Optimization of the purine operon and energy generation in Bacillus amyloliquefaciens for guanosine production.

ABSTRACT

OBJECTIVES: To deregulate the purine operon of the purine biosynthetic pathway and optimize energy generation of the respiratory chain to improve the yield of guanosine in Bacillus amyloliquefaciens XH7. RESULTS: The 5'-untranslated region of the purine operon, which contains the guanine-sensing riboswitch, was disrupted. The native promoter Pw in B. amyloliquefaciens XH7 was replaced by different strong promoters. Among the promoter replacement mutants, XH7purEP41 gave the highest guanosine yield (16.3 g/l), with an increase of 23% compared with B. amyloliquefaciens XH7. The relative expression levels of the purine operon genes (purE, purF, and purD) in the XH7purEP41 mutant were upregulated. The concentration of inosine monophosphate (IMP), the primary intermediate in the purine pathway, was also significantly increased in the XH7purEP41 mutant. Combined modification of the low-coupling branched respiratory chains (cytochrome bd oxidase) improved guanosine production synergistically. The final guanosine yield in the XH7purEP41△cyd mutant increased by 41% to 19 g/l compared with B. amyloliquefaciens XH7. CONCLUSION: The combined modification strategy used in this study is a novel approach to improve the production of guanosine in industrial bacterial strains.