A new probe to measure molecular crowding in vivo

COUTO PM Y CARAMELO J

Buenos Aires

Workshop; New trends in advanced fluorescence microscopy techniques. Third South American Workshop; 2011

Fluorescence Foundation

Cellular interior is a rich matrix in which macromolecular concentration can reach up to 30% w/v. Macromolecular crowding can deeply affect several biophysical aspects of the cell physiology. For instance, macromolecular crowding can alter binding constant up to 3 orders of magnitude, depending on the volume change upon binding. By a serendipitous observation, we recently found a GFP-based protein whose emission properties were markedly influenced by crowding levels. This sensor is conformed by YFP and CFP endowed with an intervening natively unfolded protein. The FRET efficiency of this construction showed a marked and cooperative increment when incubated with 20% w/v PEG, a condition similar to that found in the cytoplasm of eukaryotic cells. The possibility of tailoring the response range of this protein and targeting it to the desired cellular location, will allow to study the effect of crowing on several physiological processes.