Abstract

The extracellular collagenase, produced by Bacillus licheniformis F11.4, was purified by ammonium sulfate precipitation followed by DEAE Sephadex A-50. The purified collagenase showed a 26.3-fold increase in specific activity being 1.0 U/mg and 2.6% recovery. The collagenase has an apparent molecular weight of 124 and 26 kD as analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and zymography. The optimal temperature and pH were 50°C and pH 7.0, respectively. The collagenase activity was inhibited by Fe²⁺ (1 mM), Mg²⁺ (1 mM), Mn²⁺ (1 mM), Co²⁺ (1 mM), EDTA (1 mM), and β-mercaptoetanol (1 mM). However, Ca²⁺ (1 mM) and Cu²⁺ (1 mM) increased its activity. The collagenase from B. licheniformis F11.4 was capable of hydrolyzing other protein substrates such as casein, gelatin, and fibrin. The K_m and V_max of the enzyme for collagen were 0.26 mg/ml and 0.27 U, respectively.

Key words: Collagenase, Bacillus licheniformis F11.4, purification, characterization.