Abstract
Sensor proteins based on FRET between two fluorescent proteins allow real-time imaging of mol. events in living cells. Here we report new FRET-based sensor approaches for the detection of Zn(II) and protease activity. Zn(II) sensors with a very high and tunable affinity (Kd = 30 fM -1.4 pM) have been created by connecting two fluorescently labeled metal binding domains using a series of flexible peptide linkers. Modeling the conformational distribution of the linker allows a quant. understanding of the ratiometric changes and the Zn(II) affinity. In a different sensor approach, de novo Zn(II) binding sites are introduced directly on the surface of both fluorescent proteins. This sensor displays an impressive 9-fold increase in emission ratio and allows detection of Zn(II) from 10 nM to 1 mM. Finally, we present a new concept for FRET-based protease sensors that is based on intramol. complex formation between donor and acceptor fluorescent domains. [on SciFinder (R)]
Original language | English |
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Title of host publication | Proceedings of the 234th American Chemical Society National Meeting, Boston, MA, United States, August 19-23 |
Pages | BIOT-347 |
Publication status | Published - 2007 |