title: Heterologous expression and functional characterization of matrix metalloproteinase-11 from canine mammary tumor.
abstract: Matrix metalloproteinases (MMPs) are reported to be involved in tumor growth, apoptosis, angiogenesis, invasion, and development of metastases. These are zinc containing metalloproteases, known for their role in extracellular matrix degradation. MMP-11 (stromelysin3) is reported to be highly expressed in breast cancer, therefore it may act as marker enzyme for breast cancer progression. The present work was carried out to produce recombinant canine (Canis lupus familiaris) MMP-11 lacking the signal and propeptide in E. coli by optimizing its expression and purification in biologically active form and to functionally characterize it. A bacterial protein expression vector pPROEX HTc was used. The MMP-11 mature peptide encoding gene was successfully cloned and expressed in E. coli and the purified recombinant enzyme was found to be functionally active. The recombinant enzyme exhibited caseinolytic activity and could be activated by Trypsin and 4-Amino phenyl mercuric acetate (APMA). However Ethylene diamine tertra acetate (EDTA) inhibited the enzyme's caseinolytic activity. The recombinant enzyme degraded extracellular matrix constituents and facilitated migration of MDCK (Madin-Darby canine kidney) cells through BD Biocoat Matrigel invasion chambers. These results suggest that in vivo MMP-11 could play a significant role in the turnover of extracellular matrix constituents.
authors: IVRI, Biochemistry, IVRI, Izatnagar, Bareilly, India.
authors: B V
authors: BV
authors: Sunil Kumar
authors: K Aswani
authors: KA
authors: Kumar
authors: K
authors: K
authors: Padmanath
authors: Bhaskar
authors: B
authors: Sharma
authors: Meena
authors: M
authors: Kataria
mesh_terms: D000818:Animals; D015153:Blotting, Western; D003001:Cloning, Molecular; D003584:Cytological Techniques; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D004273:DNA, Neoplasm / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D004285:Dogs / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005260:Female; D061985:Madin Darby Canine Kidney Cells; D015674:Mammary Neoplasms, Animal / Q000737:chemistry / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism; D053507:Matrix Metalloproteinase 11 / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
chemical_list: D018076:DNA, Complementary; D004273:DNA, Neoplasm; D011994:Recombinant Proteins; D053507:Matrix Metalloproteinase 11