Usina bioloaical catalvsts
'or chemical synthesis
Dr Marc Struhalla, managing director of c-LEcta, looks at how biocatalysis has emerged as a key
technology for the manufacture of speciality chemicals
B
iotechnology is one of today's
tiomer is often undetectable, so very
most rapidly developing tech-
high
nology areas, leading to new
achieved.
products being developed in many
different
industries.
One
chiral
purities
are
being
In addition to high specificity, bio-
very
catalytic transformations also offer
dynamic application area is the use
the advantage of being run in very
of biotechnological processes for the
moderate
production of speciality chemicals.
Usually they run at atmospheric
The use of enzymes or microorganisms for the synthesis of chemical
pressure,
reaction
at
low
conditions.
temperatures
(<100'O and in aqueous reaction
products is referred to as biocatalysis.
systems. Toxic substances can be
These products range from
avoided and solvent use reduced.
high-
value chiral intermediates for the
Finally, the use of biocatalysis often
pharmaceuticals industry to fine and
allows a significant reduction in raw
speciality chemicals for agrochemical,
material costs and of the number of
cosmetic and food applications. Figure
total synthesis steps needed within a
1 shows the number of biocatalytic
specific route.
Unfortunately, making use of the
processes implemented in industry
since 1960. About 65-70% of these
potential advantages of biocatalysis
ciency with respect to space/time-
in order to secure rapid access to fast
have been used in pharmaceutical
in practise is not that simple. Finding
yields, inhibition effects and the eco-
feasibility evaluations.
and agrochemical applications.
the right biocatalyst for the desired
nomics of producing the enzyme
The use of biocatalysis in chemical
applications is a challenge, especially
itself is likewise of great importance.
enzymes are not often adapted to
Nonetheless,
synthesis can be highly advanta-
when the relevant industries have
Enzyme activity, in terms of its
the specific needs of specific industri-
geous by comparison with chemical
short development times.
specific activity, turnover rates, pH
al applications. They might not show
All biocatalytic processes, whether
and temperature profile, also plays a
the
processes make use of the high
they use microorganism as biocata-
crucial role. And, last but not least,
desired substrate molecule or they
stereo- and enantio-selectivity of the
lysts in whole-cell transformations or
enzyme specificity with regard to its
might bring along insuffident stere-
enzymes, which can act in a highly
isolated enzymes, depend on the
substrate spectrum, enantio- and
oselectivities.
provision of suitable enzymes with
regioselectivity needs to be considered. This complex matrix of enzyme
properties consisting of activity,
compounds.
Some very successful examples of
the direct use of enzymes for chiral
defined properties. This search for
the ideal enzyme is a multi-parameter process, depending on a number
of enzyme properties.
Enzymes have complex
three-
syntheses have been the use of
dimensional protein structures, which
ketone reductases and transaminases
might be unstable under certain cir-
to transform ketones into high-value
chiral alcohols and chiral amines
specificity, stability and efficiency
parameters makes the search for the
ideal enzyme for defined biocatalytic
turnover
available
syntheses. Very often biocatalytic
specific way on defined chemical
molecules, for instances allowing
direct chiral synthesis of high value
necessary
the
on
the
At the end of the day, enzymes
are usually catalysts doing a very
specific job in their natural surroundings; they were not evolved by
nature to serve the demands of
industrial chemical synthesis. In these
instances, technologies are needed
applications a challenging task.
The enzymes used in industrial
to develop a new enzyme in a cus-
cumstances, Thus, parameters like
biocatalysis are usually recombinant
tomised way. One option, of course,
temperature, pH and the presence of
enzymes produced in microorgan-
is to screen natural biodiversity for
respectively. In such applications, the
solvents, additives and by-products
isms. These organisms are genetical-
formation of the undesired enan-
must all be considered. Enzyme effi-
ly modified in a way that they produce the desired enzyme with high
efficiency and large yields. Very
new enzyme activities.
