Rational design of ultrastable and reversibly photoswitchable fluorescent proteins for super-resolution imaging of the bacterial periplasm

• DOI: 10.1038/srep18459
• Publication Date: 2016-01-06T09:52:50Z
• Authors: Mariam El Khatib; Alexandre Martins; Dominique Bourgeois; Jacques-Philippe ...; Virgile Adam
• Keywords: Rational design; Chromophore; Fluorescent protein; Photoswitch

Abstract Phototransformable fluorescent proteins are central to several nanoscopy approaches. As yet however, there is no available variant allowing super-resolution imaging in cell compartments that maintain oxidative conditions. Here, we report the rational design of two reversibly switchable able fold and photoswitch bacterial periplasm, rsFolder rsFolder2. was designed by hybridisation Superfolder-GFP with rsEGFP2 inherited fast folding properties former together rapid switching latter, but at cost a reduced contrast. Structural characterisation mechanisms revealed different scenarios for chromophore cis - trans isomerisation allowed designing rsFolder2, exhibits improved contrast amenable RESOLFT nanoscopy. The rsFolders can be efficiently expressed E. coli opening door nanoscale investigation localised hitherto non-observable cellular compartments.