Functional and structural correlations of individual alphaIIbbeta3 molecules.
The divalent cation Mn(2+) and the reducing agent dithiothreitol directly shift integrins from their inactive to their active states. We used transmission electron microscopy and laser tweezers-based force spectroscopy to determine whether structural rearrangements induced by these agents in the integrin alphaIIbbeta3 correlate with its ability to bind fibrinogen. Mn(2+) increased the probability of specific fibrinogen-alphaIIbbeta3 interactions nearly 20-fold in platelets, and both Mn(2+) and dithiothreitol increased the probability more than 2-fold using purified proteins. Of 3 alphaIIbbeta3 conformations, closed with stalks touching, open with stalks separated, and globular without visible stalks, Mn(2+) and dithiothreitol induced a significant increase in the proportion of open structures, as well as structural changes in the alphaIIbbeta3 headpiece. Mn(2+) also increased the number of complexes between fibrinogen and purified alphaIIbbeta3 molecules, all of which were in the open conformation. Finally, Mn(2+) induced the formation of alphaIIbbeta3 clusters that resulted from interactions exclusively involving the distal ends of the stalks. These results indicate that there is a direct correlation between alphaIIbbeta3 activation and the overall conformation of the molecule. Further, they are consistent with the presence of a linked equilibrium between single inactive and single active alphaIIbbeta3 molecules and active alphaIIbbeta3 clusters.
Litvinov
Rustem I
RI
Department of Cell and Developmental Biology, University of Pennsylvania School of Medicine, 421 Curie Blvd, 1054 BRB II/III, Philadelphia, PA 19104-6058, USA.
Nagaswami
Chandrasekaran
C
Vilaire
Gaston
G
Shuman
Henry
H
Bennett
Joel S
JS
Weisel
John W
JW
D001665:Binding Sites; D001792:Blood Platelets / Q000502:physiology; D004229:Dithiothreitol / Q000494:pharmacology; D005340:Fibrinogen / Q000378:metabolism; D006801:Humans; D008345:Manganese / Q000494:pharmacology; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism*; D011487:Protein Conformation
D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D008345:Manganese; D005340:Fibrinogen; D004229:Dithiothreitol