Breaking down the microbiology world one bite at a time
From Antarctica to our freezers.
Antarctica is a continent that people, and even scientists, do not know much about. With the gigantic ice shelves, dry air, and lowest temperatures ever registered, it is hard to imagine any life forms to survive in these conditions. Nevertheless, there is a great diversity of microorganisms present there. They are called psychrophiles (or cryophiles) and, in the course of evolution, they developed several growing and adaptation strategies that allow them to thrive even during Antarctic winters.
Antifreeze proteins are the secret to their survival. Synthesized by bacteria, these ice-binding proteins cause ice to grow in between antifreeze proteins, thereby pushing the freezing point1 and preventing ice from forming big crystals that can be damaging for the cells. In addition, during the freeze-thaw cycles, antifreeze properties of this protein can inhibit ice recrystallization2. Therefore, it’s not surprising that antifreeze proteins are pretty common among species that inhabit cold regions of Earth. Forms of this protein were earlier identified in marine fish3 and even in snow mold fungi4.
In his study, Muñoz5 and his colleagues were determined to show that Antarctic bacteria use antifreeze proteins for their survival. The researchers conducted the experiment in freeze-thaw conditions and isolated three viable microorganisms from the following genera: genus Plantibacter, genus Sphingomonas, and genus Pseudomonas. In order to establish whether or not these organisms possess antifreeze properties, scientists needed to figure out which genetic sequences regulate the production of these proteins. The authors reported that Plantibacter had two distinct sequences (gu3A and gu3B), Pseudomonas had only one unique sequence (afp5A), and Sphingomonas appeared to have a match with a known antifreeze sequence from Antarctic bacteria Marinomonas primorynsis6.
To test whether or not these sequences were, indeed, directing biological pathways to produce the antifreeze proteins, the researchers tested proteins made from these sequences on two food samples (Figure 2): a cucumber (images A and B) and a zucchini (images C and D). The underlying idea behind this experiment was to demonstrate that the antifreeze proteins extracted from bacteria would protect cells of food samples from the damage caused by ice. The red arrow on image A shows a cucumber cell that has been damaged because the antifreeze proteins were not present in the solution. On the other hand, image B shows a cell with no ruptures or damage because an extracted antifreeze protein was applied. The same results were observed on a zucchini sample, where image C represents a sample with no antifreeze proteins and image D represents a sample with applied antifreeze protein. These experiments showed that antifreeze proteins prevent cell ruptures caused by ice crystallization.
A. without antifreeze proteins
B. with antifreeze proteins
C. without antifreeze proteins
D. with antifreeze proteins
Red arrows indicate a damaged region of the cell, where ice crystals caused disruption of a cellular structure
But, what is the hidden magic trick that antifreeze proteins have to prevent the formation of ice in the place where ice dominates the environment? It definitely must be something powerful, and it truly is! The researchers determined that each extracted sequence contains an amino acid: threonine. Threonine is one of the 20 essential amino acids, and it can be found in every single organism on planet Earth (so yes, microbes also have it!). This amino acid has an -OH hydroxyl group, making it a polar substance. When a compound is polar, it means that it attracts water and is easily dissolvable. So, the presence of threonine in the antifreeze protein brings the “ice-threonine-antifreeze protein” interaction altogether. More specifically, the hydroxyl group protrudes from the center of the antifreeze protein in a way that makes it accessible for the interaction with an ice crystal7. Figure 3 shows the arrangement of these threonine residues in yellow (“T” stands for threonine, an abbreviation used in biochemistry).
Studying antifreeze proteins can revolutionize many technological processes used nowadays. They possess potential applications in cryopreservation8, the process of conservation of cells, tissues, or any other biological structures by cooling the samples to very low temperatures9. Because stem cells and other viable tissues cannot be stored under cooling temperatures, cryopreservation can significantly extend the longevity of these samples.
And let’s not forget about the food industry and frozen food preparation. For a long time, antifreeze proteins have been used to preserve the structure of vegetable and animal foods. By preventing the accumulation of big ice crystals during freeze-thaw cycles, antifreeze proteins maintain the crispiness and freshness of frozen food. More importantly, the more studies are conducted on antifreeze proteins, the better the understanding scientists will get on how psychrophilic bacteria survive and adapt to such a harsh environment like Antarctica.
4. Hoshino, T., Kiriaki, M. , Ohgiya, S. , Fujiwara, M. , Kondo, H. , Nishimiya, Y. , Yumoto, I. , Tsuda, S. 2003. Antifreeze proteins from snow mold fungi. Can J Bot. (81), 1175–81. 5. Muñoz, P.A. et al. 2017. Structure and application of antifreeze proteins from Antarctic bacteria. doi: 10.1186/s12934-017-0737-2.
5. Garnham, C. , Gilbert, J. , Hartman, C. , Campbell, R. , Laybourn‐Parry, J ., Davies, P. 2008. A Ca2+‐dependent bacterial antifreeze protein domain has a novel beta‐helical ice‐binding fold. Biochem J. 411(171):180.
8. Jang, T.H. , Park, S.C. , Yang, J.H. , et al. 2017. Cryopreservation and its clinical applications. Integr Med Res. 6(1):12-18. doi:10.1016/j.imr.2016.12.001