Rapid Diversification of BetI-Based Transcriptional Switches for the Control of Biosynthetic Pathways and Genetic Circuits

ACS Synth Biol. 2016 Nov 18;5(11):1201-1210. doi: 10.1021/acssynbio.5b00230. Epub 2016 Mar 29.

Abstract

Synthetic biologists are in need of genetic switches, or inducible sensor/promoter systems, that can be reliably integrated in multiple contexts. Using a liquid-based selection method, we systematically engineered the choline-inducible transcription factor BetI, yielding various choline-inducible and choline-repressive promoter systems with various input-output characteristics. In addition to having high stringency and a high maximum induction level, they underwent a graded and single-peaked response to choline. Taking advantage of these features, we demonstrated the utility of these systems for controlling the carotenoid biosynthetic pathway and for constructing two-input logic gates. Additionally, we demonstrated the rapidity, throughput, robustness, and cost-effectiveness of our selection method, which facilitates the conversion of natural genetic controlling systems into systems that are designed for various synthetic biology applications.

Keywords: Boolean logic gates; carotenoid; directed evolution; genetic switch; induction system; liquid-handling; lycopene; operon; selection.

MeSH terms

  • Bacterial Proteins / genetics*
  • Bacterial Proteins / metabolism
  • Biosynthetic Pathways
  • Choline / chemistry
  • Escherichia coli / genetics
  • Gene Regulatory Networks*
  • Promoter Regions, Genetic
  • Synthetic Biology / methods*
  • Transcription Factors / genetics*
  • Transcription Factors / metabolism

Substances

  • Bacterial Proteins
  • Transcription Factors
  • Choline