EmPowerPutida

Could this beastie replace the fossil fuel industry?

Technological innovation for agricultural advancement

Biochemistry could eventually replace the fossil fuel industry. Photo: Shutterstock

Estimated reading time: 9 minutes

A bacterium as a chemicals factory, refinery or waste processor? Pseudomonas putida is going to be competing with the fossil fuel industry if Wageningen professor Vitor Martins dos Santos has anything to do with it. He and his European colleagues investigated the possibilities.

This story starts in 1928, when the British doctor Alexander Fleming discovered by accident that the mould Penicillium notatum secretes a substance that kills bacteria. However, the mould only makes very small quantities of penicillin. The discovery therefore did not seem particularly useful, and Fleming lost interest. It was only fifteen years later that people realised the true potential of penicillin as an antibiotic. That was when it became clear that if you let the mould produce large amounts of penicillin, it can be used effectively to combat infections.

But how do you get a mould, bacterium or other organism to secrete more of the molecules you want? Or to do so in very different conditions to those to which the organism is accustomed? Or, taking things even further, to produce completely new molecules so you can compete with the fossil fuel industry? Because that is the major challenge we are now facing - finding alternatives for products that currently use oil, such as plastics, paint, rubber and polystyrene.

Pseudomonas putida can make the building blocks for various products that currently require oil

The Pseudomonas putida has generous boundaries, making it suitable to tinker with the cellular factory. Photo: Shutterstock

Scientists in the laboratory of Vitor Martins dos Santos, professor of Biomanufacturing & Digital Twins at Wageningen University & Research (WUR), are working precisely on those issues. Martins dos Santos uses bacteria and other microbes as cellular factories to make all kinds of substances. Sometimes he gets those ‘factories’ to work faster, sometimes he gets them working in different conditions and sometimes he persuades them to make completely different products.

Impossible competition

Martins dos Santos gives beer as an example. It contains yeast that makes alcohol. But in normal conditions, the alcohol the organisms produce doesn’t get above 5 to 10 per cent. That is fine for a flavoursome beer but not much use if you need large amounts of alcohol. “The same applies when making industrial products such as bioplastics or coatings. You need yields a hundred times bigger with the same number of microbes. Otherwise it’s impossible to compete with the traditional petrochemical industry.”

The aim of Martins dos Santos’ research is to help find a solution for one of the biggest challenges humankind is facing: the transition from a fossil fuel economy to a green economy. If we no longer need oil or other finite resources to make products such as plastics or lubricants, that will lead to a huge reduction in both CO₂ emissions and pollution.

Genetic pipeline

To get those microbes working faster and in a greater variety of ways, Martins dos Santos is coordinating the Biomanufacturing research line This research involves making microbes capable of producing a wide range of molecules under a wide range of conditions. He achieves this by reprogramming them at the genetic level.

EmPowerPutida is exploiting native endowments by re-factoring, re-programming and implementing novel control loops in Pseudomonas putida for bespoke biocatalysis. This video briefly explains how they do it.

A microbe’s genes give instructions for the production of thousands of enzymes. Those enzymes in turn are responsible for the formation of molecules or for catalysing their production. So if you change the genes, that will lead to new enzymes and consequently new molecules. Together they make up the cell’s ‘factory’. “As in any factory, you want to make sure all the supply and discharge pipelines are connected up properly. I create those links within the microbes by tinkering with the genes and their expression. That lets me create a detour in the biochemical pipelines.”

Favourite conditions

Each microbe has its own favourite conditions for each production process: with oxygen or without, in acidic or alkaline surroundings, at a high temperature or a low one and so on. The Biomanufacturing researchers can modify those ‘favourite conditions’. “We can reprogram the microbes so that they are suddenly able to function without oxygen, or become resistant to toxins that are released during the production process.”

