Selection Of Alkaline Protease Producing Strain Vibrio Metschnikovii DL33-51 And Its Enzymological Properties

An alkaline protease producing strain DL33 is isolated from a soda lake and identified as Vibrio metschnikovii. By combination treatment of DL33 with UV and DBS, a mutant DL33-51 is obtained and its enzyme activity is in the 5.74 fold of the original strain, reaching to 2010U/mL, in 48h shaking flask. The fermentation medium and condition of enzyme production under flask shaking is investigated. Under the obtained optimum condition, the mutant strain give a maximum alkaline protease activity of 3268U/mL in shaking flask after 40h cultivation when growing in a medium containing(g/l) casein, 10; corn meal, 1; NaCl, 14; NaNO3, 10; KH2P04, 1; MgSO4-7H2O, 0.5; CaCl2, 0.1 and compound promoting agent 2.5 (pH 10.4).A novel surfactant- and oxidation-stable alkaline protease from Vibrio metschnikovii DL 33-51 was purified using acetone precipitation, Q-sepharose ion exchange and Sephacryl S-100 gel filtration chromatographic techniques. The enzyme was shown to have a relative molecular weight of 29.5 kDa by SDS polyacrylamide gel electrophoresis. The optimal temperature and pH were found to be 60 癈 and 12.0 respectively. The protease was stable between a pH range of 9.0 and 11.0. The half-lives of protease recorded at 50 and 60 were 7.5 h and 30 min, respectively. The enzyme activity could be strongly activated by several commercially available laboratory surfactants (Triton X-100, tween-80) and bleaches (H2O2, sodium hypochlorite), and showed a valuable stability towards them when their concentrations ranged up to 1.0%. The enzyme activity was increased up to 19% and 18%, respectively, by Mn2+ and Ba2+.The alkaline protease shows compatibility with commercial detergents such as Tide and Jieba. This enzyme improves the cleaning power of commercial detergents, and could remove the blood, milk and protein strains in the cold washing solutions effectively.