Biology Reference
In-Depth Information
sequences constructed from operator sites, or parts, of well-characterized naturally occurring
E. coli
promoters and their complementary regulatory proteins. Over the last decade many
additional systems have been designed and built based on a similar methodology.
16,23
35
For example, one family of systems, the AND gates, is a set of synthetic promoters
composed of multiple operator sites. The sites are organized in such a way that the resulting
promoters follow AND logic in response to inducer molecules.
16
Although our understanding of synthetic biological systems is growing, how to optimally
construct and tune them still remains challenging. A major challenge that researchers face
throughout the development of new synthetic systems is finding which parts are most
suitable for constructing a particular system with a specific, targeted phenotypic behavior.
36
The complexity of synthetic constructs may quickly overwhelm the intuition of even the
most seasoned of practitioners.
Mathematical modeling has been widely used in explaining the behavior of various
synthetic systems, and can contribute significantly to their full characterization. As the
interactions and dynamics governing the biological systems are significantly nonlinear, and
therefore arduous to be predicted experimentally, building such systems benefits from
modeling. Mathematical modeling of synthetic biological systems has therefore become a
necessary ally of experimental work.
37
41
Remarkable examples of synthetic systems that
have been computationally described include protein production in an oscillatory or
bistable manner,
21,22,42,43
tight regulation of gene expression,
15,44
and logical gate-like
behavior.
16,45
Importantly, we are now transitioning from relatively simple biological
constructs to complex synthetic biological systems which integrate several simple systems
and are capable of robustly performing multiple user-defined functions.
46
These functions
have also been captured using mathematical modeling and include, but are not limited to,
cells that efficiently synchronize with one another
47
and cells that kill or rescue each
other.
48
50
Synthetic biology technologies resulting from these systems have culminated in
important applications in producing biofuels with high efficiency, in targeting and
destroying cancer cells, in preventing and treating infections, and in developing inexpensive
vaccines.
51
53
124
To date, a gamut of software products have been developed that allow synthetic biologists
to model the behavior of biological systems of interest. These packages include, but are not
limited to, SynBioSS,
54,55
CellDesigner,
56
GenNetDes,
57
COPASI,
58
TinkerCell,
59
and
others.
60,61
Each of the aforementioned products has its own limitations and advantages. In
what follows, we discuss the Synthetic Biology Software Suite (SynBioSS) and its various
components. We first revisit two sets of experimentally constructed synthetic biological
switches to illustrate the usefulness of SynBioSS and its significance in designing and
characterizing synthetic biological systems. The first is a set of AND gates and the second is
a set of inducible activators. Combined, these systems offer a wide spectrum of regulating
devices: a repressor; a derepressor; an activator; and a deactivator. All these result in varying,
well-characterized protein expression amounts.
SYNTHETIC LOGICAL-AND GATES AND PROTEIN DEVICES
AND Gates
The AND gate systems were built using components of the lactose and tetracycline operons,
and then tested both experimentally and computationally.
16
Inducible synthetic promoters
were constructed by placing the operator sites
lacO
and
tetO
downstream, between and
upstream of the
E. coli
10 consensus sequences. Three operator sites were
included for each promoter, one
lacO
and two
tetO
, or two
lacO
and one
tetO
. A strong
ribosomal binding site (RBS) and a transcriptional start site were included to complete the
functional promoter. This construction, as illustrated in
Figure 7.1
, provides the AND gate
35 and
2
2
