Cloning and expression of the levanase gene in Alcaligenes eutrophus H16 enables the strain to hydrolyze sucrose

Abstract

Genetic engineering methods were used to enhance the substrate spectrum of Alcaligenes eutrophus H16, a poly-β-hydroxybutyric acid (PHB) producer. Using parts of the vector pMMB33 and a 2.5 kb DNA fragment of the Bacillus subtilis chromosome a plasmid was constructed bearing the gene for levanase, an enzyme able to hydrolyze various saccharides. After transfer of the levanase gene by triparental conjugation, the gene, controlled by its own Bacillus subtilis promoter, is expressed in Alcaligenes eutrophus H16 and enables the strain to hydrolyze sucrose. However, growth on sucrose is limited; i.e. the sucrose is not transported efficiently into the cell and/or the levanase is not secreted into the medium.