Cloning and Expression of a Thermostable Pullulanase Gene from Thermotoga maritima MSB8 in Bacillus subtilis WB600

The pulA gene enconding a thermostable pullulanase of 844 amino acids was cloned by PCR amplification from the genomic DNA of Thermotoga maritima MSB8. We successfully expressed the pulA gene in a protease-deficient strain Bacillus subtilis WB600, and the expressed pullulanase was secreted into the culture broth. Preliminary results showed that the pullulanase exhibited optimal activity at 90°C, and was stable at the temperature range from 30 to 80°C for 30 min. The stable pH range for this pullulanase was from 5.5 to 7.0, with an optimum at pH 6.0.