Isolation and characterization of jararaca GPIb-BP, a snake venom antagonist specific to platelet glycoprotein Ib.
A platelet glycoprotein Ib-binding protein (GPIb-BP) was isolated from the snake venom of Bothrops jararaca. Jararaca GPIb-BP showed a single band with M(r) of 30,000, and two distinct bands with M(r) of 17,000/13,000 under non-reducing and reducing conditions, respectively, on SDS-polyacrylamide gel electrophoresis. Jararaca GPIb-BP itself induced neither platelet aggregation nor serotonin release from platelets, but specifically bound to GPIb (40,629 +/- 2,521 molecules per normal platelet, with Kd 39.1 +/- 2.4 nM at saturation). The purified venom protein completely inhibited ristocetin- or botrocetin-induced von Willebrand factor (vWF) binding, and blocked the bovine vWF binding to GPIb, with IC50 values ranging from 28 to 42 nM, without affecting the platelet aggregation induced by ADP or alpha-thrombin. 125I-jararaca GPIb-BP binding to GPIb was not altered by the presence of human alpha-thrombin. Jararaca GPIb-BP at a final concentration of 104 nM totally abolished vWF-dependent shear-induced platelet aggregation (SIPA) at a high shear stress, but had no effect on SIPA at a low shear stress. Reduced and S-carboxyamido-methylated jararaca GPIb-BP lost its inhibitory activity on SIPA. The NH2-terminal amino acid sequences of the subunits revealed a high degree of homology with those of several Ca(2+)-dependent lectins, especially to those of two functionally opposite venom proteins, botrocetin (a vWF-modulator) and alboaggregin-B (a GPIb-modulator).
Fujimura
Y
Y
Department of Blood Transfusion, Nara Medical University, Japan.
Ikeda
Y
Y
Miura
S
S
Yoshida
E
E
Shima
H
H
Nishida
S
S
Suzuki
M
M
Titani
K
K
Taniuchi
Y
Y
Kawasaki
T
T
D000595:Amino Acid Sequence; D000818:Animals; D017837:Bothrops / Q000378:metabolism*; D002417:Cattle; D016384:Consensus Sequence; D003435:Crotalid Venoms / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000302:isolation & purification* / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010084:Oxidation-Reduction; D010975:Platelet Aggregation Inhibitors / Q000302:isolation & purification* / Q000494:pharmacology; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000037:antagonists & inhibitors*; D010980:Platelet Membrane Glycoproteins; D011485:Protein Binding / Q000187:drug effects; D011956:Receptors, Cell Surface / Q000302:isolation & purification* / Q000378:metabolism; D012310:Ristocetin / Q000037:antagonists & inhibitors / Q000494:pharmacology; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014841:von Willebrand Factor / Q000378:metabolism
D003435:Crotalid Venoms; D010975:Platelet Aggregation Inhibitors; D019038:Platelet Glycoprotein GPIb-IX Complex; D010980:Platelet Membrane Glycoproteins; D011956:Receptors, Cell Surface; C032193:glycoprotein receptor GPIb-IX; D014841:von Willebrand Factor; D012310:Ristocetin; C030961:botrocetin