von Willebrand factor (VWF) is a large multimeric protein whose monomers are of the domain structure D’-D3-A1-A2-A3-D4-B1-B2-B3C1-C2. The three tandem A domains mediate several important functions of the protein including supporting platelet adhesion, through an interaction of A1 with the platelet GP Ib-IX-V complex, and binding collagen, through A3 and A1. Further, the A2 domain contains the cleavage site for the plasma metalloprotease ADAMTS-13, which processes the newly synthesized and extremely adhesive ultra-large forms of VWF (ULVWF) to the forms normally found in plasma. Our goal was to compare the functions of the individual A domains as isolated domains to their function in the context of the A1-A2-A3 fragment. All recombinant domains were produced in bacteria. The A1-A2-A3 fragment contained an N-terminal thioredoxin-tag and a C-terminal 6Xhis tag and was purified in 2 steps using nickel and heparin columns. The recombinant protein was correctly folded, as indicated by the fact that it was recognized by a number of conformation-specific antibodies. Using ELISA, we examined the interaction of the fragment with both glycocalicin (the soluble extracellular domain of GP Iba) and collagen. At equivalent concentrations, A1-A2-A3 bound collagen better than did the isolated A1 and A3 domains, but the binding to glycocalicin was approximately 7.5 times lower than the A1 domain. We then tested the ability of the fragment to support platelet adhesion under flow, by coating the fragment onto a coverslip that made up the bottom of a parallel-plate flow chamber. In contrast to the findings with immobilized glycocalicin, immobilized A1-A2-A3 supported the attachment, rolling, and firm adhesion of the platelets to the surface, whereas A1 did not support firm adhesion. Finally, we tested cleavage of the fragment by plasma ADAMTS-13, using both ELISA and western blotting to detect cleavage. The rate of cleavage of the A1-A2-A3 fragment was much slower than that of the recombinant A2 domain, needing a longer incubation time to achieve a similar percentage of cleavage in a 15 fold higher concentration of plasma. These results show clearly that within the triplicate A domains of VWF, adjacent A domains influence each other’s functions, possibly by binding each other or by conformational modulation. The fact that the fragment is able to support firm platelet adhesion in isolation suggests that it may contain a second, previously unidentified binding site for platelet receptors.