Human von Willebrand factor: a multivalent protein composed of identical subunits

Abstract

A large-scale method for the isolation of von Willebrand factor (vWF) from human factor VIII concentrates was developed in order to study the structure of this protein and its platelet binding activity. vWF is composed of a number of glycoprotein subunits that are linked together by disulfide bonds to form a series of multimers. These multimers appear to contain an even number of subunits of 270K. Two minor components of Mr 140K and 120K were also identified, but these chains appear to result from minor proteolysis. The smallest multimer of vWF contained nearly equimolar amounts of the 270K, 140K, and 120K subunits, while the largest multimers contained less than 20% of the two minor components. Amino acid sequence analysis, amino acid composition, and cleavage by cyanogen bromide indicate that the 270K subunits are identical and each is a single polypeptide chain with an amino-terminal sequence of Ser-Leu-Ser-Cys-Arg-Pro-Pro-Met-Val-Lys and a carboxyl-terminal sequence of Glu-Cys-Lys-Cys-Ser-Pro-Arg-Lys-Cys-Ser-Lys. Platelet binding in the presence of ristocetin was 8-fold greater with multimers larger than five (i.e., containing more than 10 subunits of 270K) as compared to multimers less than three (containing less than six subunits of 270K). However, partially reduced vWF (Mr 500K), regardless of whether it was prepared from large or small molecular weight multimers, gave platelet binding similar to that of the smallest multimers. Likewise, partial proteolysis by elastase, thermolysin, trypsin, or chymotrypsin produced small "multimer-like" proteins with platelet binding properties similar to either partially reduced vWF or to the smallest multimers. We conclude that human vWF contains identical 270K subunits assembled into a multivalent structure. Disassembly by either partial reduction or partial proteolysis produces essentially monovalent protein with platelet binding properties similar to that of the smallest multimers. Multivalency is likely the primary factor responsible for the increase in biological activity with multimer size.

Specific von Willebrand factor–cleaving protease in thrombotic microangiopathies: a study of 111 cases

“…25,26 In the present study, neither UL nor increased LMW multimers of vWF were consistently found in TMA. This discrepancy concerning the LMW multimers may be methodologic because we chose the first 5-mers to define the LMW forms, 29 whereas Galbusera and colleagues chose the 2 fastest migrating bands of the multimeric profile. 25,26 Furthermore, searching for statistical relationships between vWFcleaving protease and other laboratory features may be interesting to establish a pathogenic link between vWF-cleaving protease deficiency and the variation of the multimeric profile of vWF.…”

A monoclonal antibody recognizes a von Willebrand factor domain within the amino-terminal portion of the subunit that modulates the function of the glycoprotein IB- and IIB/IIIA-binding domains.

“…vWf has a central role in platelet adhesion and additionally is a carrier protein for coagulation factor VIII (1). It is a large multimeric protein built from identical subunits (2). The assembly of subunits gives rise to a series of multimers with molecular masses varying between 500 and 20,000 kD.…”

Differential proteolytic activation of factor VIII-von Willebrand factor complex by thrombin.

“…were BJV-VIIIcp, 28 kDa (by NaDodSO4/PAGE); fVIII, 200 kDa (average of the heterodimeric mixture by NaDodSO4/ PAGE); fVIIIaIia and fVIIIaBJ, 160 kDa (7); thrombin, 37 kDa (14); vWf subunit, 270 kDa (19).…”