3. Strategies to modulate expression using synthetic biology
Synthetic biology encompasses a broad curation of methods aimed at
reprogramming different cell types including microbial and mammalian
cells for the purposes of producing compounds used in biofuels,
pharmaceuticals, and materials-based applications. The synthetic
processes necessary to produce these compounds requires the introduction
of metabolic pathways not native to the original host species in many
cases. One challenge synthetic biology has the potential to address is
producing high-quality MPs for structural characterization and other
biochemical analyses. Progress in MP research is often stymied by a lack
of high-throughput, robust methods to produce high-quality material. The
following summarizes and highlights some of the most effective tools
available that can be used to tune MP expression and production inS. cerevisiae , Much of the progress made in synthetic biology has
been possible using E. coli and S. cerevisiae – two of
the best studied microbes. Application of these tools is slowly being
introduced into other microbes. Processes in yeast will be the focus for
the remainder of the review. The intracellular processes involved in
protein production include transcriptional regulation through adaptation
of genetic elements such as gene promoters, terminator regions, and
other mechanisms of transcriptional control including transcription
factors. A brief discussion on the use of fusion tags to boost protein
yields follows along with whole genome editing strategies to generate
selective knockout (KO) strains and down-regulate undesired proteolytic
pathways. Conversely, metabolic engineering methods are discussed as a
way of introducing novel pathways and genes identified to support MP
production along with future insights into how this technology can be
used to tune processes involved in expression. Innovative culturing
methods that include comparative insights into differential protein
induction methods, constitutive versus inductive, are discussed. Lastly,
the latest developments in cell-free (CF) expression, a field that has
seen significant advances in protein production applications, is
presented with an emphasis on advantages over whole cell expression and
future directions.