Proteolytic activities of human ADAMTS-5: comparative studies with ADAMTS-4.
Aggrecanases have been characterized as proteinases that cleave the Glu373-Ala374 bond of the aggrecan core protein, and they are multidomain metalloproteinases belonging to the ADAMTS (adamalysin with thrombospondin type 1 motifs) family. The first aggrecanases discovered were ADAMTS-4 (aggrecanase 1) and ADAMTS-5 (aggrecanase 2). They contain a zinc catalytic domain followed by non-catalytic ancillary domains, including a disintegrin domain, a thrombospondin domain, a cysteine-rich domain, and a spacer domain. In the case of ADAMTS-5, a second thrombospondin domain follows the spacer domain. We previously reported that the non-catalytic domains of ADAMTS-4 influence both its extracellular matrix interaction and proteolytic abilities. Here we report the effects of these domains of ADAMTS-5 on the extracellular matrix interaction and proteolytic activities and compare them with those of ADAMTS-4. Although the spacer domain was critical for ADAMTS-4 localization in the matrix, the cysteine-rich domain influenced ADAMTS-5 localization. Similar to previous reports of other ADAMTS family members, very little proteolytic activity was detected with the ADAMTS-5 catalytic domain alone. The sequential inclusion of each carboxyl-terminal domain enhanced its activity against aggrecan, carboxymethylated transferrin, fibromodulin, decorin, biglycan, and fibronectin. Both ADAMTS-4 and -5 had a broad optimal activity at pH 7.0-9.5. Aggrecanolytic activities were sensitive to the NaCl concentration, but activities on non-aggrecan substrates, e.g. carboxymethylated transferrin, were not affected. Although ADAMTS-4 and ADAMTS-5 had similar general proteolytic activities, the aggrecanase activity of ADAMTS-5 was at least 1,000-fold greater than that of ADAMTS-4 under physiological conditions. Our studies suggest that ADAMTS-5 is a major aggrecanase in cartilage metabolism and pathology.
Gendron
Christi
C
Kennedy Institute of Rheumatology Division, Imperial College London, London W6 8LH, United Kingdom.
Kashiwagi
Masahide
M
Kennedy Institute of Rheumatology Division, Imperial College London, London W6 8LH, United Kingdom.
Lim
Ngee Han
NH
Kennedy Institute of Rheumatology Division, Imperial College London, London W6 8LH, United Kingdom.
Enghild
Jan J
JJ
Department of Molecular Biology, University of Aarhus, Science Park, DK-8000 Aarhus C, Denmark.
Thøgersen
Ida B
IB
Department of Molecular Biology, University of Aarhus, Science Park, DK-8000 Aarhus C, Denmark.
Hughes
Clare
C
Connective Tissue Laboratories, Cardiff School of Biosciences, University of Cardiff, Cardiff CF1 3US, United Kingdom.
Caterson
Bruce
B
Connective Tissue Laboratories, Cardiff School of Biosciences, University of Cardiff, Cardiff CF1 3US, United Kingdom.
Nagase
Hideaki
H
Kennedy Institute of Rheumatology Division, Imperial College London, London W6 8LH, United Kingdom. Electronic address: h.nagase@imperial.ac.uk.
D051722:ADAM Proteins / Q000737:chemistry / Q000502:physiology*; D000071121:ADAMTS4 Protein; D000071116:ADAMTS5 Protein; D000409:Alanine / Q000737:chemistry; D001665:Binding Sites; D020134:Catalytic Domain; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D017353:Gene Deletion; D018698:Glutamic Acid / Q000737:chemistry; D006801:Humans; D006863:Hydrogen-Ion Concentration; D009154:Mutation; D011348:Procollagen N-Endopeptidase / Q000737:chemistry / Q000502:physiology*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D014162:Transfection
D018698:Glutamic Acid; D051722:ADAM Proteins; D000071116:ADAMTS5 Protein; C121532:ADAMTS5 protein, human; D011348:Procollagen N-Endopeptidase; D000071121:ADAMTS4 Protein; C000603918:ADAMTS4 protein, human; D000409:Alanine