Comparison of CO-dependent H production with strong promoters in Thermococcus onnurineus NA1.

Lee, Seong; Kim, Min-Sik; Bae, Seung; Choi, Ae; Lee, Jin-Won; Kim, Tae; Lee, Jung-Hyun; Lee, Hyun; Kang, Sung

To overproduce biotechnologically valuable products, the expression level of target genes has been modulated by using strong promoters. In a hyperthermophilic archaeon Thermococcus onnurineus NA1, two promoters, P and P, which drive expression of the genes encoding the S-layer protein and glutamate dehydrogenase were inserted in front of a gene cluster encoding a carbon monoxide dehydrogenase, a hydrogenase and a Na/H antiporter. Two promoters exhibited strong activity by increasing the transcription and translation levels of the gene cluster in the mutant strains by 2.5- to 49-folds and 1.4- to 3.3-folds, respectively, than the native promoter in the wild-type strain. While KS0413 with P promoter exhibited 2.7 to 4.7 times higher transcript level than KS0157 with P promoter, the levels of proteins were a little different between them. The biomass concentrations and H production rates of two mutants were 2- to 3-fold higher than those of the wild-type strain in a bioreactor where CO was supplied at a flow rate of 120 ml min. Two mutants showed differential response to the higher CO flow rate, 240 ml min, in terms of growth pattern and product formation, indicating two promoters were regulated by culture conditions. The results demonstrate that not only promoter strength but also product-forming conditions should be considered in promoter engineering.

CARBON monoxide; HYDROGEN production; PROMOTERS (Genetics); THERMOPHILIC microorganisms; GENETIC code; BIOMASS

Applied Microbiology & Biotechnology, 2014, Vol 98, Issue 2, p979

0175-7598

Academic Journal

10.1007/s00253-013-5448-y