Repressor-Like On-Off Regulation of Protein Expression by the DNA-Binding Transcription Activator-Like Effector in T7 Promoter-Based Cell-Free Protein Synthesis

Masafumi Sakono*, Ryoto Hayakawa

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

4 Scopus citations

Abstract

The aim of this study was to develop a transcription activator-like effector (TALE)-based technology to regulate protein synthesis in cell-free systems. We attempted to regulate the T7 promoter system, which has no natural mechanism of expression control, and sought to arbitrarily induce protein expression through the formation and dissociation of TALE and target DNA complexes. Protein synthesis was performed in a cell-free system in the presence of TALE, which recognized and bound to a sequence upstream of the T7 promoter, and protein expression was suppressed by approximately 80 % compared to in the absence of TALE. This suggests that masking part of the promoter region strongly suppresses protein synthesis. Additionally, competitive inhibition of TALE binding to the target DNA template led to protein synthesis levels that were equivalent to the levels in the absence of TALE. Our results demonstrate that DNA recognition by TALE can regulate the expression of the T7 promoter system.

Original languageEnglish
Pages (from-to)888-893
Number of pages6
JournalChemBioChem
Volume22
Issue number5
DOIs
StatePublished - 2021/03/02

Keywords

  • DNA-binding protein
  • T7 promoter system
  • cell-free protein synthesis
  • transcription activator-like effector

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Medicine
  • Molecular Biology
  • Organic Chemistry

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