Description
Description
MW = 18.0 kDa. Recombinant matrix metalloproteinase-3 (MMP-3, stromelysin-1, transin) cloned from human cDNA, expressed in E.coli. The enzyme consists of the catalytic domain of human MMP-3, residues 105-265 (UniProtKB accession P08254) with the mutation F171D. The protein has been mutated to increase its stability, as the mutation drastically reduces the enzyme’s rate of autoproteolysis. The catalytic activity rates are not affected by the mutation.
Sequence
100 110 120
M-F RTFPGIPKWR KTHLTYRIVN
130 140 150 160 170 180
YTPDLPKDAV DSAVEKALKV WEEVTPLTFS RLYEGEADIM ISFAVREHGD DYPFDGPGNV
190 200 210 220 230 240
LAHAYAPGPG INGDAHFDDD EQWTKDTTGT NLFLVAAHEI GHSLGLFHSA NTEALMYPLY
250 260 265
HSLTDLTRFR LSQDDINGIQ SLYGP
Purity
> 95% by SDS-PAGE. The protein is observed, in denaturing conditions, as a single band migrating at a molecular weight between 18.4 and 25.0 kDa.
Suppied As
0.2 mg/mL solution in Tris 20 mM, pH 7.2, CaCl2 10 mM, ZnCl2 0.1 mM, NaCl 0.3 M, acetohydroxamic acid (AHA) 0.2 M. The concentration is calculated by the analysis of the absorbance at 280 nm (ε280 = 28420 M-1cm-1 calculated).
Specific activity
> 30 U/μg. Activity described as U=100 pmol/min at 37°C using a colorimetric assay with thiopeptolide Ac-Pro-Leu-Gly-[2- mercapto-4-methyl-pentanoyl]-Leu-Gly-OC2H5 (Biomol) as substrate.
Storage
-80°C. After initial defrost, aliquot the product into individual tubes and refreeze at -80°C.
Avoid repeated freeze/thaw cycles.
Usage
Enzyme kinetic studies, cleavage of target substrates and screening of inhibitors.
RELEATED RESEARCH FIELDS
References
Johnson, L.L. et al. J. Biol. Chem. 275 (15), 11026-11033 (2000).
Chen, L. et al. J. Mol. Biol. 293 (3), 545-557 (1999).
Weingarten, H. & Feder, J. Anal. Biochem. 147 (2), 437-440 (1985).
Weingarten, H., Martin, R. & Feder, J. Biochemistry 24 (23), 6730-6734 (1985).
