Fibrinogen-elongated gamma chain inhibits thrombin-induced platelet response, hindering the interaction with different receptors.
The expression of the elongated fibrinogen gamma chain, termed gamma', derives from alternative splicing of mRNA and causes an insertion sequence of 20 amino acids. This insertion domain interacts with the anion-binding exosite (ABE)-II of thrombin. This study investigated whether and how gamma' chain binding to ABE-II affects thrombin interaction with its platelet receptors, i.e. glycoprotein Ibalpha (GpIbalpha), protease-activated receptor (PAR) 1, and PAR4. Both synthetic gamma' peptide and fibrinogen fragment D*, containing the elongated gamma' chain, inhibited thrombin-induced platelet aggregation up to 70%, with IC(50) values of 42+/-3.5 and 0.47+/-0.03 microm, respectively. Solid-phase binding and spectrofluorimetric assays showed that both fragment D* and the synthetic gamma' peptide specifically bind to thrombin ABE-II and competitively inhibit the thrombin binding to GpIbalpha with a mean K(i) approximately 0.5 and approximately 35 microm, respectively. Both these gamma' chain-containing ligands allosterically inhibited thrombin cleavage of a synthetic PAR1 peptide, of native PAR1 molecules on intact platelets, and of the synthetic chromogenic peptide D-Phe-pipecolyl-Arg-p-nitroanilide. PAR4 cleavage was unaffected. In summary, fibrinogen gamma' chain binds with high affinity to thrombin and inhibits with combined mechanisms the platelet response to thrombin. Thus, its variations in vivo may affect the hemostatic balance in arterial circulation.
Lancellotti
Stefano
S
Institute of Internal Medicine and Geriatrics, and Haemostasis Research Centre, Catholic University School of Medicine, 00168 Rome, Italy.
Rutella
Sergio
S
De Filippis
Vincenzo
V
Pozzi
Nicola
N
Rocca
Bianca
B
De Cristofaro
Raimondo
R
D001665:Binding Sites; D001667:Binding, Competitive; D001792:Blood Platelets / Q000378:metabolism*; D005340:Fibrinogen / Q000737:chemistry*; D006801:Humans; D006868:Hydrolysis; D007700:Kinetics; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D044463:Receptor, PAR-1 / Q000378:metabolism; D018179:Receptors, Thrombin / Q000737:chemistry; D013050:Spectrometry, Fluorescence / Q000379:methods; D013917:Thrombin / Q000737:chemistry*
D010455:Peptides; D044463:Receptor, PAR-1; D018179:Receptors, Thrombin; C027539:fibrinopeptides gamma; D005340:Fibrinogen; D013917:Thrombin; C112908:protease-activated receptor 4
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