The long term objective of this study is to understand the mechanism of activation of the platelet integrin GPIIb-IIIa. GPIIb-IIIa is key to platelet adhesion and aggregation. An understanding of its regulation is likely to be important for treating many cardiovascular diseases including myocardial infarction and stroke. GPIIb-IIIa exists on resting platelets in a dormant conformation unable to bind soluble fibrinogen. Upon platelet stimulation, IIb-IIIa becomes capable of binding fibrinogen and mediating platelet aggregation. Although activation of IIb-IIIa is key to platelet function, the mechanism its activation has not been solved. One hypothesis of the study is that proteolytic cleavage of the cytoplasmic domains of IIb-IIIa control the activation state of the integrin. This hypothesis will be tested by characterizing the structural differences in the cytoplasmic domain of two purified forms of IIb-IIIa which differ in activation state. A second goals of the study is to understand the structural basis of activation-dependent ligand binding to IIb-IIIa. Phage-display will be used to select ligands that bind preferentially to the dormant and active forms of the integrin. Results from this study are likely to provide a structure-activity series explaining activation-dependent ligand binding.
A third aim of the study is to understand the kinetic aspects of integrin activation. Does activation result from an increase in ligand association rate or a decrease in ligand dissociation rate? These studies will be performed on whole platelets and with purified forms of dormant and active IIb-IIIa. A final goal is to understand how the divalent ion binding sites on IIb- IIIa influence the activation event. Binding studies will be performed between Ca2+ and purified conformers of dormant and active IIb-IIIa. Results from this analysis should determine which class of ion binding sites regulate activation of the integrin.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
1R01HL058925-01A1
Application #
2605622
Study Section
Hematology Subcommittee 2 (HEM)
Project Start
1998-05-01
Project End
2002-04-30
Budget Start
1998-05-01
Budget End
1999-04-30
Support Year
1
Fiscal Year
1998
Total Cost
Indirect Cost
Name
Sanford-Burnham Medical Research Institute
Department
Type
DUNS #
009214214
City
La Jolla
State
CA
Country
United States
Zip Code
92037
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Yan, B; Hu, D D; Knowles, S K et al. (2000) Probing chemical and conformational differences in the resting and active conformers of platelet integrin alpha(IIb)beta(3). J Biol Chem 275:7249-60
Hu, D D; White, C A; Panzer-Knodle, S et al. (1999) A new model of dual interacting ligand binding sites on integrin alphaIIbbeta3. J Biol Chem 274:4633-9

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