Mechanistic insights from a refined three-dimensional model of integrin alphaIIbbeta3.
The integrin alpha(IIb)beta(3) 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 alpha(V)beta(3), to date, there are no high resolution structures of integrin alpha(IIb)beta(3) available in the literature. Therefore, it is still unclear how specific elements of the alpha(IIb) subunit contribute to integrin alpha(IIb)beta(3) function. Here we describe a refined model of the alpha(IIb) N-terminal portion of integrin alpha(IIb)beta(3) obtained by using the alpha(V)beta(3) template combined with a new method for predicting the conformations of the unique alpha(IIb) 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 alpha(IIb) contacts Glu-171 of beta(3). To test this hypothesis experimentally, the mutant integrin chains alpha(IIb) K118C and beta(3) E171C were cotransfected into HEK 293 cells. We show that the cells expressed the mutants alpha(IIb)beta(3) on their surface as a disulfide-linked dimer, supporting the close proximity between alpha(IIb) Lys-118 and beta(3) 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 alpha(IIb)beta(3). In addition, it suggests mechanistic hypotheses pertaining to both naturally occurring mutations responsible for Glanzmann thrombasthenia and to point mutations that affect ligand binding.
Filizola
Marta
M
Department of Physiology & Biophysics, Weill Medical College of Cornell University, New York, New York 10021, USA. maf2037@med.cornell.edu
Hassan
Sergio A
SA
Artoni
Andrea
A
Coller
Barry S
BS
Weinstein
Harel
H
D000409:Alanine / Q000737:chemistry; D000595:Amino Acid Sequence; D001792:Blood Platelets / Q000378:metabolism; D002460:Cell Line; D018360:Crystallography, X-Ray; D003545:Cysteine / Q000737:chemistry; D018076:DNA, Complementary / Q000378:metabolism; D019281:Dimerization; D004220:Disulfides / Q000737:chemistry; D005340:Fibrinogen / Q000737:chemistry; D005434:Flow Cytometry; D006801:Humans; D039302:Integrin alphaVbeta3 / Q000378:metabolism; D008239:Lysine / Q000737:chemistry; D008958:Models, Molecular; D015233:Models, Statistical; D008969:Molecular Sequence Data; D009154:Mutation; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D012984:Software; D013329:Structure-Activity Relationship; D013915:Thrombasthenia / Q000235:genetics; D014162:Transfection
D018076:DNA, Complementary; D004220:Disulfides; D039302:Integrin alphaVbeta3; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D005340:Fibrinogen; D008239:Lysine; D003545:Cysteine; D000409:Alanine