A mysterious protein, SGGW and world recognition

Each organism is made up of thousands of different proteins that it needs to live. For example, a man has over 20,000 of them. One fourth of them has not yet been tested, so we do not know what they are for. We will probably not find out soon. Research into completely unknown proteins is very risky and it is difficult to obtain funds for them. There is a risk that the work put in and the money invested will not contribute to any discovery. Dr hab. Krzysztof Pawłowski from the Department of Experimental and Bioinformatics, Faculty of Agriculture and Biology, Warsaw University of Life Sciences, took a risk and succeeded! The Polish research team he headed managed to obtain a grant to research a previously unknown protein. The team was composed of PhD student Monika Karasiewicz-Urbańska from the Independent Department of Microorganism Biology at the Faculty of Agriculture and Biology of the Warsaw University of Life Sciences, dr hab. Róża Kucharczyk from the Institute of Biochemistry and Biophysics of the Polish Academy of Sciences, dr hab. Małgorzata Łobocka from the Independent Department of Microorganism Biology at the Faculty of Agriculture and Biology of the Warsaw University of Life Sciences and the Institute of Biochemistry and Biophysics of the Polish Academy of Sciences, as well as a PhD student Sylwia Pilch from the Institute of Biochemistry and Biophysics of the Polish Academy of Sciences. The unknown protein has just been discovered by Dr. Hab. Krzysztof Pawłowski. Then an international research team was formed, consisting of two Polish and American ones, which investigated the newly discovered protein. The American team was led by Dr. Vincent Tagliabracci of the University of Texas Southwestern Medical Center in Dallas, USA, and he is the lead author of the publication in the journal Cell.

Discovery of protein

Where did it all start? From the observation of the protein sequence, i.e. the sequence of amino acid residues. A protein is a long chain that can take many shapes. Most often, its shape is associated with a twisted serpentine, but is actually much more complicated. The type of "ball" the protein will roll in and how it will work depends on the amino acids it contains and the order in which they are arranged.

- "To make it easier to think about proteins, they are written in the form of a sequence of letters, because there are about 20 amino acids in proteins" - explains Dr. Krzysztof Pawłowski. - “Each of them can be assigned a different letter. The sequence of amino acids in the chain is a string of letters. In the average protein of these letters there will be from several dozen in the shortest proteins to several thousand - in the longest. Interestingly, the beginning of new discoveries in protein biology is often born from "looking" at those long strings of letters. " -

The effect of "observing" strings of letters carried out with the use of bioinformatics methods was the discovery of interesting properties of a hitherto unknown family - selenoproteins O (selO).

- "I observed a few things" - explains Dr. hab. Krzysztof Pawłowski. - First of all, it is a protein that has changed very little since the ancestors of humans, fungi, protozoa, bacteria lived on earth, that is, for a very, very long time. It is quite rare. Normally, proteins change so much as they evolve that finding similarities is difficult. On the other hand, if the similarity is high, the natural conclusion is that it is something very important to the body. So there had to be compelling reasons why such an important and perfect tool would not evolve much in the course of evolution. "

This was the first reason to research a mysterious protein and discover why it is so important.

“Second, selenium is a fairly rare element,” she adds. - “For the body, building a selenoprotein is an expensive 'investment'. It's easier to make protein without selenium. Man has over 20,000. proteins, just over 20 of them are selenoproteins. We use expensive things for special purposes. In addition, I found SelO's evolutionary similarity to kinases, very important enzymes used in cell signaling that turn various processes on and off. The fact that the protein discovered is a selenoprotein and presumably a kinase that has behaved well throughout evolution has further suggested that it is an important object. From various bioinformatic considerations we have concluded that this protein is active in response to the so-called oxidative stress, that is, it helps in the appropriate response to difficult environmental conditions related to the harmful effects of reactive oxygen species ”.

Resistance to oxidative stress

To confirm their assumptions, scientists conducted an experiment using Escherichia coli bacteria. A strong disinfectant - hydrogen peroxide was added to a drop of a solution in which there were bacteria. After a while, bubbles appeared because the bacteria "felt uncomfortable" in the solution and began to break down the hydrogen peroxide. To this end, E. coli bacteria turned on various enzymes that made it possible to mitigate the effects of this lethal substance. When a strain of bacteria was genetically engineered by switching off the SelO gene, it turned out that the handicapped bacteria fared worse, with far fewer bubbles. The experiment showed that the selO gene influences the body's ability to cope with oxidative stress.

However, not kinase?

The next step was to see how this unknown protein works. The team of Dr. Vincent Tagliabracci, who determined the three-dimensional structure of the SelO protein. First, he obtained a crystal of the studied protein, and then scattered X-rays on it. In this way, he obtained a diffraction pattern and recreated the position of each protein atom. This knowledge means getting to know the structure of a protein in detail and greatly facilitates understanding of its function.

The result of the study surprised the scientists. It turned out that the structure of the protein is indeed similar to a kinase, but in the image, the ATP (adenosine triphosphate) molecule that the kinase uses for phosphorylation is associated with the SelO protein in a very unusual way. It was concluded from this, and then it was shown biochemically, that the secret protein SelO AMPylates other proteins, i.e. AMP (adenosine monophosphate) joins them, thus regulating their action. This means that SelO is an AMPylase.

- “This result is quite a sensation. While there are over five hundred human kinases, only one AMPylaase has been known so far. The one we discovered is the second. In addition, it is very well preserved in terms of evolution. We have shown that, indeed, AMPylation means "attachment" to other proteins of an AMP residue and is needed to regulate the response to oxidative stress. This is what our article in "Cell" was about, but our discovery opens up a lot of different research topics "- summed up Dr. Krzysztof Pawłowski.

The material was prepared by: Anita Kruk, Promotion Office of SGGW

Substantive consultation: dr hab. Krzysztof Pawłowski, Department of Experimental and Bioinformatics, Faculty of Agriculture and Biology, Warsaw University of Life Sciences