Molecular characteristics of equine stromelysin and the tissue inhibitor of metalloproteinase 1.
Abstract: To clone the entire coding sequence of equine matrix metalloproteinase-3 (MMP-3, stromelysin) and tissue inhibitor of metalloproteinase-1 (TIMP-1) and compare their nucleotide and amino acid sequences with those of MMP-3 and TIMP-1 from other species. Methods: Articular cartilage harvested from the joints of 4 foals, 2 yearlings, and 3 adult horses. Methods: A cDNA library was constructed from mRNA extracted from equine chondrocytes. The library was screened and clones selected that contained the cDNA for MMP-3 and TIMP-1. The cDNA was sequenced and the nucleotide and deduced amino acid sequences compared with known sequences in other species. Northern blot analysis was performed, using the resulting cDNA clones. Results: An 1803-bp cDNA for MMP-3 including the entire coding sequence of 1434 bases was cloned and sequenced. A 744-bp cDNA for TIMP-1 including the entire coding sequence of 624 bases was cloned and sequenced. Northern analysis revealed MMP-3 to hybridize to a single mRNA species at approximately 2.1 kb. TIMP-1 hybridized to a single mRNA species at approximately 0.8 kb. Conclusions: MMP-3 and TIMP-1 were highly homologous to that of other species at the nucleotide and amino acid level although each had unique residues in part of the peptide that is generally conserved. Conclusions: Understanding the molecular structure of MMP-3 and TIMP-1 and the availability of their cDNA should allow a more detailed understanding of their balance in cartilage and the degradative processes in joint disease.
Publication Date: 1998-12-19 PubMed ID: 9858406
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- Journal Article
- Research Support
- Non-U.S. Gov't
- Research Support
- U.S. Gov't
- P.H.S.
Summary
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This research is about the cloning, sequencing, and comparison of equine matrix metalloproteinase-3 (MMP-3 – also known as stromelysin) and tissue inhibitor of metalloproteinase-1 (TIMP-1) to understand their role in horse joint health.
Methodology
The researchers followed a procedural approach that included:
- Harvesting articular cartilage from diverse age groups of horses. Four foals, two yearlings, and three adult horses were used in the study.
- Constructing a cDNA library from mRNA extracted from equine chondrocytes, cells from the cartilage.
- Screening the cDNA library and selecting clones containing the cDNA for MMP-3 and TIMP-1.
- Sequencing the cDNA of the selected clones and comparing their nucleotide sequences and inferred amino acid structures with known sequences in other species.
- Carrying out Northern blot analysis for the cloned cDNA. This technique is used to observe specific RNA molecules among a mixture of other RNA.
Findings
The research yielded some crucial findings through the experimental methods:
- An 1803-bp cDNA for MMP-3 including the entire coding sequence of 1434 bases was cloned and sequenced.
- A 744-bp cDNA for TIMP-1 including the entire coding sequence of 624 bases was cloned and sequenced.
- Through the Northern analysis, MMP-3 and TIMP-1 each hybridized to a unique mRNA species at approximately 2.1 kb and 0.8 kb respectively.
- MMP-3 and TIMP-1 appeared highly alike to other species at the nucleotide and amino acid level, even though each had unique residues at part of the generally conserved peptide.
Conclusions
The significance of these findings can be summarized as follow:
- The process helped in understanding the molecular structure of MMP-3 and TIMP-1, enzymes linked to joint health in horses.
- The availability of their cDNA will allow a more detailed understanding of their balance in cartilage and their role in the degradative processes in joint disease in horses and potentially other species.
Cite This Article
APA
Richardson DW, Dodge GR.
(1998).
Molecular characteristics of equine stromelysin and the tissue inhibitor of metalloproteinase 1.
Am J Vet Res, 59(12), 1557-1562.
Publication
Researcher Affiliations
- Department of Clinical Studies, New Bolton Center, School of Veterinary Medicine, University of Pennsylvania, Kennett Square 19348-1692, USA.
MeSH Terms
- Amino Acid Sequence
- Animals
- Base Sequence
- Cartilage, Articular / enzymology
- Cartilage, Articular / metabolism
- Conserved Sequence
- DNA, Complementary
- Gene Library
- Horses
- Humans
- Matrix Metalloproteinase 3 / biosynthesis
- Matrix Metalloproteinase 3 / chemistry
- Matrix Metalloproteinase 3 / genetics
- Molecular Sequence Data
- RNA, Messenger / genetics
- RNA, Messenger / isolation & purification
- Recombinant Proteins / biosynthesis
- Recombinant Proteins / chemistry
- Sequence Alignment
- Sequence Homology, Amino Acid
- Tissue Inhibitor of Metalloproteinase-1 / biosynthesis
- Tissue Inhibitor of Metalloproteinase-1 / chemistry
- Tissue Inhibitor of Metalloproteinase-1 / genetics
Grant Funding
- AR42417 / NIAMS NIH HHS
Citations
This article has been cited 1 times.- Tung JT, Fenton JI, Arnold C, Alexander L, Yuzbasiyan-Gurkan V, Venta PJ, Peters TL, Orth MW, Richardson DW, Caron JP. Recombinant equine interleukin-1beta induces putative mediators of articular cartilage degradation in equine chondrocytes.. Can J Vet Res 2002 Jan;66(1):19-25.
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