Purification, characterization, and gene cloning of a cold-adapted thermolysin-like protease from Halobacillus sp. SCSIO 20089.
Marine sediment is a distinctive habitat of cold enzyme producing bacteria. A protease producing strain Halobacillus sp. SCSIO 20089 was isolated from a marine sediment of South China Sea. Using chromatographic techniques, the extracellular protease was purified to homogeneity from the culture supernatant. The purified protease exhibited maximal activity at 30°C, pH 8.0, and remained more than 20% of its activity at 0°C. Its activation energy was calculated to be 34.4 kJ/mol, suggesting it is a cold-adapted protease. Based on the N-terminal amino acid sequence of the purified enzyme, full gene encoding the enzyme was obtained by combination of degenerate primer PCR and hiTAIL-PCR. The deduced amino acid sequence showed 57% and 52% identity with mesothermal and thermophilic protease in thermolysin family respectively. All these indicate the enzyme is a unique cold-active thermolysin-like protease with potential in both basic research and industrial application areas.
Key Laboratory of Marine Bio-resources Sustainable Utilization, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, People's Republic of China.
Jian
J
Yang
Jie
J
Li
Zhimao
Z
Mai
Xinpeng
X
Tian
Si
S
Zhang
D000220:Adaptation, Biological; D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D003080:Cold Temperature; D058678:Halobacillus / Q000145:classification / Q000201:enzymology*; D008666:Metalloendopeptidases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D013379:Substrate Specificity
D008666:Metalloendopeptidases; C044194:microbial metalloproteinases