Biology Reference
In-Depth Information
Minimal Cells
Minimal cell construction seeks to assemble the minimal number of cellular components
such as DNA, RNA, and protein encapsulated in a lipid membrane necessary for life.
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The
goals are fundamental knowledge, testing our understanding of life and its origins, and
facilitating engineering for biotechnology applications, including evolutionary optimization
of natural and unnatural biopolymers.
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Minimal cell construction is proceeding via
two approaches: (1) the top-down approach and (2) the bottom-up approach. The top-
down approach attempts to identify the minimal number of genes required for the
organism to live by genome reduction. Pioneering efforts from the J Craig Venter Institute
have made tremendous progress towards experimentally determining essential genes.
Specifically, they identified that 387 out of 482 protein-coding genes of
Mycoplasma
genitalium
and 43 RNA-coding genes were essential for life.
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Further efforts to minimize
this genome have led to the creation of the first synthetic cell controlled by a genome
synthesized from scratch.
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However, this topic chapter is focused on
synthetic biology, and therefore we will
not delve further into the top-down approach, which was reviewed recently by Jewett and
Forster.
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Instead we will focus on the bottom-up approach, which involves creating a
minimal cell by bringing together essential purified biological macromolecules, their genes,
and their small molecule substrates (
Fig. 15.5A
). The main goal of an artificial cell built
from the bottom-up is its ability to self-replicate with efficient removal of byproducts being
exchanged with the outside environment.
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We will focus on attempts to construct a
DNA
cell-free
'
'
protein-based system derived from current biological systems, although it
should be noted that there are efforts and recent advances centered on modeling an RNA
world. In a breakthrough study last year, for example, in vitro evolution was used to
develop an RNA polymerase ribozyme that was capable of synthesizing a hammerhead
ribozyme from an RNA template, demonstrating that RNA can self-replicate and create
functional RNAs in the process.
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RNA
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Key developments in the construction of minimal cells have centered on integrating the
subsystems of biology necessary for self-replication. This includes creating the compartment
(or membrane vesicles), enabling substrates and waste products to flow in and out of
compartments, and activating complex biochemical reactions and networks inside
compartments.
The membrane of the minimal cell must be capable of growth and division, must be
permeable to allow for entry and exit of substrates and waste products, and must be
stable under the conditions of replication and gene expression.
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Conditions and timing of
membrane replication must be ultimately similar to conditions for replication of the
enclosed nucleic acid for the cell to successfully replicate and survive. Core and shell
reproduction (i.e. reproduction of all the components needed to sustain a minimal cell
along with the surrounding lipid vesicle) is currently not possible, but several important
developments have been made, three of which are detailed below. For a more detailed
review of this process refer to Stano et al.
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. In one example, Luisi
s group was able to
construct a synthetic cell that consisted of the reconstituted translation system (PURE)
encapsulated in a liposome that expressed two enzymes/membrane proteins involved in
phospholipid biosynthesis.
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This was the first successful demonstration that membrane
proteins could be synthesized in lipid vesicles of proper lipid composition. Although the
authors initially aimed at observing cell growth and division, low product yield and issues
in the reaction biochemistries did not allow for an observation of morphological changes.
In a second example, researchers were able to chemically link amplification of DNA and
self-reproduction of a giant vesicle, thereby creating a self-reproducing supramolecular giant
(
Fig. 15.5B
).
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While a huge advance for the field, this system has two limitations: (1) the
self-reproduction is limited as the percentage of the phosopholipid in the membrane
'