The purpose of this research is to determine the structure of the thermal stable fluorescent proteins TGP-E and YTP-E. The starting point of these proteins came from the thermostable green fluorescent protein (TGP) which was developed at Los Alamos National Lab and a yellow thermostable protein (YTP).  YTP was developed by mutating a histidine located under the chromophore into a tyrosine (H197Y). The properties in which we are targeting improvement are thermal stability, pH stability, and overall fluorescence. To accomplish this overall improvement, we altered the chromophore using sited directed mutagenesis to mutate glutamine 66 to glutamate protein in both YTP and TGP proteins. We named the mutants YTP-E and TGP-E. The YTP-E protein improved the thermal stability and the overall pH stability compared to TGP-E. We have obtained the crystal structure of TGP-E that has a resolution range of 20.42 to 2 Å to explain why the Q66E mutation improved quantum yield. We are optimizing YTP-E crystals to also obtain a crystal structure of this protein. We can then use the structure to guide improvement of the YTP-E quantum yield. The mutation in YTP-E improved the rate at which the protein crystals grew, and the quality of the crystals compared to YTP.

Characterization and protein structures of thermal stable fluorescent proteins TGP-E and YTP-E

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