Flavocetin-A and -B, two high molecular mass glycoprotein Ib binding proteins with high affinity purified from Trimeresurus flavoviridis venom, inhibit platelet aggregation at high shear stress.
Two high molecular mass proteins, flavocetin-A and flavocetin-B, were purified from Trimeresurus flavoviridis venom. On polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, the apparent molecular mass of flavocetin-A and -B were 149 and 139 kDa, respectively, under nonreducing conditions. On reduction, flavocetin-A showed two distinct subunits (17 and 14 kDa), and flavocetin-B three distinct subunits (17, 15 and 14 kDa). At 1 microgram/ml, flavocetin-A and -B (flavocetins) inhibited the von Willebrand factor (vWF)-dependent aggregation of fixed human platelets. However, flavocetins (10 micrograms/ml) had no effect on ADP- and collagen-induced platelet aggregation in PRP. Flavocetins (3 micrograms/ml) also inhibited shear-induced platelet aggregation at high shear stress. Furthermore, flavocetin-A completely inhibited the aggregation of and ATP release from washed platelets stimulated with a low concentration of thrombin. Flavocetin-A specifically bound to platelet with high affinity (Kd = 0.35 +/- 0.13 nM) at 21,500 +/- 1760 binding sites per platelet. The N-terminal amino acid sequences of the subunits of flavocetin-A show a high degree of homology with those of echicetin, botrocetin, alboaggregin-B and factor IX/factor X-binding protein. These results suggest that flavocetins may be a useful tool for further investigation of the GPIb-vWF interaction.
Taniuchi
Y
Y
Institute for Drug Discovery Research, Yamanouchi Pharmaceutical Co., Ltd., Ibaraki, Japan.
Kawasaki
T
T
Fujimura
Y
Y
Suzuki
M
M
Titani
K
K
Sakai
Y
Y
Kaku
S
S
Hisamichi
N
N
Satoh
N
N
Takenaka
T
T
D000255:Adenosine Triphosphate / Q000097:blood; D000595:Amino Acid Sequence; D000818:Animals; D055592:Biophysical Phenomena; D001703:Biophysics; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism; D002352:Carrier Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*; D003435:Crotalid Venoms / Q000737:chemistry*; D004912:Erythrocytes / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009504:Neutrophils / Q000378:metabolism; D010974:Platelet Aggregation; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology*; D016415:Sequence Alignment; D013917:Thrombin / Q000494:pharmacology; D017841:Trimeresurus; D014841:von Willebrand Factor / Q000378:metabolism / Q000494:pharmacology
D002352:Carrier Proteins; D003435:Crotalid Venoms; D010975:Platelet Aggregation Inhibitors; D014841:von Willebrand Factor; D000255:Adenosine Triphosphate; D013917:Thrombin