Discovery and Characterization of Native Deinococcus radiodurans Promoters for Tunable Gene Expression

摘要

The potential utilization of extremophiles as a robust chassis for metabolic engineering applications has prompted interest in the use of Deinococcus radiodurans for bioremediation efforts, but current applications are limited by the lack of availability of genetic tools, such as promoters. In this study, we used a combined computational and experimental approach to identify and screen 30 predicted promoters for expression in D. radiodurans using a fluorescent reporter assay. The top eight candidates were further characterized, compared to currently available promoters, and optimized for engineering through minimization for use in D. radiodurans. Of these top eight, two promoter regions, PDR_1261 and PrpmB, were stronger and more consistent than the most widely used promoter sequence in D. radiodurans, PgroES. Furthermore, half of the top eight promoters could be minimized by at least 20 (to obtain final sequences that are approximately 24 to 177 bp), and several of the putative promoters either showed activity in Escherichia coli or were D. radiodurans specific, broadening the use of the promoters for various applications. Overall, this work introduces a suite of novel, well-characterized promoters for protein production and metabolic engineering in D. radiodurans.