A Fluorogenic Red Fluorescent Protein Heterodimer

作者：Alford Spencer C; Abdelfattah Ahmed S; Ding Yidan; Campbell Robert E^*

来源：Chemistry & Biology, 2012, 19(3): 353-360.

DOI：10.1016/j.chembiol.2012.01.006

The expanding repertoire of genetically encoded biosensors constructed from variants of Aequorea victoria green fluorescent protein (GFP) enable the imaging of a variety of intracellular biochemical processes. To facilitate the imaging of multiple biosensors in a single cell, we undertook the development of a dimerization-dependent red fluorescent protein (ddRFP) that provides an alternative strategy for biosensor construction. An extensive process of rational engineering and directed protein evolution led to the discovery of a ddRFP with a K-d of 33 mu M and a 10-fold increase in fluorescence upon heterodimer formation. We demonstrate that the dimerization-dependent fluorescence of ddRFP can be used for detection of a protein-protein interaction in vitro, imaging of the reversible Ca2+-dependent association of calmodulin and M13 in live cells, and imaging of caspase-3 activity during apoptosis.

• 出版日期2012-3-23