Biology Reference
In-Depth Information
Q
β
Replicase
β
-Subunit rna
Purified translation factors
Q
β
Replicase
β
-subunit protein
294
Liposome membrane
FIGURE 15.5D
Illustration of a basic self-encoding system: RNA for the β-subunit of the Qβ replicase as well as purified translation factors
are compartmentalized into a liposome. The β-subunit of the replicase is then translated into protein by the purified
translation system, following which it forms the hetero-tetramer Qβ replicase composed of the β-subunit as well as EF-Tu,
EF-Ts, and ribosomal protein S1 which are present in the purified translation system. The reconstituted Qβ replicase can
then replicate the RNA template, leading to a basic self-encoding system whereby the template RNA is replicated by the
protein it encodes.
level to the 100-liter scale.
2,5
In addition to analogous product protein yields, the
production rates at the
L, mL, and L scale were virtually identical.
2
Although scalability
over six orders of magnitude to the 100 L scale has been demonstrated, increasing to the
1000 L or even 10 000 L scale would enhance the attractiveness of cell-free biology for
commercial applications. Moreover, the development of a scalable eukaryotic system is
needed.
μ
Development of Complex Biocatalytic Networks
The cell-free system provides an environment where the expression rate of multiple products
can be controlled in parallel to assess the impact of coproduction on an individual
product
s activity. This is particularly important when assembling complex macromolecular
assemblies such as the ribosome or virus-like particles. As recently demonstrated, small
changes in expression levels of one protein of multiprotein macromolecular complex can
significantly impact the efficiency and activity of the assembled macromolecular complex.
25
In addition, researchers have noted the engineering advantages of employing in vitro
'
