Návrh a příprava biosenzoru kinázy Src

• Main Author: Pavlík, Vojtěch
• Other Authors: Rösel, Daniel
• Format: Dissertation
• Language: English
• Published: 2011
• Online Access: http://www.nusl.cz/ntk/nusl-313045

Src kinase is a well-known proto-oncogene that contributes to cell migration and proliferation and is often found deregulated in tumors. Yet, biology of Src is not fully understood and great effort is made to find new tools for broadening knowledge about the kinase. In this thesis is described design and construction of a novel Src biosensor that exploits genetically encodable fluorophores derived from green fluorescent protein and Förster/fluorescence resonance energy transfer (FRET). The fluorophores are inserted directly into the structure of full-length c-Src in the way that should not impair the inner regulatory mechanisms of Src. The created biosensor proved to be sensitive to various stimuli, which also activate c-Src, by increase of activating autophosphorylation on Tyr4162 and decrease in FRET. Preliminary experiments indicate that the Src biosensor can be used to reflect Src activation in fixed cell as well.