A Disintegrin and Metalloproteinase with Thrombospondin Motifs-5 (ADAMTS-5) Forms Catalytically Active Oligomers |
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Authors: | Hansen J Kosasih Karena Last Fraser M Rogerson Suzanne B Golub Stephanie J Gauci Vincenzo C Russo Heather Stanton Richard Wilson Shireen R Lamande Paul Holden Amanda J Fosang |
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Institution: | From the ‡Department of Paediatrics, University of Melbourne, Parkville 3052, Australia.;the §Murdoch Childrens Research Institute, Royal Children''s Hospital, Parkville 3052, Australia.;the ‖Department of Orthopaedics & Rehabilitation, Oregon Health & Science University, Portland, Oregon 97239, and ;the ¶University of Tasmania, Hobart 7000, Australia |
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Abstract: | The metalloproteinase ADAMTS-5 (A
disintegrin and metalloproteinase with thrombospondin motifs) degrades aggrecan, a proteoglycan essential for cartilage structure and function. ADAMTS-5 is the major aggrecanase in mouse cartilage, and is also likely to be the major aggrecanase in humans. ADAMTS-5 is a multidomain enzyme, but the function of the C-terminal ancillary domains is poorly understood. We show that mutant ADAMTS-5 lacking the catalytic domain, but with a full suite of ancillary domains inhibits wild type ADAMTS activity, in vitro and in vivo, in a dominant-negative manner. The data suggest that mutant ADAMTS-5 binds to wild type ADAMTS-5; thus we tested the hypothesis that ADAMTS-5 associates to form oligomers. Co-elution, competition, and in situ PLA experiments using full-length and truncated recombinant ADAMTS-5 confirmed that ADAMTS-5 molecules interact, and showed that the catalytic and disintegrin-like domains support these intermolecular interactions. Cross-linking experiments revealed that recombinant ADAMTS-5 formed large, reduction-sensitive oligomers with a nominal molecular mass of ∼400 kDa. The oligomers were unimolecular and proteolytically active. ADAMTS-5 truncates comprising the disintegrin and/or catalytic domains were able to competitively block full-length ADAMTS-5-mediated aggrecan cleavage, measured by production of the G1-EGE373 neoepitope. These results show that ADAMTS-5 oligomerization is required for full aggrecanase activity, and they provide evidence that blocking oligomerization inhibits ADAMTS-5 activity. The data identify the surface provided by the catalytic and disintegrin-like domains of ADAMTS-5 as a legitimate target for the design of aggrecanase inhibitors. |
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Keywords: | cartilage cartilage biology metalloprotease oligomer oligomerization ADAMTS ADAMTS5 aggrecan |
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