Attenuated colicin-based screening to discover and create novel resistance genes

I formulated a systematic approach to screen for colicin resistance-associated genes by using an attenuated colicin mutant and demonstrated its utility in a screen of genes related to colicin E5 resistance. Screening of an Escherichia coli genome library revealed rstA as a partial resistance gene to colicin E5. Transcript expression of BtuB and OmpF, proteins responsible for translocation of nuclease E colicins, was clearly inhibited in an rstA-overexpressing strain. In addition, the tatA::recN fusion gene provides resistance to an attenuated colicin E5 mutant. Improving tatA::recN by directed coevolution, I created a novel gene with enhanced resistance to colicin E5 and concluded that attenuated colicin-based screening is useful for the discovery and creation of novel colicin resistance-associated genes.