MW = 17.6 kDa calculated. Recombinant Matrix Metalloproteinase-8 (MMP-8, Collagenase-2, Neutrophil collagenase) cloned from human cDNA, expressed in E. coli. The enzyme consists of the catalytic domain of human MMP-8 (residues 105-262, UniProtKB accession P22894).
110 120 130 140 NPKWER TNLTYRIRNY TPQLSEAEVE RAIKDAFELW 150 160 170 180 SVASPLIFTR ISQGEADINI AFYQRDHGDN SPFDGPNGIL 190 200 210 220 AHAFQPGQGI GGDAHFDAEE TWTNTSANYN LFLVAAHEFG 230 240 250 260 HSLGLAHSSD PGALMYPNYA FRETSNYSLP QDDIDGIQAI YG
> 95% by SDS-PAGE. The protein is observed, in denaturing conditions, as a single band migrating at a molecular weight between 14.4 and 18.4 kDa.
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) 0.5 M. The concentration is calculated by the analysis of the absorbance at 280 nm (ε280 = 28420 M-1cm-1 calculated).
> 80 U/μg. Activity described as U=100 pmol/min at 25°C using a colorimetric assay with thiopeptide Ac-Pro-Leu-Gly-[2-mercapto-4-methyl-pentanoyl]-Leu-Gly-OC2H5 (Biomol) as substrate.
-80°C. After initial defrost, aliquot the product into individual tubes and refreeze at -80°C.
Avoid repeated freeze/thaw cycles.
Enzyme kinetic studies, cleavage of target substrates and screening of inhibitors.
Betz, M. et al. Eur. J. Biochem. 247 (1), 356-363 (1997).
Murphy, G. & Knäuper, V. Matrix Biol. 15 (8-9), 511-518 (1997).
Ho, T.F. et al. Gene 146 (2), 297-301 (1994).