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Padron was derived from rsFastLime with the following mutations: T59M/V60A/N94I/P141L/G155S/M159Y/F190S
To generate RSFPs with different switching properties, we used rsFastLime (Dronpa-V157G) as a template because of its favorable switching behavior, and its large dynamic range (67:1) in the fluorescence signal between the on- and the off-state.
At 37 °C, Padron* behaves as a true monomer, although it has a slight tendency for dimerization at lower temperatures (∼15% dimer at 4 °C). The dimerization tendency was removed by exchanging hydrophobic amino acid residues that are located in potential dimer interfaces, namely Ile94, Ile100 and Leu141...The variant obtained [ Padron*(star) ] exhibits a slightly reduced dynamic range in the switchable signal (off-state fluorescence is 2% instead of 0.7% of the on-state signal), and its bleaching per cycle amounts to 3% instead of 2%. Hence Padron*(star) is particularly useful for applications at low temperatures; for most in vivo applications that require elevated temperatures, Padron* is superior.