Effect of Carbohydrate Modifications of Factor VIII/ von Willebrand Factor on Binding to Platelets

Abstract

SummaryThis 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 α-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 β-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 β-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.

Role of A and B blood group antigens in the expression of adhesive activity of von Willebrand factor

“…Sialic acid or galactose moieties have been suspected to play a role in the adhesive activity of VWF (Sodetz et al, 1977;Gralnick, 1978;Kao et al, 1980). However, the observed decrease may have been due to proteolysis caused by contaminating proteases in the enzyme preparations because other studies failed to demonstrate a change in VWF activity by either neuraminidase or b-galactosidase when the experiments were conducted in the presence of protease inhibitors (Federici et al, 1984;Goudemand et al, 1985). On the other hand, recombinant VWF specifically lacking O-linked carbohydrates exhibits less binding to platelets and a diminished capacity to promote ristocetindependent platelet agglutination (Carew et al, 1992).…”

Decreased Sialic Acid Content of Plasma von Willebrand Factor in Precapillary Pulmonary Hypertension

“…A major limitation of the present study however, is that the galactose content of vWF carbohydrate moiety was not measured. The importance of galactose for the maintenance of multimer organization and ability of vWF to bind to platelets has been emphasized (22). Preliminary unpublished data from our laboratory do not suggest any important decrease of vWF-associated galactose residues in pulmonary hypertensive patients.…”

Primary structure of a new tetraantennary glycan of the N-acetyllactosaminic type isolated from human factor VIII/von Willebrand factor

“…The primary structure of a glycopeptide released from .factor VIII/von Willebrand factor by alkaline treatment, based on the results of H-NMR and methylation analysis acid residue, giving the glycopeptide a conformation allowing the lectin to recognize its specific determinant. Such externally exposed galactose residues may play an important role in the FVIII/vWF binding to platelets [23]. It is interesting to note that, like the previously described biantennary glycan of FVIII/vWF [7], the proposed structure possesses galactose residues in nonreducing terminal position and only one sialic acid and fucose residue.…”