Fluorescence Resonance Energy Transfer

Interactions between one protein and another may be monitored by fluorescence  resonance energy transfer (FRET) by labeling the 2 proteins of interest with various fluorochromes.  The emission spectrum of 1 fluorochrome overlaps along the excitation spectrum of the other like, while the two proteins are in close proximity, light may be transferred from one fluorochrome to the other.

By Fluorescence resonance energy transfer (FRET) Interactions between one protein and another may be monitored. with a different  Fuorochrome  (tagged  with different  variants  of GFP, see above), the two proteins of interest are each labeled  selected so that the emission spectrum  of one ?uorochrome  overlaps with  the excitation  spectrum  of the other.  If the two proteins come into very near proximity (closer than 2 nm), the energy of the absorbed light may be directly transferred from one ?uorochrome  to the other.  therefore,  at  the  excitation   wavelength   of  the  ?rst  ?uorochrome when  the sample  is illuminated,  at the  emission  wavelength  of the  second light  is emitted  .  If the two proteins fail to come into near proximity then no transfer of ?uorescence occurs.

Fluorescence resonance energy transfer (FRET). the proteins are ?rst tagged with two different variants of GFP to determine whether two proteins interact inside the cell,. (a) In this instance, protein X is coupled to cyan ?uorescent protein (CFP), that is excited at 440 nm and emits blue light at 490 nm, while protein Y is coupled to yellow ?uorescent protein (YFP), that is excited at 490 nm and emits yellow light at 527 nm. (b) If protein X and Y do not interact, only illuminating the sample at 440 nm yields ?uorescence at 490 nm from CFP. (c) When protein X and Y interact, FRET now takes place. Illuminating the sample at 440 nm excites CFP, in turn excites YFP whose emission, resulting in the emission of yellow light at 527 nm.