title: The efficient expression of human fibroblast collagenase in Escherichia coli and the discovery of flavonoid inhibitors.
abstract: Human skin fibroblast collagenase also known as Matrix Metalloproteinase-1 (MMP-1) is a key enzyme in remodeling and degradation of extracellular matrix, and the inhibitors of human MMP-1 are effective drug candidates for the treatment of cancer. In this study, we report an improved method for high-level expression of soluble human MMP-1 catalytic domain (cd-MMP-1) in E.coli. The enzymatic activity is found maximum at pH 7.5 and temperature 40°C with a Km value of 13.02 µM. Effects of 17 structure-related flavonoids on MMP-1 activity are evaluated using a fluorescent assay, 6 inhibitors are identified with IC₅₀ < 10 µM. Fisetin is the most active agent with an IC₅₀ value of 1.35 µM and is identified as a mixed type inhibitor. Our improved soluble cd-MMP-1 expression method provides a basis for inhibitors identification and may be beneficial to discover novel anti-cancer agent targeting human MMP-1.
authors.affiliation: Shanghai Key Laboratory of Chemical Biology, School of Pharmacy, East China University of Science and Technology, Shanghai, China.
authors.forename: Weiqiang
authors.initials: W
authors.lastname: Lu
authors.forename: Junsheng
authors.initials: J
authors.lastname: Zhu
authors.forename: Shien
authors.initials: S
authors.lastname: Zou
authors.forename: Xi
authors.initials: X
authors.lastname: Li
authors.forename: Jin
authors.initials: J
authors.lastname: Huang
mesh_terms: D003001:Cloning, Molecular; D004305:Dose-Response Relationship, Drug; D055808:Drug Discovery; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005419:Flavonoids / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects* / Q000235:genetics; D006801:Humans; D020784:Matrix Metalloproteinase 8 / Q000235:genetics / Q000378:metabolism*; D015394:Molecular Structure; D013329:Structure-Activity Relationship
chemical_list: D004791:Enzyme Inhibitors; D005419:Flavonoids; D020784:Matrix Metalloproteinase 8