Author: Darío Sánchez, PhDHave you ever heard the word “extremophile”? It refers to an organism that is able to live in extreme conditions, like very hot, cold, or high-pressure environments. These are usually microorganisms as well, like bacteria, archaea, fungi… And because of them, life and society have changed in the last 50 years. In fact, if it was not for extremophiles, you might even be dead.
COVID, DNA polymerases, and PCR.
A few years ago, the world watched as a virus spread throughout the world. Causing panic, death, and chaos across cities big and small. The COVID-19 pandemic changed how we all see society.
While vaccines were being manufactured and provided, its spread was curbed by a simple but tough to implement system: detection, isolation, recovery.
The detection of this virus and almost any other pathogen on Earth is done the same way: PCR, or Polymerase Chain Reaction, which you most likely have heard about. This method, fully developed in 1986, works by amplifying existing DNA in an exponential manner, using an enzyme known as DNA polymerase.
There are many kinds of DNA polymerases, but PCR worked terribly when using “normal” DNA polymerases. The process relies on the two strands of DNA separating, each being copied on the next cycle, then separated again, and so on. This requires pretty high temperatures, close to 100 °C.
Normal proteins cannot withstand those temperatures, and DNA polymerases from normal organisms are no exceptions. After such heat, their structure changes, and they no longer work. So how do we do PCR today? To tell you, we have to go to the famous Yellowstone Park, in the US, and back in time to the 1960s.
Molecular revolution from a Yellowstone hot spring
Before PCR was invented, scientists were already growing and purifying proteins in the lab. Those proteins often came from simple bacteria, like E. coli, which is found in many places and is easy enough to grow in the lab. One of the first DNA polymerases used in in-vitro assays was, in fact, the DNA polymerase I of E. coli.
But often, standard proteins do not cut it for cutting-edge science. And in Yellowstone park, there was research being done around the hot springs. Scientists believed that there could be no life there, as proteins and other molecules would degrade at the high temperature of those hot springs.
They were proven wrong, and in 1969, a researcher working there managed to isolate and culture an extremophile for the first time, the bacteria Thermus aquaticus.
This was groundbreaking for two reasons. First, they proved that life could actually thrive at over 70 °. We now know that to be the case of many microorganisms. But most microorganisms to ever exist are impossible to grow in a lab. The percentage is even lower for extremophiles. Thermus aquaticus was not only found and isolated, but grown in a laboratory environment.
This meant that proteins that were somehow able to survive extremely high temperatures and still work could now be harvested and used for in-vitro processes.
It was Thermus aquaticus, or Tac, DNA polymerase that was the enzyme needed to make PCR work. The high temperatures of the thermal cycling did not break and inhibit this new thermostable polymerase.
Extremophile enzymes and their uses
You may believe that extremophile enzymes are relegated to a lab and molecular diagnostics. Nothing further from reality. Here are some areas where enzymes found originally on extremophiles are used:
- Laundry detergents: Most store-bought laundry detergents have enzymes. These need to survive high temperatures and harsh acidities, so they often come from extremophiles. They break down fat, protein, and sugars, cleaning the stains of your textiles.
- Paper and textile industry: Both the paper and textile industry use enzymes for bleaching and cellulose digestion. Paper pulp is whitened using cellulose-breaking enzymes, and textiles like denim are faded with peroxidases. They sometimes come from extremophiles too.
- Bioplastic production: Plastics made from renewable sources, like PHA, can be produced by halophilic bacteria (Thriving under high-salt conditions). Enzymes from these organisms can be used as well with salt to produce these bioplastics.
- Environmental bioremediation: Extremophiles that can survive high radiation, break down fossil fuels, and survive other environmental conditions where humans can’t thrive are used for bioremediation, reducing radiation, eating plastic, petroleum, or other materials, and helping restore the environment, all through special enzymes that most organisms lack.
- Therapeutics: From skin care to cancer treatments, molecules from extremophiles are extremely interesting for humans to improve their health. Their enzymes are used to manufacture therapies and even used in the therapies themselves.
Mining in the Arctic: DNA and useful genes from the past
By now it should be clear that extremophiles are very important. But finding them and growing them in a lab is hard. Fortunately, there are other ways to find key proteins and enzymes able to help us. DNA holds the information needed to make proteins, and with some simple genetic engineering, we can make everyday laboratory-grown bacteria like E. coli express and produce nearly any given protein.
But while many extremophiles are being discovered living, the TOLERATE project looks to the past. Genetic records from Arctic expeditions may hold the key to old enzymes to tackle new challenges. Because the Earth has gone through multiple changes in temperature and different climactic events, organisms (and especially microorganisms) have evolved to survive said climates, then died.
Ancestral sequence reconstruction
Among the tools to do this, ASR, or ancestral sequence reconstruction, is of important relevance to the TOLERATE project.
This technique is used to find enzymes that are similar or prior to existing ones, matching genetic sequences from today’s organisms to those in ancient DNA. This means that scientists are not peering blindly into the ancient genetic archives found in the archive, but can use current genetic sequences to trace their ancestors or extinct brothers and sisters.
This way, mutants of currently used enzymes could be found. The activity and characteristics of these mutant enzymes can be predicted, filtering for those with better activity in extreme climates, like very warm, dry, cold, etc.
TOLERATE’s aims: ancient molecules and enzymes to better today’s world.
One of the goals of TOLERATE is to find molecules, proteins, and enzymes, coded in ancient DNA from the Arctic, to improve today’s society. Using ASR they can understand what ancient proteins can do, based on similarities from today’s known proteins. Then, using lab-grown bacteria, they can engineer them to introduce genes to produce relevant ancient molecules and proteins.
The project aims to use these on several areas. Sugars and small molecules can be used for 3D printing and drug delivery systems, and large sugars for biological nanofibers. Enzymes can be used for industrial cleaning and detergents (soaps, laundry detergents, etc.). Lastly, engineered bacteria with enhanced resilience to heat and drought can be used to improve crops, adding these bacteria to the plants’ soil microbiome.
Overall, TOLERATE’s approach might result in completely new scientific breakthroughs that could transform even your everyday life!
