The C-terminal domains of ADAMTS1 contain exosites involved in its proteoglycanase activity

Minns, Alexander Frederick, Qi, Yawei, Yamamoto, Kazuhiro ORCID: 0000-0002-8481-775X, Lee, Karen and Santamaria, Salvatore
(2022) The C-terminal domains of ADAMTS1 contain exosites involved in its proteoglycanase activity. JOURNAL OF BIOLOGICAL CHEMISTRY, 299 (4). 103048-.

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A disintegrin-like and metalloproteinase with thrombospondin type 1 motifs (ADAMTS1) is a protease involved in fertilization, cancer, cardiovascular development, and thoracic aneurysms. Proteoglycans such as versican and aggrecan have been identified as ADAMTS1 substrates, and Adamts1 ablation in mice typically results in versican accumulation; however, previous qualitative studies have suggested that ADAMTS1 proteoglycanase activity is weaker than that of other family members such as ADAMTS4 and ADAMTS5. Here, we investigated the functional determinants of ADAMTS1 proteoglycanase activity. We found that ADAMTS1 versicanase activity is approximately 1000-fold lower than ADAMTS5 and 50-fold lower than ADAMTS4 with a kinetic constant (k<sub>cat</sub>/K<sub>m</sub>) of 3.6 × 10<sup>3</sup> M<sup>-1</sup> s<sup>-1</sup> against full-length versican. Studies on domain-deletion variants identified the spacer and cysteine-rich domains as major determinants of ADAMTS1 versicanase activity. Additionally, we confirmed that these C-terminal domains are involved in the proteolysis of aggrecan as well as biglycan, a small leucine-rich proteoglycan. Glutamine scanning mutagenesis of exposed positively charged residues on the spacer domain loops and loop substitution with ADAMTS4 identified clusters of substrate-binding residues (exosites) in β3-β4 (R756Q/R759Q/R762Q), β9-β10 (residues 828-835), and β6-β7 (K795Q) loops. This study provides a mechanistic foundation for understanding the interactions between ADAMTS1 and its proteoglycan substrates and paves the way for development of selective exosite modulators of ADAMTS1 proteoglycanase activity.

Item Type: Article
Uncontrolled Keywords: Animals, Mice, Aggrecans, Versicans, ADAMTS1 Protein, ADAMTS4 Protein, ADAMTS5 Protein
Divisions: Faculty of Health and Life Sciences
Faculty of Health and Life Sciences > Institute of Life Courses and Medical Sciences
Depositing User: Symplectic Admin
Date Deposited: 03 Mar 2023 10:53
Last Modified: 28 Jun 2023 22:31
DOI: 10.1016/j.jbc.2023.103048
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