181Synthetic Biology: Parts, Devices and Applications, First Edition. Edited by Christina Smolke.
© 2018 Wiley-VCH Verlag GmbH & Co. KGaA. Published 2018 by Wiley-VCH Verlag GmbH & Co. KGaA.
9
9.1 Introduction Yohei Yokobayashi
Engineering small molecule‐responsive RNA switches in bacteria was motivated
by the discovery of natural prokaryotic riboswitches in 2002 [1–3]. These
en dogenous RNA cis‐regulatory elements are usually found in the 5′ untrans-
lated region (UTR) of prokaryotic mRNAs and modulate gene expression in
response to various metabolites [4]. A typical riboswitch contains an aptamer
domain that is responsible for metabolite binding and an expression platform
that facilitates a ligand‐dependent structural change that influences gene
expression. For example, a metabolite‐mediated structural change may alter the
accessibility of the ribosome binding site (RBS), which results in a change in
translation efficiency, or dictate the formation of a transcription terminator
structure (a stem‐loop followed by a short poly(U) tract) that results in prema-
ture termination of the transcript. The ability of these riboswitches to control
gene expression in response to small molecules of biological or synthetic origin
can be very useful in synthetic biology and metabolic engineering. This section
provides a brief overview of the previous major efforts to engineer small
molecule‐ responsive RNA switches in bacteria.
9.2 Design Strategies
9.2.1 Aptamers
An RNA aptamer that specifically binds to a desired small molecule ligand is a
prerequisite to engineering riboswitches. Most published synthetic riboswitches
have used known aptamers selected in vitro or metabolite‐binding aptamers
found in natural riboswitches. At least one group has performed in vitro selection
to develop novel aptamers specifically for riboswitch applications in bacteria [5].
While researchers have successfully performed in vitro selection to discover
Small Molecule-Responsive RNA Switches (Bacteria):
Important Element of Programming Gene Expression
in Response to Environmental Signals in Bacteria
Yohei Yokobayashi
Okinawa Institute of Science and Technology Graduate University, Nucleic Acid Chemistry and Engineering Unit
Onna-son, Okinawa 9040415, Japan