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Projekt Druckansicht

Chemische Werkzeuge zur Untersuchung von Phosphatasen

Antragstellerin Professorin Dr. Maja Köhn
Fachliche Zuordnung Biologische und Biomimetische Chemie
Förderung Förderung von 2010 bis 2016
Projektkennung Deutsche Forschungsgemeinschaft (DFG) - Projektnummer 165935751
 
Erstellungsjahr 2016

Zusammenfassung der Projektergebnisse

Within intracellular signaling networks, phosphatases are counter players of kinases and play crucial roles in health and disease, ranging from cancer and diabetes to bacterial infection. However, to date inhibitors that directly target phosphatases are not used in the clinic due to limited specificity and bioavailability, with only very few exceptions. Chemical modulators of phosphatase function and other tools to decipher phosphatase signaling are limited or not available for the majority of phosphatases. Such tools are, however, crucial to understand the functions and regulation of phosphatases and their contribution to signaling networks. To this end, I proposed to develop new chemical tools that will enable studying protein tyrosine phosphatases (PTPs). These tools were supposed to be based on (1) phosphatidylinositol phosphates (PIP), (2) phosphonopeptides and (3) protein engineering. (1) There are few chemical tools available to study PIP biology, including PIP phosphatases. Structure-activity-relationship studies are largely absent due to the challenges in the synthesis of PIPs. We also encountered severe challenges, but nevertheless successfully designed and established the first solid phase chemistry strategy for PIP analogues, enabling the parallel synthesis of these compounds without purification of intermediates. We published first in vitro studies regarding the influence of different modifications at the position of the lipid in the PIPs on the activity of PIP phosphatases to dephosphorylate them. Library synthesis of PIP analogues modified at the head group or the lipid tail is ongoing. (2) We devised a strategy for the chemical tool design for PTPs. These tools can act as in-cell active inhibitors, pull-down baits and fluorescence in-cell detection probes. They are based on phosphopeptide substrates of PTPs chemically altered to achieve cell penetration and cellular stability. We published the proof-of-principle using the prototypic PTP1B. The tools are easily accessible to biologists through standard peptide synthesis companies. (3) It is highly difficult to find substrates and interacting proteins of small Dual Specificity Phosphatases (DSPs). Different family members of the DSPs can dephosphorylate substrates ranging from protein to non-protein substrates and their substrates are therefore hardly predictable. The interaction between the partners is transient, and these DSPs carry no specific interaction domain that would allow a prediction of protein-protein interactions. We addressed this challenge by applying protein engineering. We created photocrosslinkable interactor-trapping mutants of DSPs by introducing a photocrosslinkable amino acid at specific positions in the protein sequence. We showed that this approach enables the detection of transient interactions between proteins and small DSPs; in particular we studied the phosphatase VHR. New chemical tools will help to achieve a better understanding of phosphatase biology. In this project we established design strategies for such tools and applied them to specific questions, delivering enabling tools for PTP research. Both, chemical lead compounds and biological understanding are required to enable targeting PTPs in diseases in the future.

Projektbezogene Publikationen (Auswahl)

 
 

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