Recognition by the platelet integrin alphaiibB3 of macromolecular ligands, particularly fibrinogen, is a key step in hemostasis. Activation of alphaiibB3 involves some of the largest conformational changes known in proteins, including leg extension and head opening. Several therapeutics that target alphaiibB3 are used during percutaneous interventions;however, they also induce the active conformation and can lead to thrombocytopenia. Understanding the structure of integrin alphaiibB3, how its conformation is regulated during platelet activation, and how it binds to macromolecular ligands and small molecule therapeutics, is key to a better understanding of vascular processes and developing improved therapeutics. Furthermore, alphaiibB3 is a common target of antibodies that are responsible for immune thrombocytopenia purpura (ITP), and understanding the molecular basis of ITP is important for improvements in therapy.
Four aims address these needs. 1. Crystal structures and molecular dynamics examine the atomic pathway between the closed and open headpieces, including four intermediate states, and demonstrate opening by RGD and therapeutic RGD-mimetics of the headpiece. Structures of alphaiibB3 bound to RUC-2 advance second-generation integrin antagonists that do not induce opening. 2. How alphaiibB3 recognizes macromolecular ligands will be examined with complexes with one or more ligands including the fibrinogen yC module, fibronectin Fn3 modules 9 and 10, Del-1 EGF domains, and a disintegrin. The structures will show how the non-RGD portions of ligands contribute specificity and affinity. 3. We examine the biology of integrin activation by measuring monomeric affinity for the fibrinogen yC module on resting and activated platelets. The effects of function-perturbing or function-dependent Fab including LIBS Fab and ligand-mimetic Fab are correlated with the conformation of intact alphaiibB3 in complex with the Fab in EM. 4. We will examine the structural basis of ITP. Crystal structures of drug-dependent complexes between Fab and alphaiibB3 are examined in model systems using a drug with no known propensity for alphaiibB3 (quinine) or a specific antagonist (eptifibatide). A new concept is advanced that drugs can perturb the backbone conformation of antibody variable region loops. Complexes with intact alphaiibB3 in EM examine specificity of drug-dependent Fab from patients with ITP.

Public Health Relevance

Structures of molecules on platelets are important to understand disorders of bleeding and thrombosis, and immune destruction of platelets. The structures determined in this work will be important for development of drugs to prevent thrombosis, and improved treatments of patients that have bleeding disorders due to abnormally low numbers of platelets.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Program Projects (P01)
Project #
5P01HL103526-05
Application #
8695446
Study Section
Heart, Lung, and Blood Program Project Review Committee (HLBP)
Project Start
Project End
Budget Start
2014-07-01
Budget End
2015-06-30
Support Year
5
Fiscal Year
2014
Total Cost
Indirect Cost
Name
Children's Hospital Boston
Department
Type
DUNS #
City
Boston
State
MA
Country
United States
Zip Code
02115
Xu, Shutong; Wang, Jianchuan; Wang, Jia-Huai et al. (2017) Distinct recognition of complement iC3b by integrins ?X?2 and ?M?2. Proc Natl Acad Sci U S A 114:3403-3408
Chen, Bing; Chou, James J (2017) Structure of the transmembrane domain of HIV-1 envelope glycoprotein. FEBS J 284:1171-1177
Li, Jing; Springer, Timothy A (2017) Integrin extension enables ultrasensitive regulation by cytoskeletal force. Proc Natl Acad Sci U S A 114:4685-4690
Fu, Hongxia; Jiang, Yan; Yang, Darren et al. (2017) Flow-induced elongation of von Willebrand factor precedes tension-dependent activation. Nat Commun 8:324
Piai, Alessandro; Fu, Qingshan; Dev, Jyoti et al. (2017) Optimal Bicelle Size q for Solution NMR Studies of the Protein Transmembrane Partition. Chemistry 23:1361-1367
Li, Jing; Su, Yang; Xia, Wei et al. (2017) Conformational equilibria and intrinsic affinities define integrin activation. EMBO J 36:629-645
Lin, Fu-Yang; Zhu, Jianghai; Eng, Edward T et al. (2016) ?-Subunit Binding Is Sufficient for Ligands to Open the Integrin ?IIb?3 Headpiece. J Biol Chem 291:4537-46
Fu, Qingshan; Fu, Tian-Min; Cruz, Anthony C et al. (2016) Structural Basis and Functional Role of Intramembrane Trimerization of the Fas/CD95 Death Receptor. Mol Cell 61:602-613
Dev, Jyoti; Park, Donghyun; Fu, Qingshan et al. (2016) Structural basis for membrane anchoring of HIV-1 envelope spike. Science 353:172-175
Chen, Xinping; Zhao, Chunyue; Li, Xiaolong et al. (2015) Terazosin activates Pgk1 and Hsp90 to promote stress resistance. Nat Chem Biol 11:19-25

Showing the most recent 10 out of 31 publications