Cloning，expression and characterization of alcohol dehydrogenase and aldehyde dehydrogenase from Bacillus pseudofirmus OF4

Abstract:

Abstract:［Objective］ To clone and characterize the alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) from Bacillus pseudofirmus OF4． ［Methods］Genes of adh and aldh were cloned by PCR; expression vectors pET-Ahd and pET-Aldh were constructed and expressed in Escherichia coli BL21 (DE3)．After Ni-NTA column chromatography purification，the protein was characterized．［Results］The optimal temperature and pH of ALDH was 35℃ and 8. 0，the specific activities of ALDH was 979.6 U/mg protein，the thermostability at 25℃ and 35℃ was better than at 45℃ ． Although the expression level of ADH was too low to purify，but it was found that ADH had high catalytic activities by experiments of co-expression and ethanol tolerance．［Conclusion］Adh and aldh from B． pseudofirmus OF4 were cloned successfully． Co-expression of double genes could greatly increase the host strain on ethanol tolerance.