(175h) Refactoring the Partial Photosynthesis Gene Cluster for the Biosynthesis of the Key Pigment, Bacteriochlorophyll a

In
prokaryotes, the genes responsible for complex biological functions are often
physically clustered on the chromosome for co-regulation. As a goal for
synthetic biology, it is important to design and build manipulable gene
clusters for the further construction of biological systems with higher
hierarchy and useful biological functions. In order to achieve this goal,
microbial photosynthesis machinery was chosen as a target for refactoring.
Taking the purple non-sulfur bacterium Rhodobacter sphaeroides 2.4.1 as
an example, almost all the genes responsible for photosynthesis are clustered
on the chromosome spanning in an approximately 40 kb region. We are using a
systematic approach to re-design this photosynthesis gene cluster by rebuilding
it from the bottom-up using synthetic and well-characterized genetic parts.
Focusing on the biosynthetic operons for the key pigment bacteriochlorophyll a (BChl a)
biosynthesis, all the essential genes will be chosen for knock-out and
complementation, with the exception of regulatory genes. An architecture with mono-cistronic expression
of each gene will be adopted for the construction of the synthetic gene cluster. Each gene
is being designed to be under the control of tunable genetic parts, including
promoter, ribosome binding site, transcription terminator and insulator. The
codons for the genes are adjusted to be as different as possible from the wild
type genes, thus excluding possible internal regulation sequences. Finally, a simplified
environment sensor will be implemented in a genetic circuit to regulate the
gene expression profile. The synthetic gene cluster for the biosynthesis
of BChl a may
provide a modular platform for further combinatorial biosynthesis of BChl a derivatives, which may expand the
chemical space for additional and novel photosynthetic pigments. Via this
refactoring process, this gene cluster may also be easily transferred to other
non-photosynthetic microorganisms, facilitating the engineering of artificial
photosystems.