authors.forename: Chantal
authors.initials: C
authors.lastname: Melchior
authors.forename: Sebastien
authors.initials: S
authors.lastname: Plançon
authors.forename: Christiane
authors.initials: C
authors.lastname: Margue
authors.forename: Olivier
authors.initials: O
authors.lastname: Pradier
authors.forename: Pierre
authors.initials: P
authors.lastname: Fondu
authors.forename: Nelly
authors.initials: N
authors.lastname: Kieffer
authors.forename: Timothy A
authors.initials: TA
authors.lastname: Springer
authors.forename: Hans
authors.initials: H
authors.lastname: Deckmyn