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Veronika Bilanovicova, Tomasz Nodzynski and his team described the structure of the hydrophilic loop (HL), a central part of PIN1 protein, that serves as a transporter of auxin phytohormone. The research team showed that the hydrophilic loop is an intrinsically disordered protein (IDP). The findings were recently published in the International Journal of Molecular Sciences and will most probably influence the future direction of biochemical research concerning PIN auxin transport. Auxins play a crucial role in the coordination of many growth processes in the plant life cycle and are essential for plant body development.
Proteins in plants are built from basic structural units such as helices and beta-pleated sheets. In the case of PIN1 hydrophilic loop there was no clarity regarding its structure. Some theories claimed that it has a secondary structure and some claimed that it is unstructured. Tomasz Nodzynski and his colleagues have shown that it is a little bit of both. The research confirmed that HL falls into the category of intrinsically disordered proteins (IDP), which is a specific group of proteins that lacks a fixed or ordered three-dimensional structure.
“Anybody who boiled an egg long enough can attest, that its insides become irreversibly hard and cannot return to fluidity once the egg is removed from hot water, as the structure of proteins in the egg yolk and white lost the normal, native structure irreversibly due to heat. Interestingly, for the hydrophilic loop in higher temperatures, which should cause protein structure degradation, we saw more structures appearing. This was puzzling. The first author of the study, PhD student Veronika Bilanovicova made sense of those results concluding that the hydrophilic loop is an intrinsically disordered protein (IDP) domain, and its structural elements are not fixed as it is typical for the globular soluble protein, that behaves like an egg yolk. The HL changes structure dynamically. To explain it very simply, try to imagine the egg becoming hard inside the boiling water but then returning to its liquid state once placed at room temperature. This realisation stimulated a whole new way of thinking about HL, which is the central domain of PIN1, explains Tomasz Nodzynski, the corresponding author of this study.
“Acknowledging the intrinsically disordered nature of PIN1 hydrophilic loop drove some of our other experiments. You see, since we know that IDP can switch their consistency back and forth, they can interact and work together with many more proteins. They can adapt and fill into the spaces and be part of solidifying interactions. You can imagine it like liquid cement and its ability to fill the spaces between the steel and pre-manufactured elements of concrete constructions. When we saw the thermal stability results indicating the IDP nature, we realised that the PIN1 hydrophilic loop might be a hub for protein interactions. And indeed, we have shown that it forms homo-dimerises, this means that it pairs with itself. And this feature is likely important for its localisation on the cell membrane enabling the whole PIN1 protein to function as directional auxin transporter. One must now consider that HL as an IDP is most probably interacting with many more proteins than previously anticipated. But not all was uncovered yet, and we suspect that our discovery is just the tip of the iceberg as more of its interactors now have to be verified. We have some hints there, but I would not like to divulge them at this stage. The IDP field is still rapidly developing, and our contribution will be certainly further stimulating. It will also surely influence the biochemical research concerning PIN auxin transporters,” adds Tomasz Nodzynski.
Many techniques and multidisciplinary expertise were used to come to this conclusion. The core work was performed by Veronika Bilanovicova, supported by Nikola Rydza, Lilla Koczka and Martin Hess, who were helping a lot with cell biology, microscopy and yeast assays. The team received external support from Elena Feraru from BOKU Austria that provided some key tools for the research. We also benefited unique expertise of Jiří Friml from IST Austria. Another important factor behind this research was the expertise and equipment provided by CEITEC Core Facilities such as Biomolecular Interactions and Crystallization Core facility, CELLIM and Plant Sciences Core Facility.