Directed evolution of a transcription factor PbrR to improve lead selectivity and reduce zinc interference through dual selection.

ABSTRACT: Directed evolution has been proven as a powerful tool for developing proteins and strains with novel or enhanced features. In this study, a dual selection system was designed to tune the binding specificity of a transcription factor to a particular ligand with the ampicillin resistance gene amp (ON selection) as the positive selection marker and the levansucrase gene sacB (OFF selection) as the negative selection marker. It was applied to the lead responsive transcription factor PbrR in a whole-cell lead biosensor previously constructed in our lab (Jia et al. in Fems Microbiol Lett 365:fny157, 2018). After multiple rounds of ON-OFF selection, two mutants with higher specificity for lead were selected. Structural analysis revealed that the mutation C134 located on the metal-binding loop at

SUBMITTER: Jia X 

PROVIDER: S-EPMC7148400 | biostudies-literature | 2020 Apr

REPOSITORIES: biostudies-literature