Research is helping the transition from fossil fuel economy to a green economy

Removing such thresholds and limits in the cellular factory requires an awful lot of tinkering. That is why Martins dos Santos uses computer models. They can tell you that to make substance X, you can use microbe Y but need to modify gene Z. The next step is putting the computer model into practice in the genes of an actual microbe, for example by using CRISPR-Cas, a relatively simple technique that lets you add new genes in an organism and remove other genes very precisely.

However, the model outcomes do not necessarily match reality. “A model always produces some kind of output so you need to test it with real-world data. By feeding those results back into the computer, you get an interactive model that is self-learning.” Martins dos Santos also uses artificial intelligence to improve the predictions of his models. And he deploys ‘digital twins’, which are digital copies of a microbe where he can try out what happens if he moves or replaces one of the genetic pipelines.

Tinkering

A starring role in his work tinkering with cellular factories is reserved for Pseudomonas putida. This bacterium is found all over the place in nature, it is unaffected by many toxins, can convert a broad spectrum of molecules and can tolerate a wide range of conditions. Martins dos Santos’ research group has managed to extend that range even further – and in some cases eliminate the limits altogether. Their version of P. putida can produce substances without oxygen (which it normally needs) and survive at higher temperatures than it would normally find comfortable (which means that production can be speeded up).

Our role is to introduce innovations and perform tests. If they work, we pass them on to the market

The P. putida is now able to produce the building blocks that could replace palm oil, amongst other things. Photo: Dede Sudiana / Shutterstock

Pseudomonas putida is now capable of making the building blocks for various products that currently require oil. You can use it to make biobased plastics or an alternative to palm oil. What is more, the bacterium can break down toxic substances, which means it can be used to purify contaminated wastewater. The microbe can also convert lignin, a waste material of plants that is difficult to break down, into nylon, which would therefore no longer need to be made from oil. These results have led not just to numerous scientific papers but also to patents for the transformations in question.

Obstacle

So can we say goodbye to the fossil fuel industry? No, we are not yet that far. First because numerous technical issues need to be resolved in order to scale production of valuable chemicals (or the breakdown of chemicals) by Pseudomonas putida up to industrial levels. But a bigger obstacle is the fact that it is hard for the traditional industries to completely overhaul their existing processes.

“Fossil, oil-based processes have been investigated thoroughly as they have been used for over a century. People know exactly how they work. Everything is perfectly aligned and optimised.” That means the products can be manufactured very cheaply, very efficiently and with a high level of productivity. The cellular factories of Martins dos Santos are much more expensive, are less stable and they are slower. “Biological production involves many unknown factors and entails many risks. Furthermore, it is not yet suitable for the infrastructure that industries have at present. You would have to build new factories.”

The Tesla for biochemistry

That is why the hope is that a Tesla for biochemistry will appear. After all, when the Tesla car company showed that electric cars really can compete with the fossil fuel variant, traditional car manufacturers made the switch too. “The chemicals industry will only make the change when it becomes unavoidable because it is very expensive and will require a lot of adaptations. That is why you need serious financial incentives, or regulations that make it compulsory. So it is a combination of market factors, external pressure and technical possibilities.”

Which does not mean there is no sign of any movement. Martins dos Santos is already collaborating with industrial giants such as BASF, DSM and Mitsubishi, as well as public bodies such as the Dutch National Institute for Public Health and the Environment (RIVM) and Ministry of Agriculture, Nature and Food Quality, plus other research institutions. “We always use the ‘golden triangle’ approach of working with governments, the private sector and science institutes. Our role is to introduce innovations in the concepts and models. We test them. If they work, we pass them on to the market.” Returning to the penicillin story, Martins dos Santos suspects it could be another fifteen years before we see the results of his work on the market. That will be thanks to Pseudomonas putida, as versatile as it is minuscule.

European research context

EmPowerPutida addresses the following European policy challenge: The transition from a fossil-based economy to a biobased economy

Wageningen University & Research groups involved: Bioprocess engineering

European and other countries involved: Belgium, Denmark, France, Germany, Greece, the Netherlands, Portugal, Spain and United Kingdom, plus multinationals from Japan, South America and the United States

Duration: 2018 – 2022

Share this article