Cloning and expression of a novel (−)γ-lactamase gene from Bacillus thuringiensis

Abstract:

[Background] γ-lactam is an important pharmaceutical intermediate and its chiral property makes against drug synthesis. The preparation of optically pure γ-lactam can be realized by γ-lactamases’ selective resolution. [Objective] To explore the gene of γ-lactamase from Bacillus thuringiensis and the technology of optically pure γ-lactam preparation. [Methods] The genome sequence of Bacillus thuringiensis was searched by the key words “acetamidase/formamidase”, two sequences were selected and heterologously expressed. The degradation of γ-lactam by recombinant bacteria was monitored by HPLC method to determine whether the cloned gene was a γ-lactamase gene. The application of 5 L fermentation, substrate separation and product recovery were also performed using the constructed recombinant bacteria. [Results] The skA2 recombinant strain had activity of (?)γ-lactamase and showed good performance in small scale production. [Conclusion] The A gene is an unreported new (?)γ-lactamase gene, which enriches producers’ enzyme toolbox. The skA2 strain can meet the need of industrial production of (+)γ-lactam and lay a solid foundation for the preparation of optically pure drugs.

Key words:γ-lactamase, γ-lactam, gene discovery, chiral separation