| Application notes
(see key) | Recommended dilutions
WB: 1/1000 when using colorimetric substrates such as BCIP/NBT, and 1:5000 for chemiluminescent substrates. EDTA/EGTA treatment of tissues or lysates is required to see latent zymogen. Predicted molecular weight: 102 kDa. This antibody recognizes the zymogen of ADAMTS5 at 120 kD in reduced Western blots, activated forms at 73 kD (major bands), and breakdown products at 50 kD and 40 kD in cell lysates. Dilution optimised using Chromogenic detection. Not yet tested in other applications. Optimal dilutions/concentrations should be determined by the end user. |
| Relevance | ADAMTS5, also known as Implantin or Aggrecanase 2, is a member of the larger family of ADAM (A Disintegrin And Metalloproteinase) metalloproteinases containing thrombospondin (TS) repeats. ADAMTS5 (A Disintegrin And Metalloproteinase with Thrombospondin-5 motif) was first described as ADAMTS5, a protein elevated in mice during the peri-implantion period. At the same time, another group identified Aggrecanase 11, a protein elevated in arthritic synovium. The name was later changed to ADAMTS5. ADAMTS5 is expressed in human and mouse. It has been found in heart, lung, cervix, uterus, ovary, brain, cartilage, and numerous other tissues, as well as chondroblastoma cell lines. Initial observations indicated a role for ADAMTS5 in aggrecan cleavage and cartilage destruction, especially in arthritis, and potentially a role in embryo implantation. ADAMTS5 contains the canonical HExxHxxxxxH zinc metalloproteinase motif, and has been shown to efficiently cleave Aggrecan. In addition to the metalloprotease domain, ADAMTS5 has a propeptide domain, a prohormone convertase (PC, furin) cleavage site, a cysteine-rich domain, a spacer domain, and two thrombospondin-1 like domains. ADAMTS5 is inhibited by the endogenous MMP inhibitors (TIMP1, 2, 3, and 4) but most efficiently by TIMP3. Unlike many of the ADAMs proteases, ADAMTS5 does not have a transmembrane domain, and is a secreted protein. Full length ADAMTS5 is a 930 amino acid protein with a predicted molecular mass is 101.7 kDa, but glycosylation and the abundance of cysteine residues gives ADAMTS5 a greater apparent molecular weight on reduced SDS PAGE gels. When ADAMTS5 is secreted, it is cleaved at the furin cleavage site (predicted molecular mass 73.2 kDa) and then further cleaved to generate a range of smaller forms. |
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