Alkaline protease production by thermotolerant Bacillus sp. KU-K2, from non-rubber skim latex through the non-sterile system and its enzymatic characterization

Abstract

The non-rubber skim latex was used as a substrate for alkaline protease production under a non-sterile system. Among 108 isolates of aerobic spore-forming bacteria, the isolate of KU-K2 was selected as a potent bacterial strain for alkaline protease production. The 16s rRNA gene sequence analysis showed that the KU-K2 strain was closely related to the B. licheniformis with 99.39% similarity. The alkaline protease production from non-rubber skim latex serum was optimized using the Plackett-Burman design (PB) and response surface methodology based on central composite design (CCD). The results showed that the optimized medium consisted of 9.82 g/L skim milk, 0.97 g/L glucose, 0.82 g/L K2HPO4, and 0.02 g/L CoCl2·6H2O gave the highest enzyme production at 6.14 ± 0.13 U/mL. The upscale production in the 5.0 L stirrer fermenter was achieved at 6.78 ± 0.32 U/mL when incubated at 37 °C, 250 rpm with 2.0 vvm of aeration rate for 48 h, which increased by 2.90 folds compared to the un-optimized medium. The optimal pH and temperature for alkaline protease activity were 10.0 and 60 °C, respectively. Zn2+, Fe3+, and Hg2+ metal ions strongly inhibited the crude alkaline protease. This study showed the potential of thermotolerant Bacillus sp. For alkaline protease production using non-rubber skim latex as a substrate which could be applied to reduce the industrial waste from latex industries.