Effect of carbohydrate modifications of factor VIII/von Willebrand factor on binding to platelets.
This study compares the ability of unmodified and carbohydrate-modified forms of factor VIII/von Willebrand factor (FVIII/vWF) protein to bind to platelets in the presence of ristocetin or thrombin. Treatment of intact FVIII/vWF with alpha-D-neuraminidase results in more than 95% desialylation. Asialo FVIII/vWF retains total activity in ristocetin- and thrombin-mediated binding to platelets as demonstrated by direct and competitive binding assays. Examination of its multimeric pattern by sodium dodecyl sulfate-agarose electrophoresis reveals a normal multimeric structure. Treatment of intact FVIII/vWF with beta-D-galactosidase results in the removal of 20% of galactose (agalacto FVIII/vWF) whereas 55% of galactose is released from asialo FVIII/vWF (asialo agalacto FVIII/vWF). Agalacto and asialo-agalacto FVIII/vWF are both unable to bind to platelets in the presence of ristocetin. In contrast, they still bind to thrombin-stimulated human (except thrombasthenic) platelets. Removal of either ultimate (agalacto FVIII/vWF) or ultimate and penultimate (asialo-agalacto FVIII/vWF) galactose results in the same loss of the larger molecular weight multimers and in an increase of smaller multimers. These results suggest (1) that sialic acid does not play a significant role in ristocetin- or thrombin-mediated FVIII/vWF-platelets interactions and multimeric structure of FVIII/vWF (2) that ultimate beta-linked galactose residues are essential for the maintenance of a normal multimer organization (3) that ristocetin- and thrombin-mediated binding of FVIII/vWF to platelets differ in FVIII/vWF galactose requirement.
Goudemand
J
J
Mazurier
C
C
Samor
B
B
Bouquelet
S
S
Montreuil
J
J
Goudemand
M
M
D001234:Aspergillus niger / Q000201:enzymology; D001792:Blood Platelets / Q000378:metabolism*; D002241:Carbohydrates; D002462:Cell Membrane / Q000378:metabolism; D003016:Clostridium perfringens / Q000201:enzymology; D005169:Factor VIII / Q000502:physiology; D005690:Galactose / Q000032:analysis; D006801:Humans; D007700:Kinetics; D009439:Neuraminidase / Q000378:metabolism; D010944:Plants / Q000201:enzymology; D010974:Platelet Aggregation; D010980:Platelet Membrane Glycoproteins; D011956:Receptors, Cell Surface / Q000378:metabolism*; D012794:Sialic Acids / Q000032:analysis; D001616:beta-Galactosidase / Q000378:metabolism; D014841:von Willebrand Factor / Q000378:metabolism*
D002241:Carbohydrates; D010980:Platelet Membrane Glycoproteins; D011956:Receptors, Cell Surface; D012794:Sialic Acids; D014841:von Willebrand Factor; C028254:von Willebrand factor receptor; D005169:Factor VIII; D009439:Neuraminidase; D001616:beta-Galactosidase; D005690:Galactose