Fluorescence from Multiple Chromophore Hydrogen-Bonding States in the Far-Red Protein TagRFP675

Far-red fluorescent proteins are critical for in vivo imaging applications, but the relative importance of structure versus dynamics in generating large Stokes-shifted emission is unclear. The unusually red-shifted emission of TagRFP675, a derivative of mKate, has been attributed to the multiple hydrogen bonds with the chromophore N-acylimine carbonyl. We characterized TagRFP675 and point mutants designed to perturb these hydrogen bonds with spectrally resolved transient grating and time-resolved fluorescence (TRF) spectroscopies supported by molecular dynamics simulations. TRF results for TagRFP675 and the mKate/M41Qvariant show picosecond time scale red-shifts followed by nanosecond time blue-shifts. Global analysis of the TRF spectra reveals spectrally distinct emitting states that do not interconvert during the S-1 lifetime. These dynamics originate from photoexcitation of a mixed ground-state population of acylimine hydrogen bond conformers. Strategically tuning the chromophore environment in TagRFP675 might stabilize the most red-shifted conformation and result in a variant with a larger Stokes shift.1122sciescopu

Is local journalism failing? Local voices in the aftermath of the Grenfell and Lakanal fire disasters

When the Grenfell fire tragedy struck in 2017, the world watched in horror. Social media buzzed with comments, questions and demands for meaningful intervention. The local press was also blamed for failing to identify a disaster ‘foretold’. This chapter compares local press treatment of the previous major fire in London at Lakanal House in 2009 with its response to the Grenfell fire. After Lakanal House voices were amplified by local media creating a running story and investigating the causes and consequences of the fire. None of this translated into an adequate public policy response to fire safety. It asks if an emerging fifth estate can fill a news deficit and broaden public discourse to effect change? And whether local voices are, or will be, heard any more clearly to find remedies to disaster? Significantly, it considers whether local journalism can really matter if public authorities ignore the evidence it puts before the public