Mechanistic insights from a refined three-dimensional model of integrin αIIbβ3

Abstract

The integrin αIIbβ3 plays an important role in platelet function, and abnormalities of this protein result in a serious bleeding disorder, known as Glanzmann thrombasthenia. Although crystallographic data exist for the related integrin αVβ 3, to date, there are no high resolution structures of integrin αIIbβ3 available in the literature. Therefore, it is still unclear how specific elements of the αIIb subunit contribute to integrin αIIbβ3 function. Here we describe a refined model of the αIIb N-terminal portion of integrin αIIbβ3 obtained by using the αVβ3 template combined with a new method for predicting the conformations of the unique αIIb loop regions comprising residues 71-85, 114-125, and 148-164. The refined model was probed based on a structural prediction that differentiates it from standard homology models: specifically, that Lys-118 of αIIb contacts Glu-171 of β3. To test this hypothesis experimentally, the mutant integrin chains αIIb K118C and β3 E171C were cotransfected into HEK 293 cells. We show that the cells expressed the mutants αIIbβ3 on their surface as a disulfide-linked dimer, supporting the close proximity between αIIb Lys-118 and β3 Glu-171 predicted from the refined model. This validated model provides a specific structural context for the analysis and interpretation of structure-function relations of integrin α IIbβ3. In addition, it suggests mechanistic hypotheses pertaining to both naturally occurring mutations responsible for Glanzmann thrombasthenia and to point mutations that affect ligand binding.