Expanding the functionality of proteins with genetically encoded dibenzo[,][1,4,5]thiadiazepine: a photo-transducer for photo-click decoration

• Published in: Chemical science (Cambridge) 2022-03, Vol.13 (12), p.3571-3581
• Main Authors: Xiong, Qin, Zheng, Tingting, Shen, Xin, Li, Baolin, Fu, Jielin, Zhao, Xiaohu, Wang, Chunxia, Yu, Zhipeng
• Format: Article
• Language: English
• Subjects: Amino acids; Chemical reactions; Chemistry; Converters; E coli; Genetic code; Isomerization; Proteins; Residues; Selectivity
• ISSN: 2041-6520; 2041-6539
• DOI: 10.1039/d1sc05710c

Genetic incorporation of novel noncanonical amino acids (ncAAs) that are specialized for the photo-click reaction allows the precisely orthogonal and site-specific functionalization of proteins in living cells under photo-control. However, the development of a r&cmb.b.line;ing-strain i&cmb.b.line;n situ l&cmb.b.line;oadable d&cmb.b.line;ipolarophile (RILD) as a genetically encodable reporter for photo-click bioconjugation with spatiotemporal controllability is quite rare. Herein, we report the design and synthesis of a photo-switchable d&cmb.b.line;ib&cmb.b.line;enzo[ b , f ][1,4,5]t&cmb.b.line;hiad&cmb.b.line;iazepine-based a&cmb.b.line;lanine (DBTDA) ncAA, together with the directed evolution of a pyrrolysyl-tRNA synthetase/tRNA CUA pair (PylRS/tRNA CUA ), to encode the DBTDA into recombinant proteins as a RILD in living E. coli cells. The fast-responsive photo-isomerization of the DBTDA residue can be utilized as a converter of photon energy into ring-strain energy to oscillate the conformational changes of the parent proteins. Due to the photo-activation of RILD, the photo-switching of the DBTDA residue on sfGFP and OmpC is capable of promoting the photo-click ligation with diarylsydnone (DASyd) derived probes with high efficiency and selectivity. We demonstrate that the genetic code expansion (GCE) with DBTDA benefits the studies on the distribution of decorated OmpC-DBTD on specific E. coli cells under a spatiotemporal resolved photo-stimulation. The GCE for encoding DBTDA enables further functional diversity of artificial proteins in living systems. Via directed evolution of the tRNA synthetase, genetic encoding of a unique DBTD derived ncAA into proteins is realized. The DBTD residue is capable of transducing photon energy into ring-strain energy in situ for photo-clicking with diarylsydnone.