has around 150 biocatalytic processes, based on enzymes. This is likely to be an underestimate because new enzyme technologies are developing all the time and there is also a large number of niche, highly specialised, enzymes made for one-off jobs. Chemists have exploited the range of different building blocks available to them to add to the properties of the natural catalysts. However, the biggest innovations over the past two decades have come from the use of genetic modification techniques, which were introduced in the late 1980s. These helped to raise enzyme yields, which lowered production costs, and broadened their specificity.
Nonetheless, these genetic methods
did not radically alter the characteristics of natural enzymes. The introduction of methodologies, such as ‘gene shuffling’ or ‘directed evolution’ using high throughput screening in the late 1990s, triggered much bigger changes because they allowed chemists to make enzymes not found in nature. However, chemists maintain the enzymes are still natural because the new techniques merely speeded up evolutionary processes in enzymes that have been selected from those found in nature. ‘Our (development) work begins with
the search for suitable enzymes from natural sources,’ says Andre Koltermann, head of the biotech and renewables centre of Clariant, which took over a 10- year old R&D operation in enzymes when it recently acquired Sud-Chemie, the German catalyst specialist. ‘We first look
$3.4bn
The estimated value of the global market for industrial enzymes, which represent around 10% of the total catalyst market
for naturally occurring microorganisms that can utilise the raw material that we want to process – straw, for example [for biofuels production],’ he explains. Besides wanting microorganisms that have the attributes needed for applications on specific substrates, enzyme makers also want ones that can be manufactured in large amounts and will then perform efficiently in an industrial processing plant. ‘You can’t always be sure that the enzyme you have selected is the best one for manufacturing on a mass scale,’ says Neilsen. ‘Then the big issue in its application is stability. There are so many stability requirements with enzymes in relation to processing conditions like temperature and other factors.’
High throughput screening involves
repeated cycles of random mutagenesis (changing the genetic information) or gene recombination followed by screening to pick the most suitable mutant for a specific application. Large numbers of mutations are produced in a very short time – far faster than would occur in nature.
In a typical automated screening
procedure, a microtitre plate – usually with 96 wells – is used to grow a
30 Chemistry&Industry • November 2012
Over the past 20 years, researchers have considerably raised yields in two areas – in the mass-scale manufacture of the enzymes themselves and in their application processes. Since the 1980s, there has been a 100-fold increase in yields of fermentation processes for making enzymes, which has also driven down costs.
In July 2012, Clariant opened a €28m demonstration plant at Straubing,
generation of mutants, which are screened for desired characteristics. Hundreds of thousands of variants are compared with one another. The most suitable mutant then becomes the template for the next round of mutagenesis, with the process repeated until the microorganism with as many of the right specifications as possible emerges.
A crucial part of the scaling-up
exercise is the transfer of a selected enzyme to a pilot plant where deficiencies might be revealed requiring further automated screening to make further modifications so that the enzyme works as effectively as possible with the chosen substrate. ‘The goal is to get the enzyme to the point where it is tailored to the process conditions and does the job every efficiently,’ says Koltermann.
Using rational design, chemists can make superior enzyme mimics for industry
Novozymes has about half the indus- trial enzymes market