Dimerization-Dependent Green and Yellow Fluorescent Proteins

作者：Alford Spencer C; Ding Yidan; Simmen Thomas; Campbell Robert E^*

来源：ACS Synthetic Biology, 2012, 1(12): 569-575.

DOI：10.1021/sb300050j

• dimerization-dependent fluorescent protein
• protein engineering
• directed evolution
• biosensor
• caspase-3
• mitochondria-associated membrane

Dimerization-dependent fluorescent proteins (ddFP) are a recently introduced class of genetically encoded reporters that can be used for the detection of protein interactions in live cells. The progenitor of this class of tools was a red fluorescent ddFP (ddRFP) derived from a homodimeric variant of Discosoma red fluorescent protein. Here, we describe the engineering and application of an expanded palette of ddFPs, which includes green (ddGFP) and yellow (ddYFP) variants. These optimized variants offer several advantages relative to ddRFP including increased in vitro contrast and brightness for ddGFP and increased brightness and a lowered plc for ddYFP. We demonstrate that both variants are useful as biosensors for protease activity in live cells. Using the ddGFP tool, we generated a highly effective indicator of endomembrane proximity that can be used to image the mitochondria-associated membrane (MAM) interface of endoplasmic reticulum (ER) and mitochondria.

• 出版日期2012-12