Chapter 2
Extension of substrate range.
In many industrial processes it is interesting to extend the
substrate range for the applied microorganism in order to have a more efficient utilization
of the raw material. Initially, it is necessary to insert the necessary pathway (or enzyme)
for utilization of the substrate of interest. Subsequently, it is important to ensure that the
substrate is metabolized at a reasonable rate, and that the metabolism of the new substrate
does not result in the formation of undesirable by-products. This may in some cases
involve extensive pathway engineering.
Pathways leading to new products.
It is often of interest to use a certain host for
production o f several different products, especially if a good host system is available.
This can be achieved by extending existing pathways by recruiting heterologous
enzymes. Another approach is to generate completely new pathways through gene
shuffling or other methods of directed evolution. In both cases it is often necessary to
further engineer the organism to improve the rate of production and eliminate by-product
formation.
Pathways
for
degradation
of xenobiotics.
Many
organisms
naturally
degrade
xenobiotics, but there are few organisms that degrade several different xenobiotics. In
bioremediation it is attractive to have a few organisms that can degrade several different
compunds. This may be achieved either by inserting pathways from other organisms or
through engineering of the existing pathways.
Engineering of cellular physiology for process improvement.
In the industrial
exploitation of microorganisms or higher organisms it may be of significant interest to
engineer the cellular physiology for process improvement,
e.g.
make the cells tolerant to
low oxygen concentration, less sensitive to high glucose concentrations, improve their
morphology, or increase flocculation. In cases where the underlying mechanisms are
known this can be achieved by metabolic engineering. This may involve expression of
heterologous genes, disruption of genes or over-expression of homologous genes.
Elimination or reduction of by-product formation.
In many industrial processes by-
products are formed. This constitutes a problem due to the loss of carbon to
the by-
products, due to toxicity of these compounds, or due to interference of the by-products
with the purification of the product. In some cases the by-products can be eliminated
through simple gene disruption, but in other cases the formation of the by-product is
essential for the overall cellular function and disruption of the pathway leading to the by-
product may be lethal for the cell. In the last case it is necessary to analyze the complete
metabolic network, and based on this analysis design a strategy for reduction of the by-
product formation.
Improvement of yield or productivity.
In many industrial processes, especially in the
production of low-value added products, it is important to continuously improve the yield
and/or productivity. In some cases this can be achieved simply be increasing the activity
of the biosynthetic pathway,
e.g.
by inserting additional gene copies. In other cases the
pathways leading to the product of interest involves many steps, and it is therefore not
possible to increase the activity of all the enzymes. Analysis of the flux regulation in the
pathway is therefore required. Finally, in some cases the limitation is not in the actual
pathway, and it may therefore be necessary to engineer the central carbon metabolism,
which is generally very difficult due to the many control structures present here.
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