ADAMTS13-binding IgG are present in patients with thrombotic thrombocytopenic purpura.

Functional assays are commonly used to measure the antibodies of ADAMTS13 found in patients of thrombotic thrombocytopenic purpura (TTP). In this study we used an enzyme-linked immunoassay to analyze the ADAMTS13-binding IgG levels in six groups of individuals: normal, random hospitalized patients, acute TTP, TTP after receiving plasma therapy, TTP in remission, and other types of thrombotic microangiopathy (TMA). The results showed that ADAMTS13-binding IgG levels were elevated in 100% of the acute TTP group, 75% of the TTP group after receiving plasma therapy, and 40% of the remission group. Overall, the ADAMTS13-binding IgG levels correlated with the inhibitory activity levels againstADAMTS13 (r = -0.69, P < 0.0001). The assay also detected elevated IgG binding levels in 5% - 15% of the normal, random, and other TMA control groups. Addition of purified ADAMTS13 protein to the plasma samples suppressed the IgG binding in each of the acute TTP patients, but in none of the non-TTP groups. Serial measurement in a patient that had two exacerbations of TTP within the first three weeks revealed that the ADAMTS13 activity levels remained <0.1 U/ml during this period, and the ADAMTS13-binding IgG remained elevated, suggesting that ADAMTS13 analysis may provide valuable insight to the disease status during the course of therapy. Analysis of ADAMTS13-binding IgG is helpful for the diagnosis and management of TTP.

Crystal structure of the thrombin mutant D221A/D222K: the Asp222:Arg187 ion-pair stabilizes the fast form.

The thrombin mutant D221A/D222K (ARK) does not bind Na+ and has interesting functional properties intermediate between those of the slow and fast forms of wild type. We solved the X-ray crystal structure of ARK bound at exosite I with a fragment of hirudin at 2.4-A resolution. The structure shows a slight collapse of the 186 and 220 loops into the Na+ binding site due to disruption of the Asp222:Arg187 ion-pair. The backbone O atoms of Arg221a and Lys224 are shifted into conformations that eliminate optimal interaction with Na+. A paucity of solvent molecules in the Na+ binding site is also noted, by analogy to what is seen in the structure of the slow form. These findings reinforce the crucial role of the Asp222:Arg187 ion-pair in stabilizing the fast form of thrombin.