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Published on: April 13, 2019
O-fucosylation is required for ADAMTS13 secretion
Lindsay M Ricketts1, Malgosia Dlugosz, Kelvin B Luther
1Department of Internal Medicine, Division of Hematology, Washington University School of Medicine, St. Louis, Missouri, 63110, USA.
The Journal of Biological Chemistry
|March 31, 2007
Summary
O-fucosylation, the addition of fucose sugars to ADAMTS13 protein, is crucial for its secretion. This modification ensures proper function of ADAMTS13, preventing thrombotic thrombocytopenic purpura.
Area of Science:
- Biochemistry
- Molecular Biology
- Hematology
Background:
- ADAMTS13 is a metalloproteinase that cleaves von Willebrand factor, preventing thrombotic disorders.
- Deficiency in ADAMTS13 activity causes thrombotic thrombocytopenic purpura.
- ADAMTS13 possesses thrombospondin type 1 repeats (TSRs) with potential glycosylation sites.
Purpose of the Study:
- To investigate the role of O-fucosylation in ADAMTS13 secretion and function.
- To identify the specific O-fucose structures on ADAMTS13.
- To determine the impact of O-fucosylation on ADAMTS13 secretion.
Main Methods:
- Mass spectral analysis of tryptic peptides from human ADAMTS13.
- Metabolic labeling with radioactive fucose.
- Site-directed mutagenesis of potential O-fucosylation sites in TSRs.
- siRNA-mediated knockdown of protein O-fucosyltransferase 2 (POFUT2).
- Expression of ADAMTS13 in cell lines with altered GDP-fucose synthesis.
Main Results:
- At least six TSRs in ADAMTS13 are modified with a glucose-beta1,3-fucose disaccharide.
- Mutating serine residues in TSRs significantly reduced ADAMTS13 secretion.
- Knockdown of POFUT2 or impaired GDP-fucose synthesis decreased ADAMTS13 secretion.
- Overexpression of POFUT2 partially rescued secretion of a double mutant ADAMTS13.
Conclusions:
- O-fucosylation of ADAMTS13 is essential for its efficient secretion.
- The specific O-fucose disaccharide structure is important for this process.
- These findings highlight the functional significance of protein O-fucosylation in ADAMTS13 biology and secretion.
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