Directed evolution of GFP with non-natural amino acids identifies residues for augmenting and photoswitching fluorescence.

ABSTRACT: Genetic code reprogramming allows proteins to sample new chemistry through the defined and targeted introduction of non-natural amino acids (nAAs). Many useful nAAs are derivatives of the natural aromatic amino acid tyrosine, with the para OH group replaced with useful but often bulkier substituents. Extending residue sampling by directed evolution identified positions in Green Fluorescent Protein tolerant to aromatic nAAs, including identification of novel sites that modulate fluorescence. Replacement of the buried L44 residue by photosensitive p-azidophenylalanine (azF) conferred environmentally sensitive photoswitching. In silico modelling of the L44azF dark state provided an insight into the mechanism of action through modulation of the hydrogen bonding network sur

SUBMITTER: Reddington SC 

PROVIDER: S-EPMC5811120 | biostudies-literature | 2015 Feb

REPOSITORIES: biostudies-literature