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Membrane-type 1 MMP (MMP-14) cleaves at three sites in the aggrecan interglobular domain
1Orthopaedic Molecular Biology Research Unit, Melbourne University, Royal Children's Hospital, Parkville, Australia. fosang@cryptic.rch.unimelb.edu.au
Abstract:
An aggrecan G1-G2 substrate was used to determine sites within the interglobular domain that were susceptible to cleavage by MT1-MMP. Degradation products were identified by Western blotting with neo-epitope antibodies specific for MMP-derived N- and C-terminal sequences. The results showed that MT1-MMP cleaved at the N341-F342 and D441-L442 bonds, as shown for other MMPs, and also at a site 13 amino acids C-terminal to the N341-F342 site. The G2 product of this additional cleavage was identified by sequence analysis and revealed an N-terminus commencing T355VxxPDVELPLP. The data are consistent with MT1-MMP cleavage at three sites in the aggrecan interglobular domain; one at N342-F342, a second at D441-L442 and a third at Q354-T355.
Insights
Matrix metalloproteinase-13 (MT1-MMP) cleaves aggrecan within its interglobular domain at three specific sites. This enzyme
Area of Science:
- Biochemistry
- Molecular Biology
- Enzymology
Background:
- Aggrecan is a major proteoglycan in cartilage.
- Matrix metalloproteinases (MMPs) play a role in tissue degradation.
- MT1-MMP is a specific type of MMP involved in extracellular matrix remodeling.
Purpose of the Study:
- To identify the precise cleavage sites of MT1-MMP within the aggrecan interglobular domain.
- To characterize the degradation products generated by MT1-MMP activity on aggrecan.
Main Methods:
- Utilized an aggrecan G1-G2 substrate for enzymatic assays.
- Employed Western blotting with neo-epitope antibodies to detect MMP-generated fragments.
- Performed sequence analysis to identify the N-terminus of degradation products.
Main Results:
- MT1-MMP cleaved aggrecan at the N341-F342 and D441-L442 bonds.
- An additional cleavage site was identified 13 amino acids C-terminal to the N341-F342 site.
- Sequence analysis revealed the N-terminus of the G2 product as T355VxxPDVELPLP, indicating cleavage at Q354-T355.
Conclusions:
- MT1-MMP exhibits specific cleavage activity within the aggrecan interglobular domain.
- Three distinct cleavage sites (N341-F342, D441-L442, and Q354-T355) were confirmed.
- Understanding these cleavage patterns is crucial for studying aggrecan metabolism and cartilage degradation.