A1 Journal article – refereed
GreA and GreB Enhance Expression of Escherichia coli RNA Polymerase Promoters in a Reconstituted Transcription–Translation System




List of Authors: Lea L. de Maddalena, Henrike Niederholtmeyer, Matti Turtola, Zoe N. Swank, Georgiy A. Belogurov, and Sebastian J. Maerkl
Publisher: American Chemical Society
Publication year: 2016
Journal: ACS Synthetic Biology
Journal name in source: ACS SYNTHETIC BIOLOGY
Journal acronym: ACS SYNTH BIOL
Volume number: 5
Issue number: 9
eISSN: 2161-5063

Abstract

Cell-free environments are becoming viable alternatives for implementing
biological networks in synthetic biology. The reconstituted cell-free
expression system (PURE) allows characterization of genetic networks
under defined conditions but its applicability to native bacterial
promoters and endogenous genetic networks is limited due to the poor
transcription rate of Escherichia coli RNA polymerase in this
minimal system. We found that addition of transcription elongation
factors GreA and GreB to the PURE system increased transcription rates
of E. coli RNA polymerase from sigma factor 70 promoters up to
6-fold and enhanced the performance of a genetic network. Furthermore,
we reconstituted activation of natural E. coli promoters
controlling flagella biosynthesis by the transcriptional activator FlhDC
and sigma factor 28. Addition of GreA/GreB to the PURE system allows
efficient expression from natural and synthetic E. coli promoters
and characterization of their regulation in minimal and defined
reaction conditions, making the PURE system more broadly applicable to
study genetic networks and bottom-up synthetic biology.


Last updated on 2019-20-07 at 13:23