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New enzymes for new needs

New enzymes for new needs

The world is constantly changing and changing needs require new cleaning technology and a greener and more sustainable detergent business.

Enzymes, nature’s own building blocks, can be a solution to these challenges. Produced in fermentation processes, enzymes for industrial use are based on fungi or microbes found in nature. The challenge is to find the right enzyme for any given application and to determine gene encoding for this enzyme.

By Sara Landvik, Science manager, Department of fungal discovery, Novozymes
May 5, 2020

A growing population in a globalized world constantly calls for new technological solutions to support new lifestyles - also in the area of detergents. Changes in eating habits or food ingredients introduce new types of stains, which might be challenging to remove with traditional detergent technology. Stains and dirt might bind differently to novel types of fabrics compared to traditional fabrics. Advanced functionalities of fabrics such as UV protection, rain resistance, and wrinkle freeness, all adds further demands to modern detergent performance.

Changing eating habits, food ingredients and fabrics all create challenges for traditional detergent technology.

Even more challenging, we have to face the fact that the detergent business plays a role in the increased use of fossil fuels and other non-sustainable resources. All steps towards more sustainable detergents are therefore welcome – and have the chance to substantially contribute to a greener world. The trend of washing at lower temperatures is one move in this direction as savings in electricity is obvious. But, this again calls for adjustments of detergent technology to live up to the expected cleaning performance.

 

Enzymes: The little tools of nature and the perfect stain remover

A supreme match to both the requirements of new functionality as well as a greener technology is the use of enzymes. Detergent enzymes have been used to boost the cleaning since 1963, when the first protease was introduced to the market. The strengths of enzymes are their cost efficiency and the specific activity on their target molecules. An amylase type of enzyme only affects starch-containing stains, lipase only acts on specific types of fat stains, and so on. This means that the enzymes will not destroy the fabric or the washing machine, needless to say, unless these would be made by the target material of the enzymes. Due to their efficiency, only small amounts of the enzymes are needed. Typically, enzymes make up less than one to a few per cent of the total detergent and yet they cause a profound difference in washing results. Furthermore, enzymes are of natural origin, and completely biologically degradable. Because of this, enzymes are increasingly used within a number of different industries, not least Detergent.

Enzymes are often called ‘The little tools of Nature’. They are proteins made by all living creatures and are used to degrade or build up structures and functions, inside or outside of the cells. Enzymes are not alive themselves. Fungi and bacteria are examples of microorganisms, which often live in nature by secreting enzymes onto the substrate they nourish upon and thereafter transport the degraded products into the cells as nutrition and energy. Because they secrete these enzymes out into their environment, scientist can harvest and test them. Most enzymes used in the industry are derived either from fungi or bacteria.

Sara and her colleague Mikako Sasa from Novozymes Microbial Bioprospecting harvest fungal samples

Finding the enzyme that works

The challenge is to find the best enzyme for any given application. For this, various strategies are being used. Novozymes A/S, the largest enzyme company in the world, has since the 1950s built up a Culture Collection with a large diversity of natural microbial strains for enzyme screening purposes. This diversity covers various physiological niches(e.g. different pH and temperatures), geographical areas, taxonomic relationships, and ecology of the microbes (e.g.their lifestyles and how they feed). Each fungal strain would on an average be able to secrete a couple of hundred enzymes, bacteria typically a bit less. Depending of the specific lifestyle of the organisms, they will show a different enzyme profile. Thus, a fungus which is able to parasitize and degrade insects would need a significantly different set of enzymes compared to a fungus which is degrading plants. The extremophilic bacteria in thermal regions are known to have some remarkably thermostable enzymes. Keeping as broad a diversity of lifeforms as possible enables us to efficiently make novel discoveries and develop unique enzymes for new applications. This way, we can quickly respond to our customers’ needs, even if we today cannot know what the customers are asking for tomorrow.

Potential sources of detergent enzymes are microbes known to thrive in washing machines and laundry environments (1). Strains from arctic sources might potentially provide us with low-temperature acting enzymes. Microbes from highly alkaline areas might likewise have adapted to their environment by developing alkalotolerant enzymes. However, since it is not the use of the whole organism but the enzymes we are looking for, other sources of organisms can be equally relevant to screen. Thousands and thousands of fungi and bacteria might be tested for their enzyme potentials before an enzyme is pointed out as being interesting for a particular purpose.

