Oxidative stress on VWF proteolysis

In this issue of Blood, Chen and colleagues demonstrate that the rate of cleaving VWF by ADAMTS13 is significantly slowed when the residue Met¹⁶⁰⁶ in the VWF A2 domain is oxidized.¹  The finding adds a new dimension to the complexity of regulating VWF cleavage and links the rate of VWF proteolysis to the state of oxidative stress.

Von Willebrand factor (VWF) multimers secreted from Weibel-Palade bodies of endothelial cells are enriched in ultra-large forms that are rapidly cleaved by the zinc metalloprotease ADAMTS13. Enzyme deficiency is associated with the development of thrombotic thrombocytopenic purpura as well as with conditions associated with systemic inflammation. There have been extensive studies on how VWF proteolysis is initiated and regulated, mostly focusing on identifying the interface between the substrate and metalloprotease, and specific mutations that alter the rate of cleavage. The current study has delineated a new regulatory mechanism. The study found that Met¹⁶⁰⁶, within the peptide bond (Tyr¹⁶⁰⁵-Met¹⁶⁰⁶) targeted by ADAMTS13, is oxidized to the sulfoxide by hypochlorous acid (HOCl) in vitro. The oxidative modification significantly slows the rate of cleaving an isolate A2 domain and purified VWF multimers by the metalloprotease. The study is significant because it provides the first experimental evidence that VWF proteolysis could be regulated by oxidative stress, which results from accumulation of reactive oxygen species including HOCl.²  The study is also provocative because it raises several interesting and important questions. First, is Met¹⁶⁰⁶ the only amino acid in VWF multimers that is subjected to oxidative modification? The answer is likely to be no, even though the current study is focused on the HOCl modification of a specific methionine residue. VWF contains a high percentage of cysteine residues (8.3%), some of which are in thiol forms that could be sensitive to regulations by redox in general and HOCl in particular. Consistent with the notion, an early report has shown that thiol(s) in the VWF A3 domain is targeted by a reductase activity associated with thrombospondin-1.³  The question remains as to whether different reactive oxygen species differentially modify specific amino acids within a VWF multimer, leading to different functional outcomes. Second, does oxidation also affect VWF adhesion activity? The answer is likely to be yes. There is no direct evidence as to whether VWF with oxidized Met¹⁶⁰⁶ is more or less adhesive, but converting thiols in VWF multimers to disulfide bonds is associated with enhanced VWF binding to platelets.⁴  Third, is VWF oxidation by reactive oxygen species permanent or transient? A disulfide bond is traditionally considered to be a permanent basic posttranslational modification critical for maintaining the tertiary structure of a given protein. However, increasing evidence also suggests that the oxidative modification occurs extracellularly to nonstructural thiols in response to changes in a redox environment.⁵  The blood redox system is composed of proteins and small-molecule thiols, and its balance can be changed in a variety of (patho)physiologic conditions. One well-known example is that the plasma redox potential is approximately 13 to 1 as measured by the ratio of reduced to oxidized glutathione.⁶  However, this reducing state can be made transiently oxidizing in conditions such as oxidative stress. The question is whether the oxidative modification to VWF is reversible when the environment returns to a reducing state. Demonstrating such reversibility will answer a fundamental question as to whether VWF proteolysis can be transiently slowed or inhibited by oxidative stress. If this is indeed the case, such a transient inhibition of VWF proteolysis may explain why some patients with acquired thrombotic thrombocytopenic purpura present with only a moderately reduced ADAMTS13 activity at the time of laboratory assays. This could particularly be the case for patients whose thrombotic crisis is triggered by systemic inflammation, a known condition of oxidative stress. It will therefore be interesting to further investigate the strength and duration of oxidative stress that is required for oxidizing VWF multimers, especially in animal models.