α2-6–linked sialic acid enhances ADAMTS13 proteolysis by reducing exposure of subterminal D-galactose. (A) The effects of α2-3,6,8,9 neuraminidase treatment in regulating VWF susceptibility to ADAMTS13 proteolysis were investigated at different urea concentrations. At 2M urea, the rate of VWF proteolysis was significantly increased for both pd-VWF and Neu-VWF compared with 1.5M urea. However, the resistant phenotype of Neu-VWF became less apparent at the higher concentration of urea. (B) Removal of capping α2-6–linked sialic acid from the N-linked glycans of VWF results in exposure of penultimate D-galactose residues. To determine whether the protective effect of α2-3,6,8,9 neuraminidase treatment was mediated through loss of sialic acid or was attributable to the subsequent increased galactose exposure, we studied the effects of combined α2-3,6,8,9 neuraminidase and β-galactosidase digestion. After combined glycosidase digestion, the protective effect of loss of α2-6–linked sialic acid only was no longer observed.