Characteristics of the interaction between thrombin exosite 1 and the sequence 269-287 [correction of 269-297] of platelet glycoprotein Ibalpha.
The interaction between GPIb and thrombin promotes platelet activation elicited via the hydrolysis of the thrombin receptor and involves structures located on the segment 238-290 within the N-terminal domain of GPIbalpha and the positively charged exosite 1 on thrombin. We have investigated the ability of peptides derived from the 269-287 sequence of GPIbalpha to interact with thrombin. Three peptides were synthesized, including Ibalpha 269-287 and two scrambled peptides R1 and R2 which are comparable to Ibalpha 269-287 with regards to their content and distribution of anionic residues. However, R2 differs from both Ibalpha 269-287 and R1 by the shifting of one proline from a central position to the N-terminus. By chemical cross-linking, we observed the formation of a complex between 125I-Ibalpha 269-287 and alpha-thrombin that was inhibited by hirudin, the C-terminal peptide of hirudin, sodium pyrophosphate but not by heparin. The complex did not form when gamma-thrombin was substituted for alpha-thrombin. Ibalpha 269-287 produced only slight changes in thrombin amidolytic activity and inhibited thrombin binding to fibrin. R1 and R2 also formed complexes with alpha-thrombin, modified slightly its catalytic activity and inhibited its binding to fibrin. Peptides Ibalpha 269-287 and R1 inhibited platelet aggregation and secretion induced by low thrombin concentrations whereas R2 was without effect. Our results indicate that Ibalpha 269-287 interacts with thrombin exosite 1 via mainly electrostatic interactions, which explains why the scrambled peptides also interact with exosite 1. Nevertheless, the lack of effect of R2 on thrombin-induced platelet activation suggests that proline 280 is important for thrombin interaction with GPIb.
Laboratoire de Recherche sur l'Hémostase et la Thrombose, Faculté de Médecine Xavier Bichat, Paris, France.
M C
MC
Bouton
C
C
Thurieau
M C
MC
Guillin
M
M
Jandrot-Perrus
D000595:Amino Acid Sequence; D020134:Catalytic Domain; D003432:Cross-Linking Reagents; D006801:Humans; D008969:Molecular Sequence Data; D010455:Peptides / Q000097:blood*; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D013917:Thrombin / Q000378:metabolism*
D003432:Cross-Linking Reagents; D010455:Peptides; D019038:Platelet Glycoprotein GPIb-IX Complex; D013917:Thrombin