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approaches like the one presented by Lippow et al., 3 where computation is used to design a
library instead of a single sequence, will become more widely used.
CONCLUSION AND OUTLOOK
The recent advances in computational protein design highlighted in this chapter suggest the
likely areas in which synthetic biology could be impacted by CPD most immediately.
Generally, CPD should have an advantage over directed evolution in cases where the
sequence space related to the design problem is too large for library construction and/or
there is no high-throughput assay. Of the different available techniques discussed here, the
redesign of protein
protein interactions is currently the most robust, and thus most readily
applicable. This puts within reach the redesign of cell-signaling pathways (as presented by
Kapp et al. 19 ) or improved protein therapeutics that bind their targets with picomolar
affinity (as presented by Lippow et al. 16 ).
De novo design of binders against a given epitope on a target of interest, while shown to
work in several examples, is probably still a few years away from being routinely used for
practical applications and arbitrary targets. One problem in this area that is not yet solved
is, in cases where no existing binder is known, how to identify
'
bindable
'
epitopes on the
target of interest and corresponding productive
points on the new binder. For
example, if the hotspot-design strategy is used, it is not yet clear how best to identify
potential hotspot residues if only a structure of the target in its apo form is known.
However, in case a crystal structure of the target in complex with an already existing binder
is known, but that binder happens to have undesirable biochemical properties (i.e. size,
immunogenicity), the existing de novo design algorithms can be used out of the box to try
to design a novel binder starting from a more favorable scaffold. Specificity redesign of
enzymes, i.e. for bioremediation of pollutants or modification of existing metabolic
pathways, should also be achievable with the currently available methods. A further
complication for pathway engineering lies in the fact that, in the pathway of interest,
possibly all enzymes downstream of the first redesigned enzyme would also have to be
redesigned to transform the novel metabolite(s). De novo design of enzymes, being perhaps
the hardest problem in CPD, will likely require several more years of research effort before it
can reliably be applied for practical applications, and should be deployed in combination
with directed evolution to increase the (probably low) activity of the initial designs.
anchoring
'
'
120
References
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