Finding the right enzymes for a particular purpose can involve testing thousands of potential candidates.

Exploring the genes

When the relevant enzyme has been found, the gene encoding for this enzyme also has to be determined. Today’s affordable technology of whole genome sequencing allows us to not only identify the target gene but to gather information of all other genes of the strain in question. The total genomic information can then be repeatedly re-used in other projects on a quest for new enzymes.

Once the target gene has been identified, the task of producing the corresponding enzyme is given to another microbial strain, a production strain. This can, for instance, be a strain of Aspergillus oryzae, the type of fungi that have been used to make soya sauce for hundreds of years by secreting enzymes that act on soybeans to produce the well-known sauce flavours. In our hands, the fungus is not producing its whole original range of enzymes but mainly the enzyme in question. This way, the production of the selected enzyme becomes more economical, clean (no interfering undesired enzymes), and safe in use.

As described, the whole genome sequencing technology enables us to quickly discover gene sequences not only from selected and characterized microbial strains but also from metagenomics sources such as a soil sample. A metagenome contains a mixture of different microorganisms of which only a minor part might be culturable (and therefore more easy to test) in the laboratory. Other types of non-culturable organisms that today can be utilized in enzyme discovery too are symbiotic fungi, which cannot grow in the laboratory without its symbiotic partner. This could be for instance mycorrhizal fungi that live in symbiosis with a plant via the plant roots or fungi that form lichens together with an alga.

 

Building on the blocks of nature

Nature indeed provides us with extremely versatile enzymes. However, it can be difficult to completely match the requirements of the enzymes in some of the harsh industrial environments. It can be particularly challenging to find an enzyme with all of the combined desired properties for a certain application; specificity for the substrate; the speed of action; the right pH and temperature activity; tolerance to other factors such as surfactants; stability in product formulation; stability on the shelf, and so on.

Enzymes for detergent applications are put to the test in Novozymes Detergent Research Center

If the performance of the best-in-class enzymes is not good enough, these enzymes might be subject to Protein Engineering. Protein engineering means that scientists are inducing changes in the enzyme molecules, which are then tested for improved performance. The changes in the molecule are all natural in the sense that they could by chance all happen in nature, too. However, in nature, these changes would be slower and established in a certain microbial strain. Similar changes can be accomplished by breeding but it would take a long time to wait for the changes in the molecules to be established before we could screen and select out these variants. Instead, we use tools similar to those, which are used in the cause of evolution by the biological cell – only we use them in a controlled way.

Considering diversity as the keyword for all our activities, these above enabling technologies make sure that we are not short of interesting leads for a greener detergent industry!

 

It’s payback time fungi!

One might think of dinosaurs as old creatures of Earth – but infact, the development of fungi and bacteria far precede the dinosaurs in time. While dinosaurs originated an estimated 230 million years back in time, fungi and bacteria originated 1500 resp. 3500 million years ago.

During this period, they have continuously developed new enzymes while evolving into new life niches. Some bacteria impress by being able to live in such extreme environments as boiling water and alkaline environments up to pH11. Some bacteria are even chemotrophic, meaning they can harvest energy from other sources than carbon by oxidation of organic or inorganic molecules. On the other hand, many fungi are generally regarded as superior to bacteria in degrading some highly complex substrates, such as the lignin-cellulose complex of wood. As such, they play a huge role for life on earth by recycling this biomass back into the ecosystem.

Recent studies suggest that the development of the enzyme system, which allows fungi to degrade the lignin in wood, coincides with the ending of the carbonaceous era (2). This makes sense since there at this time was an abrupt stop of the ongoing accumulation of wood, which by time turned into coal. So, jokingly it might be fair to say that it is because of the fungi that we are running short of this kind of fossil fuel today. Perhaps, it is just as fair to say “It’s payback time, fungi – you put a stop for the accumulation of biomass and the following fossil fuel build up, now please provide us with enzymes that can help us to be less dependent on fossil biomass in our industries”. And the payoff has just begun.

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