Cloning, characterization and application of the promoter of alkaline protease gene in Bacillus pumilus

Abstract

A 797 bp promoter fragment of alkaline protease gene was cloned from the Bacillus pumilus genome by employing TAIL-PCR strategy. The sequence analysis of this promoter fragment showed that the sequence accounting for gene expression was approximately 390bp. Deletion analysis of the fragment defined the minimal required sequence of promoter for initiating transcription lies on a 160 bp region upstream of the start codon. The alkaline protease gene WApQ3 containing the cloned promoter fragment was inserted into the shuttle vector pSUGV4 and the constructed expression plasmid pSUBpWApQ3 was transformed into Bacillus subtilis and B. pumilus. Active alkaline protease was successfully expressed in both host strains. The peak value of extracellular alkaline protease activities of B. subtilis and B. pumilus recombinants reached to 465.5 U/mL and 3060 U/mL, respectively.