Bacillus amyloliquefaciens HZ-12 heterologously expressing NdmABCDE with higher ability of caffeine degradation

Caffeine is a major component of tea and coffee. Apart from the stimulatory effects, cumulative intake of caffeine has caused several diseases. In addition, the presence of caffeine brings out the deterioration of environmental pollution. Therefore, the demand for decaffeinated products, especially decaffeinated coffee and tea, is increasing. However, Pseudomonas putida as the most widely studied bacteria isn't suitable for producing decaffeinated tea and coffee due to the production of an unpleasant odor and endotoxins at the same time. A more appropriate system (Bacillus amyloliquefaciens HZ-12:ndmABCDE) is needed to produce decaffeinated products; and HZ-12:ndmABCDE was constructed by orderly integrating N-demethylase genes (ndmABCDE) in a safer strain HZ-12. Demethylases (NdmABCDE) in P. putida can degrade caffeine to xanthine, which was further degraded to CO2 and NH4+ by the purine metabolic pathway. Finally, HZ-12:ndmABCDE could degrade 52.6% of caffeine in the optimized medium, 31.5% higher than HZ-12; caffeine was demethylated in sequence to form theobromine, paraxanthine, 7-methylxanthine and xanthine; and there are no endotoxins and unpleasant odor. This study provides a safer strain (HZ-12:ndmABCDE) for producing decaffeinated tea and coffee and a new idea that B. amyloliquefaciens HZ-12 can heterologously express the demethylase genes in P. putida.