As shown in Figure
often, relatively crude enzyme preparations derived from microbial fer-
1960
1970
1980
1990
2002
2008
2020
Year
Figure 1 - Biocatatalytics process implemented in industry, 1960-2020
50
2, when
searching for adapted enzymes for
defined biocatalytic processes, fast
and efficient screening processes are
needed. Here, beginning with a
mentations are applied in the synthe-
library stored as frozen cultures, the
sis processes. Work-up needs to be
candidate enzymes are produced by
simple in order to limit the enzyme
cultivation in a micro-titreplate for-
costs in the application.
The most straightforward way of
mat, screened for interesting activities with a photometric assay, then
developing a biocatalytic process is to
further characterised in a secondary
make use initially of enzymes already
screen based on gas chromatography analytics.
known to be available. Technology
companies focusing on the develop-
One commonly applied strategy is
ment of biocatalytic processes are
to use microbial strain collections, In
building up large enzyme collections
these, microbial strain candidates are
June 2009 Speciality Chemicals Magazine
Biotechnology
cultivated and analysed for the presence of a defined enzyme activity. If
and when a good enzyme candidate
is found, it must be isolated and the
enzyme gene encoding its protein
sequence identified. Without the protein sequence, the development of
an efficient recombinant production
process for the enzyme is impossible.
A very good alternative to microbial screenings is the use of recombinant expression libraries representing natural diversity. Here, one starts
with the DNA containing the genetic
information for the enzyme candidates. DNA isolated from natural
sources is isolated, fragmented and
cloned into expression vectors.
lndividual gene fragments containing
gene candidates are translated into
the encoding proteins, which are
afterwards screened for the desired
enzymatic activities,
lnnovative screening technologies
make possible a very high throughput
in such recombinant biodiversity
screenings, analysing millions of
enzyme gene candidates in a few
days. The biodiversity addressed by
such approaches can again be derived
from microbial strain collections, for
instance, but it can also be directly isolated from environmental sources.
DNA can be extracted from interesting microbial habitats directly and
used for the production of so-called
metagenome libraries. Many microbial habitats, for instance soil samples, contain a large fraction of
micro-organisms which cannot be
cultivated and which are not accessible by microbial screenings. These
organisms can make up more than
99% in certain samples and this biodiversity can be rapidly made available through metagenome expression technologies.
Efficient technologies to provide
new enzymes from natural diversity
are powerful technologies for biocatalytic process development. Nonetheless, the enzymes that nature provides
do not often fulfill the demands of
industrial processes.
In these instances, technologies
are needed which allow for the optimisation of natural enzymes or the
development of artificial enzymes
with completely new properties. Both
options are available now by making
use of innovative enzyme engineering technologies.
New enzyme properties are developed in the laboratory, making use
of directed evolution. By using
sophisticated mutagenesis techniques, large libraries of enzyme
variants consisting of hundreds of
thousands up to millions of individual enzyme candidates are generated.
Again rapid screening procedures
are needed to analyse this diversity
in a straight forward way. By using
such approaches, new enzymes have
been evolved that are active under
unusual reaction conditions or which
are highly active on non-natural substrates.
Enzyme engineenng technologies
make it possible to bring biocatalysis
to the next level. Not only the broad
set of enzymes that nature provides
but also evolved, artificial enzyme
mutants are made available and
accessible by using highly developed
molecular biology technologies.
Conclusion
The use of biocatalysts for the synthesis of high value speciality Chemicals not only has great potential, it is
already
an
economic
reality.
lnnovative biotechnological technologies have became available in
recent years that speed up the development times and increase the
chances of success of biocatalytic
process developments.
New natural enzymes and also
artificial enzymes generated by lab
evolution are being made available
to the relevant industries. Making
use of these technologies allows
innovating synthesis route development and thereby offers significant
economic advantages to the applicants. The use of biocatalysis offers
great opportunities to improve the
competitive situation in the speciality
chemicals markets.
For more information, please
contact:
Dr Marc Struhalla
cLEcta GmbH
Deutscher Platz 5
D-04103 Leipzig
Germany
Tel: +49 341 355 214-0
E-mail: marc.struhalla@
c-lecta.de
Website: www.c-lecta.de
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